Fig. 4

Metformin regulates androgen receptor (AR) signaling and a subset of AR target genes within C4–2 cells. A C4–2 cells were treated for 24 hours with phosphate buffered saline (PBS) vehicle or metformin (0.2–10 mM). Western blot analysis was performed to determine AR and actin protein levels in treated cells. B C4–2 cells were treated with PBS vehicle or 5 mM metformin for 24 hours. qRT-PCR was performed on total RNA extracted from treated cells to measure the level of full-length AR (AR-FL) and 18S RNA. Each bar represents the mean −/+ SEM for three independent samples. *, P < 0.05 compared to PBS control. C qRT-PCR was performed to detect the level of GR and 18S RNA in C4–2 and 22Rv1 cells grown in FBS-containing media and exposed to PBS vehicle or 5 mM metformin. Each bar represents the mean −/+ SEM for three independent samples. *, P < 0.05 compared to PBS control. D Venny program was used to compare the list of metformin regulated genes identified by RNA-Seq to the AR direct target genes identified by C. Massie [30]