Figure 6
Injection of rats with EE2 reduced the enzyme activities of testosterone steroidogenesis in the Leydig cells. After treatment for 7 days, Leydig cells were isolated from the sacrificed EE2-injected rats. (A) Representative result of RIA of the concentration of testosterone in cells (1 × 105 cells per tube) treated with 25-OH-C (10−5 M) or androstenedione (10−6 M) for 1 h. At the end of the incubation, culture media were collected for testosterone RIA (n = 5). (B) Determination of the effect of EE2 treatment on the activity of P450scc in Leydig cells through evaluating the level of pregnenolone. Cells were preincubated in the absence or presence of trilostane (10−4 M) for 1 h, and then treated with or without 25-OH-C (10−5 M) in the absence or presence of trilostane for an additional 1 h. At the end of the incubation, the pregnenolone concentration in the medium was measured (n = 4). (C) Western blot and (D) quantification of P450scc and StAR of Leydig cells (3 × 106 cells) isolated from rats injected with EE2 for 7 days. Data represent means ± SEM. *P < 0.05, **P < 0.01 as compared with the EE2-untreated group. +P < 0.05, ++P < 0.01 as compared with the vehicle corresponding group. 25-OH-C, 25-hydroxycholesterol; α-Tu, α-tubulin.
