PMA enhanced c-fos expression in NP cells.

<p>(<b>A</b>) c-fos was transfected with the pGL4.74 vector into NP cells, and the cells were stimulated with PMA (200 nM) for 6–24 h (left panel). In other experiments, c-fos was transfected with the pGL4.74 vector into NP cells, and the cells were stimulated with various concentrations of PMA (0, 10, 100, and 200 nM) for 12–24 h (right panel). (<b>B</b>) Effect of activation or inhibition of the PKC pathway on c-fos promoter activity in NP cells. *<i>P</i> < 0.05 between groups. Error bars represent SD. n.s. = not significant. (<b>C</b>) Detection of c-fos protein expression by immunofluorescence microscopy. After 1-day of serum deprivation, NP cells were incubated in serum-free medium containing PMA. NP cells were cultured with or without 200 nM PMA for 24 h, fixed, and stained with an antibody against c-fos. <b>Left</b>: cells stained with antibody to c-fos. <b>Middle</b>: cells stained with DAPI to identify healthy nuclei, (blue). <b>Right</b>: cells stained with antibody to c-fos and with DAPI. Scale bar= 50 μm (original magnification 20×). (<b>D</b>) Detection of c-fos (left) and c-jun (right) protein by western blot analysis after treatment with PMA (0–200 nM) for 24 h. Densitometry analyses were performed to quantify the levels of c-fos and c-jun protein 24 h after PMA treatment, after normalization to the level of β-actin. *<i>P</i> < 0.05 indicates significant differences between groups.