Effect of ovulation synchronization program and season on pregnancy to timed artificial insemination in suckled beef cows.

This study was conducted to (i) evaluate the requirement for the administration of GnRH coincident with insertion of a progesterone-releasing intravaginal device (PRID) and (ii) the effect of supplementing with 400 IU eCG at PRID removal on pregnancy per AI (P/AI) in spring and autumn calving suckled beef cows, subjected to a 7-d CO-Synch + PRID timed artificial insemination (TAI) program. Suckled beef cows (n = 1408) on 62 commercial farms were enrolled and randomly assigned to either of three treatments: 1) cows received a PRID and 100 µg GnRH on Day -10, followed by 25 mg PGF2α at PRID removal (Day -3) and 100 µg GnRH 72 h later (Day 0) at TAI (Treatment 1; n: spring = 236, autumn = 248); 2) as Treatment 1, but without GnRH at PRID insertion on Day -10 (Treatment 2; n: spring = 232, autumn = 227); 3) as Treatment 1, but cows also received 400 IU eCG at PRID removal on Day -3 (Treatment 3; n: spring = 233, autumn = 232). At Day -10, ovaries were examined by ultrasonography to evaluate the presence or absence of a corpus luteum (CL) and follicle(s) ≥ 10 mm in diameter. Body condition score (BCS) was assessed on a scale of 1-5. Pregnancy diagnosis was carried out 30-35 d after TAI by transrectal ultrasonography. Data were analyzed using the GENMOD and LOGISTIC procedures of SAS. There was a treatment by season interaction for P/AI (P < 0.001). In spring, overall P/AI was 59.1% (414/701) and was affected by treatment (59.3 v 49.6 v 68.2%, for Treatments 1, 2 and 3, respectively P < 0.05). In contrast, in autumn, overall P/AI (51.5%, 364/707) was unaffected (P > 0.05) by treatment (50.1 v 53.7 v 48.7% for Treatments 1, 2 and 3, respectively). Overall, eCG had a positive effect on P/AI for cows lacking a CL at treatment initiation (P < 0.05). In addition, in cows with low BCS (≤2.25), eCG supplementation tended (P = 0.09) to improve P/AI. Seasonal differences in response to synchronization treatment may be reflective of different management regimens (grazing v confinement) and breed type and remain to be elucidated.

Effects of systemic progesterone during the early luteal phase on the availabilities of amino acids and glucose in the bovine uterine lumen.

The uterine histotroph provides essential nutrition to the developing conceptus during the preimplantation period of pregnancy. The objective of the present study was to examine the effects of cycle stage and progesterone (P4) concentrations in the blood on the recoverable quantities of amino acids and glucose in the histotroph during the preimplantaion period of conceptus development. Following oestrus, dairy heifers were assigned to low, control or high P4 groups (n=6 heifers per treatment and time point). The uterine horn ipsilateral to the corpus luteum was flushed on either Day 7 or Day 13. The present study quantified 24 amino acids and glucose in the uterine flushings using HPLC and fluorometry, respectively. Heifers in the low P4 group had lower plasma concentrations of P4 throughout the cycle, whereas heifers in the high group had higher plasma concentrations of P4 between Days 3 and 7 compared with the control group (P<0.05). Total recoverable neutral (Ser, Gln, Gly, Thr, Cit, ß-Ala, Tau, Ala, Tyr, Trp, Met, Val, Phe, Ile, Leu, Pro and Cys), acidic (Glu) and basic (His, Arg, Orn and Lys) amino acids were greater (P<0.05) on Day 13 than on Day 7. There was no significant difference in the amount of Asp or Asn between Day 7 and Day 13. The amount of amino acids recovered on Day 7 was similar across treatment groups. On Day 13, the amount of Asn, His and Thr was lower (P<0.05) in the low P4 heifers compared with the controls and/or high P4 heifers. Quantities of glucose were not altered by cycle stage or P4 treatment. In conclusion, the stage of oestrous cycle and P4 play important roles in modulating amino acids in the histotroph, a potentially critical factor for early embryonic and/or conceptus survival.

Alterations in systemic concentrations of progesterone during the early luteal phase affect RBP4 expression in the bovine uterus.

Systemic progesterone affects the timing and duration of uterine endometrial gene and protein expression and has significant effects on conceptus development. The objective of the present study was to examine how changes in progesterone concentrations during the early luteal phase affect retinol-binding protein (RBP4) mRNA and protein concentrations in the uterus. Endometrial tissue and uterine flushings were recovered on Days 7 and 13 of the oestrous cycle in heifers with high, normal and low progesterone concentrations. RBP4 mRNA and protein concentrations were higher (P<0.05) on Day 13 compared with Day 7 in heifers with high and control progesterone concentrations. However, there was no difference in RBP4 protein concentrations between Days 7 and 13 in heifers with low progesterone (P>0.05). On Day 7, although heifers with low progesterone had lower RBP4 mRNA expression compared with controls (P<0.05) there was no difference in protein concentrations between treatment groups. On Day 13, RBP4 mRNA was 2-fold higher (P<0.001) in heifers with high and control progesterone compared with their low-progesterone counterparts and RBP4 protein concentrations were over 2-fold higher (P<0.001) in heifers with high compared to low progesterone. In conclusion, progesterone modulates uterine RBP4 mRNA and protein abundance in a time- and concentration-dependent manner.

Proteomic characterization of histotroph during the preimplantation phase of the estrous cycle in cattle.

Uterine secretions, or histotroph, are a critical component for early embryo survival, functioning as the sole supply of vitamins, minerals, enzymes, and other myriad of nutrients required by the developing conceptus before implantation. Histotroph is therefore a promising source for biomarkers of uterine function and for enhancing our understanding of the environment supporting early embryo development and survival. Utilizing label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) shotgun proteomics, we characterized the uterine proteome at two key preimplantation stages of the estrous cycle in high fertility cattle. We identified 300 proteins on Day 7 and 510 proteins on Day 13 including 281 proteins shared between days. Five proteins were more abundant (P < 0.05) on Day 7 compared with Day 13 and included novel histotroph proteins cytokeratin 10 and stathmin. Twenty-nine proteins were more abundant (P < 0.05) including 13 unique on Day 13 compared with Day 7 and included previously identified legumain, metalloprotease inhibitor-2, and novel histotroph proteins chromogranin A and pyridoxal kinase. Functional analysis of the 34 differentially expressed proteins (including 14 novel to histotroph) revealed distinct biological roles putatively involved in early pregnancy, including remodelling of the uterine environment in preparation for implantation; nutrient metabolism; embryo growth, development and protection; maintenance of uterine health; and maternal immune modulation. This study is the first reported LC-MS/MS based global proteomic characterization of the uterine environment in any domesticated species before implantation and provides novel information on the temporal alterations in histotroph composition during critical stages for early embryo development and uterine function during the early establishment of pregnancy.