FIG. 3.

15dPGJ₂ increased PKCζ activation through a PI3-kinase-dependent process. RAW 264.7 cells were transfected for 6 h with the p110CAAX plasmid and maintained in culture for 14 h. (A) The phosphorylation at T410 of PKCζ was determined after 30 min of incubation with 2 μM 15dPGJ₂, 200 nM wortmannin, and 20 μM LY294002. The activity of PI3-kinase (in arbitrary units [a.u.]) was measured by the synthesis of PIP in immunocomplexes from cells treated for the indicated times with 15dPGJ₂ and LPS. (B) Immunoprecipitated (IP) PI3-kinase from LPS-treated cells was incubated for 10 min with 2 μM 15dPGJ₂ to evaluate the direct effect of this PG on enzyme activity. WB, Western blot; α-p85α, anti-p85α Ab. Results are from one representative experiment (of three experiments) for panel A or are the means ± standard deviations for three experiments for panel B. Values that are significantly different from the value for the corresponding condition in the absence of 15dPGJ₂ are indicated (P < 0.05 [*] and P < 0.01 [**]).