EP2 and EP4 expression of prostaglandin receptors in MoDCs.

(A-B) Total RNA was isolated from immature and mature MoDCs that had been stimulated with a cocktail of proinflammatory cytokines (TNF-α, IL-1β, and IL-6), sCD40L, or polyI:C in the presence or absence of PGE2 for 48 hours. The expression of EP2 (A) and EP4 (B) mRNA was analyzed by real time RT-PCR. After standardization to yield the same amount of GAPDH mRNA, all samples were compared with immature MoDCs (value = 1). The data shown represent the mean EP2 or EP4 mRNA level ± SEM obtained from 3 independent experiments with MoDC preparations from different donors. (C) MoDCs were stimulated for 48 hours with polyI:C alone (−PGE2), with polyI:C in combination with PGE2; or with polyI:C along with forskolin, cholera toxin, 11-deoxy-PGE1, or pertussis toxin in combination with PGE2; the MoDCs were subsequently analyzed in a transwell migration assay for their chemotactic response to CCL21. (D) MoDCs stimulated for 48 hours with TNF-α, IL-1β, IL-6, and PGE2 alone (thin line), or along with pertussis toxin (bold line), were analyzed for CCR7 surface expression by flow cytometry. The filled gray histogram represents staining with an isotype-matched irrelevant mAb. The data shown in panels C and D are from 1 out of 3 experiments that gave similar results.