Figure 7.

Expression profile of AR regulated genes. Shown are expression profiles of AR-regulated genes, with physiologic DHT versus physiologic and pharmacologic MPA concentrations in Y-AR cells compared. (a) Venn diagram comparing the number of genes regulated at least 2.0-fold by low-dose DHT versus low and high dose MPA. Y-AR cells were treated with ethanol, or 10 nmol/l progesterone, MPA, or DHT, or 1 μmol/l MPA for 6 hours in triplicate, time separated experiments. RNA was extracted, derivatized, and used to probe Affymetrix U-133 2 plus gene chips interrogating about 47,000 genes. Data were analyzed using Gene Spring software, and statistical analysis was performed using a one-way analysis of variance, with P < 0.05 considered statistically significant for genes regulated at least 2.0-fold. (b) Confirmation of the hormonal regulation of four genes identified in panel a. Four genes shown to be hormone regulated in Y-AR cells (i.e. F3, Maf-B, Krt4 and p57) were chosen for further analysis. Bar graphs: the hormonal regulation of the four selected transcripts in Y-AR cells, as assessed by microarray profiling. RT-PCR: Y-AR cells were treated for 6 hours with ethanol, 10 nmol/l progesterone, MPA, or DHT, or 1 μmol/l MPA and RNA was isolated. Primers directed against the transcripts of the four selected genes were used in RT-PCR reactions, and GAPDH was run as an internal control. (c) Venn diagrams showing number and overlap of genes regulated by 10 nmol/l MPA in AR⁺ Y-AR cells versus PR⁺ T47D cells. Microarray and data analysis were performed as described for Fig. 3a (PR⁺, AR^- cells) and panel a above (AR⁺, PR^- cells), and data for low-dose MPA in the two cell lines were compared. The 88 genes regulated by MPA through either PR or AR were tabulated (Additional files 1 and 2). The top ranking genes are shown in Table 1. AR, androgen receptor; DHT, dihydrotestosterone; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; MPA, medroxyprogesterone acetate; PR, progesterone receptor.