GSK-3 inhibits macrophage differentiation of human monocytes. CD14⁺ monocytes were cultured for 5 days in the presence of GM-CSF and IL-4. The GSK-3 inhibitors LiCl (10 mM), SB415286 (10 μM), and SB216763 (10 μM) were added on day 0. After 5 days of culture, cells were either tested in phagocytosis assay or were activated with intact E coli (1:1000; “Materials and methods”) for an additional 36 hours and examined for phenotypic maturation and cytokine secretion. (A) Photomicrographs of DC culture on day 5 in the absence (top panel) or presence (bottom panel) of the GSK-3 inhibitor (SB415286). One representative of 6 cultures is shown. Similar results were observed in the presence of LiCl and SB216763. (B) Phagocytosis of day-5 cultures with and without GSK-3 inhibitors. (Left panel) Phagocytic index (number of yeast particles per cell). (Right panel) Proportion of cells containing yeast particles. Shown are the means ± SD; n = 4. *P < .05; **P < .01. (C) Flow cytometric analysis of DC surface maturation marker expression following activation with E coli for 36 hours. One representative of 4 experiments is shown. (D) Cytokine secretion of E coli–activated cells. Supernatants were harvested 36 hours following activation. Absolute cytokine levels of E coli–activated MoDCs (set 1): IL-12p70: 5.4 ± 2.5 ng/mL; IL-12p40: 22.2 ± 7.9 ng/mL; IL-6: 28.1 ± 20 ng/mL; IL-10: 2.3 ± 2.3 ng/mL; and TNF-α: 10.6 ± 9.5 ng/mL. Shown are the means ± SD; n = 7-10. *P < .05; **P < .01.