EP-4 deficiency enhances BM-DC RALDH expression. (a and b) BM cells from EP-KO or WT C57BL/6 and 129 mice were cultured with GM-CSF alone or in the presence of skin SN. After 3 d, LPS was added and, 18 h later, cells were stained with CD11c and ALD to measure DC RALDH activity. (a) Bar graph shows the mean percentage of CD11c⁺ DCs that are ALD⁺ for each mouse genotype with SEM pooled from four experiments (***, P < 0.001). (b) A representative contour plot of CD11c and ALD expression of EP-4 KO and WT 129 BM-DCs is shown. Inset values indicate the MFI of ALD expression by CD11c⁺ DCs. (c) BM cells were cultured with GM-CSF in the presence of exogenous PGE2 or EP-specific agonists (cultures comprised 25% SN from indomethacin-treated skin stroma to enhance PGE2 responsiveness). Indomethacin was added at the beginning of culture to block endogenous PGE2 synthesis. After 3 d, LPS was added and, 18 h later, cells were stained with CD11c and ALD to measure DC RALDH activity. Graph shows the mean ± SEM of the percentage of CD11c⁺ DCs that are ALD⁺ pooled from three experiments.