Figure 7
Involvement of Nuclear Factor-kappaB (NF-κB) in mPGES-1 and COX-2 regulation. (A) Gingival fibroblasts were treated with TNFα (20 ng/ml) with or without the NF-κB pathway inhibitor Bay 11-7082 (Bay, 2.0 μM) for 24 h. Expression of mPGES-1 and COX-2 was measured by flow cytometry using specific antibodies. (B-C) Gingival fibroblasts were treated with TNFα (20 ng/ml) with or without Bay (2.0 μM) for 1, 3, 6 and 24 h (B) or 10 minutes (C). Cells were lysed and total protein was analyzed for phosphorylated NF-κB p65 (p-NF-κB) and expressed as relative to control cells at 1 h (B) or at start of incubation (C). Data is presented as mean ± s.d. Asterisks (*) indicate a significant difference (p < 0.05) between TNFα-stimulated cells and control cells at each time point, and the hash symbols (#) indicate a significant difference (p < 0.05) between TNFα-stimulated cells and cells treated with TNFα together with Bay at each time point. (D) Gingival fibroblast were cultured with the indicated doses of Bay in the absence or presence of TNFα (20 ng/ml) for 24 h. Levels of PGE2 in the culture media were measured by EIA on a Luminex system, and data is presented as mean ± s.d. Asterisks (*) indicate a significant difference from control cells not treated with TNFα or Bay (p < 0.05). (E) Gingival fibroblast were cultured with Bay (2 μM) in the absence or presence of TNFα (20 ng/ml) for 6 h. Levels of PGE2 in the culture media were measured by EIA, and data is presented as mean ± s.d. Asterisk (*) indicates a significant difference (p < 0.05). The results are representative for all three cell lines and all analyses were performed in triplicates.