The aim of this study was to determine the impact of the length of the preovulatory period and consequent influences on preovulatory plasma estradiol concentrations, on expression of mRNA for ISG-15, SPP1 and Mx1 by endometrium collected from beef cows slaughtered 17.5 d after GnRH-induced ovulation. Twenty-one non-lactating cows in 2 replicates (Rep 1, n = 10; Rep 2, n = 11) were pre-synchronized to a common day of estrus and ultrasonographic-guided follicle aspiration was performed 6.2 ± 0.4 d after estrus (d -7 of the experiment). Cows were assigned to receive prostaglandin F2α on either d -3 (high estradiol group, n = 9; HiE), or d -2 (low estradiol group, n = 12; LoE). All cows received GnRH (100 µg) on d 0 (h 0), resulting in a preovulatory period of either 3 (HiE) or 2 (LoE) days. On d 7, cows received no further treatment (Cyclic = C) or were implanted with an embryo (Pregnant = P) resulting in 4 treatments (HiE-P, n = 5; HiE-C, n = 4; LoE-P, n = 8; LoE-C, n = 4). Ovarian ultrasonography was performed on d -7, -3, 0, 2 and 6 to monitor emergence and maximum diameter of the preovulatory follicle and formation of a new corpus luteum after GnRH. Blood samples collected at 12 h intervals from h -72 to h -12 and at h -6 and 0 were used to determine preovulatory concentrations of estradiol. Progesterone concentrations were assessed in samples collected on d -3, -2, 0, 2, 4, 6, 9, 11, 13 and daily to d 17.5. All cows were slaughtered on d 17.5 and endometrium was collected and snap frozen for later analyses. Number of animals in the HiE-P and LoE-P treatments reflects those from which embryos were recovered and for the HiE-C and LoE-C treatments, those cows with luteal phase progesterone concentrations at slaughter. Gene expression was determined using RT-qPCR for ISG-15, Mx1, SPP1 and RPL19. Statistical analyses were conducted using mixed procedures of SAS with appropriate models that included treatment, pregnant/cyclic, Rep and interactions for gene expression, with the addition of d as a repeated measure for hormone concentrations. Estradiol concentrations differed between the HiE and LoE treatments (treatment x h, P <.05), being greater (P<.05) in the HiE than LoE treatment at h 48 (3.2 ± 0.32 and 1.6 ± 0.26 pg/ml, respectively). Concentrations of progesterone from d 0 to d 17 did not differ between the HiE and LoE treatment. Relative amount of mRNA for Mx1 was greater in P than C females, but did not differ between the HiE-P (10.3 ± 2.9) and LoE-P (9.4 ± 2.8) treatments. ISG-15 mRNA was undetectable in 5/8 cows in the C treatments and did not differ between the HiE-P and LoE-P treatments (4.8 ± 1.1 and 5.9 ± 1.5, respectively). For SPP1, amount of mRNA did not differ between the HiE-C and HiE-P treatments, but was was greater (P < 0.05) in the LoE-P (2.8 ± 0.3) than LoE-C treatment (1.3 ± 0.03). In Rep 1, relative amount of SPP1 was greater (P < 0.05) in the HiE-P than LoE-P treatment (4.2 ± 0.5 and 2.5 ± 0.4, respectively) whereas in Rep 2 the inverse tended (P = 0.06) to occur (treatment x Rep, P < 0.05). In conclusion, altered concentrations of estradiol during the preovulatory period did not influence amounts of endometrial mRNA for ISG-15 and Mx1 in pregnant cows, and had variable effects on amounts of mRNA for SPP1. (poster)