Figure 4.

Inhibition of biosynthesis of Crth2 endogenous ligands increases iNOS and Cox2 expression in RAW264.7 cells. A–C) Representative immunoblot (A) and quantitation of Cox2 (B) and iNOS (C) in NTC and Crth2^−/− RAW264.7 cells treated with DMSO or HQL79 (25 µM) after 8 h of LPS stimulation (100 ng/ml). β-actin is used as loading control. P value is in comparison with NTC RAW264.7 cells; n = 3 independent experiments. D, E) Real-time PCR of Cox2 (D) and iNOS (E) expression in NTC and Crth2^−/− RAW264.7 cells treated with DMSO or HQL79 (25 µM) after 8 h of LPS stimulation. P value is in comparison with NTC RAW264.7 cells; n = 3 independent experiments. F, G) Representative immunoblot (F) and quantitation (G) of iNOS and Cox2 in RAW264.7 cells lysates prepared after stimulation with LPS (100 ng/ml) for 1 h with or without DKPGD₂ (50 nM) after 1 h cells were washed and replenished with fresh medium containing either PBS or DKPGD₂ (50 nM) up to 8 h. β-actin is used as loading control. P value is in comparison with expression in cells incubated with PBS; n = 3–4 independent experiments. H) Mean fold change of mRNA levels of Cox2 (8 h) and iNOS (4 h) normalized to 18S RNA. P value is in comparison with PBS-treated group; n = 3–4 independent experiments. *P < 0.05, **P < 0.01, ****P < 0.0001 (Student’s t test).