Title: Association of circulating concentrations of estradiol during the preovulatory period and expression of steroidogenic enzymes Authors
Submitted to: International Symposium on Reproduction in Domestic Ruminants
Publication Type: Abstract Only
Publication Acceptance Date: April 16, 2014
Publication Date: N/A
Technical Abstract: Cows with greater circulating concentrations of estradiol during the preovulatory period (HighE2) have increased pregnancy success following a fixed-time AI protocol. Furthermore, these animals have an enhanced ability to produce estradiol as indicated by increased expression of CYP19A1 and LHR within granulosa cells and increased concentrations of androstenedione within the follicular fluid of the dominant follicle. This study was conducted to determine if there were changes within the steroidogenic pathway, prior to androstenedione production, associated with increased circulating concentrations of estradiol. Beef cows (Rep1; n=32, Rep2; n=20) were synchronized by an injection of GnRH (100µg; i.m.) on d -7 and an injection of PGF2a (25mg; i.m.) on d 0. In Rep1, estrus was monitored and blood samples were collected every 3 h from PGF2a on d 0 to h 33 after PGF2a and at slaughter (h 36 to 42; n=10). At slaughter, follicular fluid (FF) and granulosa cells were collected from the dominant follicle. In Rep2 the dominant follicle was aspirated via ultrasound-guided follicular aspiration (ASP) to collect FF and granulosa cells (h 38 to 46). In Rep2, estrus was monitored and blood samples were collected every 8 h from PGF2a until ASP. Circulating concentrations of estradiol and FF concentrations of progesterone were determined by RIA and cows were classified as either HighE2 (peak >/=6.0 pg/mL; n=4, and >/=7.3 pg/mL; n=6) or LowE2 (peak =4.5 pg/mL; n=5, and </=5.5 pg/mL; n=11) in Rep1 and 2, respectively. Total cellular RNA was extracted from granulosa cells and RT-PCR was performed to determine relative expression of mRNA for CYP11A1 (the gene encoding P450scc), 3beta-hydroxysteroid dehydrogenase (HSD), and GAPDH. There was a tendency for a treatment (P=0.10), and a replicate effect (P<0.01) for FF concentrations of progesterone, but there was no treatment by replicate interaction (P=0.70). Follicular fluid from slaughtered animals (Rep1) had increased progesterone (75.94±9.77 ng/mL) compared to FF collected by ASP (Rep2; 34.09±7.39 ng/mL) which was diluted in 2 mL flush media during the flush procedure. However, in both reps, HighE2 tended to have increased FF concentrations of progesterone when compared to LowE2 (65.51±9.40 ng/mL vs. 44.53±7.86 ng/mL, respectively). There was a treatment by replicate interaction (P<0.01) for expression of CYP11A1 and 3beta-HSD, therefore, data are reported separately. There was no difference (P=0.13) in expression of CYP11A1 in Rep1, but in Rep2 HighE2 had increased expression of CYP11A1 (P<0.05; 306.77±74.64) compared to LowE2 (32.69±74.64). Expression of 3beta-HSD was increased in HighE2 (P=0.01; 11.33±1.43, P<0.01; 184.84±21.49) compared to LowE2 (3.01±1.10, 16.97±21.49) in Rep1 and 2, respectively. In conclusion, beef cows with greater concentrations of circulating estradiol prior to fixed-time AI experienced an up-regulation of the steroidogenic pathway during the preovulatory period. Thus management methods to up regulate the entire steroidogenic pathway are necessary.