The Peroxisome Proliferator-Activated Receptor Gamma System Regulates Ultraviolet B-Induced Prostaglandin Production in Human Epidermal Keratinocytes
Figure 3
The specific antagonist GW9662 blocks UVB- and TBH-induced PGE2 production and UVB-induced COX-2 protein expression. (a) GW9662 inhibits UVB- and TBH-induced PGE2 production in PHKs. PHKs were pretreated with vehicle control (VEH), GW9662 (1 M) for 1 hour, or the COX-2 inhibitor NS398 (10 M) for 30 minutes, then treated with 10 M tert-butylhydroperoxide (TBH), 500 nM calcium ionophore A23187, or exposed to 600 J/m2 UVB irradiation (UV), respectively. PGE2 production was assayed in the tissue culture supernatants after 8 hours and was normalized to tissue weight. The values of PGE2 expression shown are mean SEM and are representative of three experiments , , by t-test compared with nonirradiated vehicle control , or from the respective UV- or TBH-treated cells without GW9662 pretreatment ). (b) GW9662 inhibits UVB-induced COX-2 expression by immunoblot. PHKs were pretreated with vehicle (VEH) or 1 M GW9662 (GW) 1 hour prior to irradiation with 300 J/m2 UVB (UV). After 20 hours, total cellular protein was isolated and immunoblots were performed for COX-2 expression (a) as detailed in the methods section. (b) shows a high MW nonspecific invariant band used to normalize for COX-2 expression. After film exposure, band intensities were assessed using NIH Image J. Normalized band intensities are shown at the bottom as a relative increase from VEH control cells.
