To verify whether 15d-PGJ2 could modulate the NaBut-induced P-gp expression observed, SW620
Z-FA-FMK were treated with a combination of NaBut and 15d-PGJ2, then, promoter activity, mRNA transcription and P-gp expression were evaluated. First, we investigated whether 15d-PGJ2 affects the NaBut-induced MDR1 activation. Caco-2 cells were transiently transfected with the MDR1-luc plasmid and transfected cells were treated with NaBut alone or in combination with increasing concentrations of 15d-PGJ2 then assayed for luciferase activity ( Fig. 3A). As we expected, NaBut induced luciferase activity of the pMDR1-luc construct. While 15d-PGJ2 alone did not modulate the promoter activity, the combination between NaBut and 15d-PGJ2 profoundly increased the luciferase activity suggesting that 15d-PGJ2 exacerbated the NaBut-induced effect on P-gp transcription and expression. We next wanted to determine whether 15d-PGJ2 could regulate MDR1 transcription. As shown in Fig. 3B, 15d-PGJ2 alone did not modulate MDR1 transcription. Interestingly, when SW620
cells were treated with increasing concentrations of 15d-PGJ2 in addition to NaBut, we observed a dose-dependent augmentation of NaBut-induced MDR1 transcription. Moreover, the expression of P-gp in cells treated with NaBut, 15d-PGJ2 and a combination of these compounds was determined by FACS analysis ( Fig. 3C). An up-regulation of the P-gp protein level was observed after exposure to NaBut and 15d-PGJ2. In contrast, the exposure to 15d-PGJ2 alone did not modulate P-gp expression at cell surface.
