Hormonal protocol used for cervical dilation in ewes does not affect morphological embryo quality but reduces recovery rate and temporarily alters gene expression.

BACKGROUND: Information on the impact of hormonal protocols for cervical dilation on the quality of ovine embryos is scarce. METHODS: To compare the quality of embryos after cervical dilation protocol, ewes (n = 64) were allocated into either a treated group (100 µg estradiol benzoate intravenous and 0.12 mg cloprostenol intramuscularly, 12 hours before embryo collection plus 100 iu oxytocin intravenous 15 minutes before the collection procedure) or a control group (saline). Luteal function was analysed using ultrasonography and P4 measurement. Some collected embryos were frozen/thawed for gene expression, others were cultured in vitro, frozen/thawed for gene expression, and the remaining embryos were fixed for the apoptosis test (TUNEL test). RESULTS: The treatment reduced fluid (p=0.04) and structure (p=0.03) recovery rates, but the morphological quality, development stage, and apoptosis incidence of the embryos were not affected by treatment. The corpora lutea of the control group had greater blood perfusion (p = 0.002) and greater P4 concentrations at 6, 9, and 12 h after the treatment (p < 0.0001). The expression of BAX, BCL2, PRDX1, and HSP90 genes were not affected by the treatment. However, the embryos in the treated group had fewer NANOG and OCT4 transcripts than control embryos (p = 0.008; p = 0.006, respectively). After culture, there was no difference between the groups in any gene. CONCLUSION: The hormonal protocol for cervical dilation reduced the efficiency of embryo collection. In addition, the treatment induced luteolysis and a transient alteration of embryo gene expression, however there were no detectable changes in embryo morphological quality, development stage, or incidence of apoptosis.

Hormonal protocols for early resynchronization of ovulation in ewes: The use of progestagens, eCG, and inclusion of early pregnancy diagnosis with color Doppler ultrasound.

This study aimed to evaluate 1) the effect of inserting an intravaginal sponge containing medroxyprogesterone during the late luteal phase on the corpus luteum (CL) function and endogenous production of P4; 2) the effectiveness of two different equine chorionic gonadotrophin (eCG) doses on synchronization of ovulations for a resynchronization treatment; and 3) the inclusion of an early pregnancy diagnosis in an early resynchronization protocol for ovulation in ewes. For all studies, the synchronization protocol was based on a short-term protocol of six days of progestogen treatment plus one dose of prostaglandin F2alpha, one dose of eCG, and gonadorelin acetate after sponge withdrawal. For the first study, the ewes were mated with fertile rams; a second sponge was inserted in half of these ewes 12 days later, and blood samples were collected daily for six days, until sponge withdrawal. For the second study, the ewes were not mated, and received a second sponge during the same period, after which they were divided into three groups according to eCG dose (0, 200, or 300 IU). In the third study, all ewes were artificially inseminated and received the second sponge during the same period. At sponge withdrawal, pregnancy was diagnosed by color Doppler ultrasonography (DUS) of the CL, and only non-pregnant ewes were re-inseminated two days later. In the first study, serum progesterone values were similar regardless of whether an intravaginal sponge had been inserted. In the second study, the ovulation time was more concentrated in those ewes which received 200 IU of eCG. In the third study, there was no difference between the experimental groups (with or without a previous pregnancy diagnosis) in pregnancy rate at the first insemination, accumulated pregnancy rate, and pregnancy loss. The insertion of an intravaginal sponge impregnated with medroxyprogesterone acetate did not affect the endogenous production of P4. The application of 200 IU of eCG provided the best result with regard to the synchronization of ovulations in the resynchronization treatment. Also, the inclusion of an early pregnancy diagnosis with DUS is useful and improves the general results of resynchronization programs, shortening the total working period.

Accuracy of assessment of luteal morphology and luteal blood flow for prediction of early pregnancy in goats.

This study aims to determine the best post-breeding time for an early pregnancy diagnosis in dairy goats, when using luteal morphology and vascularization assessment by B-Mode and color Doppler ultrasonography (US) in association or not with the anechoic uterine content. A total of 131 Saanen goats (2.0 ±â€¯0.5 years old) were used in the two studies. In the first study, pregnancy prediction was daily performed from Day 15-23 post-breeding in 51 does. This was based on a subjective assessment of luteal morphology (B-Mode US), luteal blood flow (color Doppler US), and the presence of anechoic uterine content (B-Mode US). In the second study, pregnancy predictions were performed in 71 does on the best post-breeding day, as determined in the first study (Day 21 using luteal blood-flow assessment and Day 23 using luteal morphological assessment and overall analysis). In both studies, pregnancy diagnosis was confirmed on Day 30 (gold standard method by B-Mode transrectal ultrasonography). The B-Mode and color Doppler US performance in reaching an early pregnancy diagnosis was evaluated by calculating sensitivity (Sens), specificity (Spec), negative and positive predictive values (NPV and PPV), accuracy (Acc), the Kappa index (κ), and the Younden index (J). In Study 1, the pregnancy prediction accuracy progressively increased from Day 17-23 using B-Mode US (D17: 50.98%; D18: 52.94%; D19: 62.75%; D20: 74.51%; D21: 86.27%; D22: 90.20%; D23: 96.08%), and from Day 17-21 using color Doppler US (D17: 49.02%; D18: 54.90%; D19: 70.59%; D20: 86.27%; D21: 96.08%). In the second study, color Doppler, B-Mode, and overall assessments at Day 21 and Day 23, respectively, presented a similar pattern of sensitivity, specificity, and accuracy as was found in the first study. Therefore, both color Doppler and B-Mode assessments can be judged as effective tools for reaching a pregnancy diagnosis in goats as early as on Day 21 and Day 23, respectively, post-breeding.