Figure 4.

Rhythmic clock gene expression triggered by 15d-PGJ₂ is independent of PPAR-γ-, MAPK-, JNK- and p38MAPK-signaling pathway. After 1 h pretreatment of DMSO (15d; upper left panel), 10 μM GW9662 (GW+15d; upper right panel), 10 μM U0126 (U+15d; middle left panel), 20 μM SP600125 (SP+15d; middle right panel), or 30 μM SB203580 (SB+15d; bottom left panel), NIH3T3 cells were stimulated by 10 μM 15d-PGJ₂ for 1 h. As a negative control, NIH3T3 cells were pretreated by DMSO and stimulated by DMSO for 1 h (DMSO; bottom right panel). Total RNAs were isolated at each time point. Quantitative real-time RT-PCR was performed using mPer2 (solid line), mBmal1 (dotted line), and 18S rRNA primers. Abscissa presents "hour", ordinate "mRNA amount", respectively. The maximum mRNA amount was set to 1. The relative levels of each mRNA were normalized to the corresponding 18S rRNA levels.