FIGURE 10.

Dex plus GnRH synergistically decreases cell proliferation by up-regulating SGK-1. A, Flot-1 and SGK-1 expression levels were decreased with 40 nm siRNA-mediated knockdown for 72 h, after which the cells were harvested for RNA extraction. Total RNA was isolated and reverse-transcribed, and relative levels of SGK-1 transcripts were determined by quantitative real-time PCR. Fold changes in SGK-1 mRNA levels were normalized to GAPDH transcripts and were calculated relative to vehicle-treated (Ctrl, control) samples. The graph is representative of three independent experiments. B, as in A except that the cells were reseeded after knockdown and stimulated for 48 h with 100 nm Dex, 100 nm GnRH, or a combination of both. During the last hour of stimulation, the thymidine analog, BrdU, was added to the cells, and newly synthesized DNA was quantified with a colorimetric immunoassay. One-way ANOVA with Dunnett's (A) or both Dunnett's and Tukey (B) post-tests were used for statistical analysis and denoted as *, **, or *** to indicate p < 0.05, p < 0.01, or p < 0.001, respectively.