Figure 8

Stat6 gene expression is induced by progesterone. Synchronized T47D cells were treated with or without progesterone (30nM) for the indicated times (A, left panel) or for 24 h (A, right panel, and B). A. Quantitative PCR analysis. mRNA levels are expressed relative to levels in vehicle-treated control cells harvested at 6 h, arbitrarily set as 1. Right panel. Cycloheximide (5 μg/ml) or actinomycin D (10 μg/ml) was added to the medium 2 h before the addition of progesterone or ethanol (vehicle). h, hour; d, day; NS, not significant; *, P ≤ 0.05; **, P ≤0.01 versus control. B. Modulation of progesterone on Stat6 was detected in T47D cells after 24 h progesterone treatment using Confocal Laser Scanning Microscope. Coverslips seeded with synchronized T47D cells treated with or without progesterone (30nM) and vehicle were triple-stained with Stat6 antibody and one secondary antibody conjugated with FITC to permit detection of Stat6 (a), phalloidin to permit detection of actin (b), and DAPI, a DNA-binding dye, to counterstain the DNA (c). In T47D cells, Stat6 accumulated in nuclear domains that colocalized with dense regions of DAPI staining (d).