FIG 6.

Effect of transient exposure of Lin⁻ progenitor cells to live C. albicans cells prior to differentiation on macrophage cytokine production. (A) Schematic protocol of in vitro Lin⁻ cell differentiation and stimulation. Lin⁻ cells were cultured in the presence or absence of live C. albicans cells at 1:0.5 or 1:2 ratio (progenitor/yeast) for 6 h and then amphotericin B (0.5 μg/ml) was added. Lin⁻ cells were transferred to a new plate and cultured with M-CSF for 7 days to obtain macrophages. (B) Macrophages were stimulated with 100 ng/ml of Pam₃CSK₄, and TNF-α and IL-6 levels in 24-h culture supernatants were assessed by ELISA. (C) Lin⁻ cells from WT, TLR2^−/−, or dectin-1^−/− mice were cultured in the presence or absence of live C. albicans cells at 1:2 ratio (progenitor/yeast) for 6 h and differentiated as indicated in the schematic protocol in panel A. (C) Macrophages were stimulated, and TNF-α, IL-6, and MIP-2 production was measured as indicated for panel B. Triplicate samples were analyzed in each assay. Results are expressed as means ± SDs of pooled data from two experiments. *, P < 0.05; ***, P < 0.001 with respect to cytokine production by macrophages derived from Lin⁻ cells unexposed to C. albicans.