FIGURE 3.

LTB₄-dependent stabilization of COX-2 mRNA was mediated through MAPK signaling. Cells were treated with vehicle (Con) or with 5.7 pmol/liter (100 pg/ml) of rhIL-1β for 16 h in the presence or absence of the COX-2 inhibitor NS-398 and increasing concentrations of LTB₄ with or without U0126 (1 μmol/liter). Monolayers were extracted for RNA and protein as follows: A, 5 μg of total RNA was analyzed for COX-2 mRNA and GAPDH mRNA by Northern hybridization using specific DIG-labeled cDNA probes; B, 50 μg of protein was analyzed for COX-2 protein by Western blotting using specific polyclonal antisera as described under “Experimental Procedures.” ANOVA of means for densitometric analysis in A and B is as follows: IL-1β + NS-398 versus IL-1β + NS-398 + LTB₄ (− log M −9 to −6), p < 0.001; IL-1β versus IL-1β + NS-398 + LTB₄ (− log M −8 to −6), not significant; IL-1β + NS-398 + LTB₄ (− M −9 to −6) versus IL-1β + NS-398 + LTB₄ (− log M −9 to −6) + U0126, p < 0.002.