Figure 2.

Regulation of CAP mRNA level by PPARγ activators. (A) Actimomycin D (5 μg/ml) was added 1 h before troglitazone treatment of 3T3-L1 adipocytes for 12 h. The relative level of CAP mRNA was determined by RNase protection assay. (B) After treatment of 3T3-L1 adipocytes with troglitazone (10 μM) for 12 h, the cells were rinsed three times, and actimomycin D (5 μg/ml) then was added. Cells were incubated further in the absence (○) and presence (•) of troglitazone for the indicated times. CAP mRNA was measure by RNase protection assay. Message levels were quantitated, and the CAP/cyclophilin mRNA ratio was calculated. Results shown are representative of three separate experiments with essentially identical results. (C) 3T3-L1 adipocytes were untreated or treated with troglitazone or rosiglitazone (10 μM) for 12 h. Nuclei were isolated, and nuclear transcription assays were performed as described in Materials and Methods. (D) 3T3-L1 adipocytes were treated with cycloheximide (5 μg/ml) for 30 min before treatment with rosiglitazone (10 μM) for 12 h. Total RNA was isolated and analyzed for CAP mRNA expression by RNase protection assay. Trog, troglitazone; Rosi, rosiglitazone; CHX, cycloheximide.