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Prokineticin 1 (PROK1) is an important factor in pregnancy establishment in pigs, acting at the embryo-maternal interface and the corpus luteum (CL). Estradiol-17β (E2) is the primary pregnancy recognition signal in pigs, and its effects are augmented by luteotropic prostaglandin E2 (PGE2). On the contrary, prostaglandin F2α (PGF2α) exerts mainly a luteolytic effect. The present study aimed to elucidate whether E2, PGE2, and PGF2α regulate the expression of PROK1 and its receptors in the porcine CL and to determine the PROK1 effect on luteal endothelial cells and pathways that may be involved in this regulation. The effects of E2, PGE2, and PGF2α on the expressions of PROK1 and its receptors in the CL were studied using an in vitro model of ultrathin luteal tissue explants model. Additionally, the effects of E2 and PGE2 on the PROK1 system were determined using an in vivo approach, in which the hormones were administered into the uterine lumen to imitate their secretion by embryos. Endothelial cell proliferation was measured using the colorimetric method. E2 acting via estrogen receptors simulated the mRNA and protein expressions of PROK1 and PROKR1 in CL explants in vitro (p < 0.05). The simultaneous action of E2 with PGE2 enhanced the expression of luteal PROK1 mRNA in vitro (p < 0.05). Estradiol-17β acting alone significantly increased PROK1 mRNA levels in vivo, whereas E2 simultaneously administered with PGE2 significantly elevated the PROK1 mRNA expression and PROKR1 mRNA and protein contents in CLs adjacent to uterine horns receiving hormonal infusion compared with CLs adjacent to placebo-treated uterine horns (p < 0.01). The PROK1 protein expression was significantly higher in the CLs of pigs treated with E2, PGE2, and E2 together with PGE2 than in the control group. PGF2α increased the PROK1 mRNA content in CLs on days 12 and 14 of the estrous cycle (p < 0.05). The expression of PROKR2 at the mRNA and protein levels remained unchanged in response to in vitro and in vivo treatments. PROK1 stimulated the proliferation of luteal endothelial cells by activating the MAPK, AKT, and mTOR pathways (p < 0.05). In summary, the luteal expressions of PROK1 and PROKR1 in early pregnancy are regulated by E2 and PGE2. PROK1 stimulates luteal angiogenesis by activating the MAPK, AKT, and mTOR pathways. The regulation of luteal PROK1 expression by PGF2α indicates PROK1's putative role during luteolysis. We conclude that PROK1-PROKR1 signaling supports luteal function during CL rescue in pregnancy in pigs., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Benzo(a)pyrene (B(a)P) is a widespread persistent organic pollutant (POP) and a well-known endocrine disruptor. Exposure to BaP is known to disrupt the steroid balance and impair embryo implantation, but the mechanism under it remains unclear. The corpus luteum (CL), the primary source of progesterone during early pregnancy, plays a pivotal role in embryo implantation and pregnancy maintenance. The inappropriate luteal function may result in implantation failure and spontaneous abortions. Therefore, this study was conducted to assess the effects and potential mechanisms of B(a)P on the CL function. Our results showed that pregnant mice received B(a)P displayed impaired embryo implantation and dysfunction of ovarian CL. The estrogen and progesterone levels decreased by B(a)P. In vitro, exposure to BPDE, which is the metabolite of B(a)P, affected the luteinization of granular cell KK-1. Additionally, melatonin and its receptors, which are important for ovarian function and anti-oxidative damage, were affected by B(a)P or BPDE. B(a)P or BPDE-treated alone impaired antioxidant capacity of ovarian granulosa cells, caused an increasing of ROS and cell apoptosis, and disrupted the PI3K/AKT/GSK3β signaling pathway in vivo and in vitro. Co-treatment with melatonin alleviated B(a)P or BPDE-induced CL dysfunction by ameliorating oxidative stress, counteracting phosphorylation of PI3K/AKT/GSK3β signaling pathway, decreasing the apoptosis of the ovarian cells. Moreover, activation of the melatonin receptor by ramelteon in KK-1 cells exhibits an analogous protective effect as melatonin. In conclusion, our findings not only firstly clarify the potential mechanisms of BaP-induced CL dysfunction, but also extend the understanding about the ovarian protection of melatonin and its receptors against B(a)P exposure., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: The main goal of the present study is to investigate the effects of retinoic acid and fibroblast growth factor-2 on serum levels of FSH and LH, histology, and apoptosis in the mouse model of Poly Cystic Ovary Syndrome (PCOS)., Materials and Methods: 80 female NMRI mice have been randomly divided into eight groups. Group 1 received normal saline as a control, and Group 2 received estradiol valerate (EV) at 4 mg/100 g of body weight. Moreover, Groups 3-4 were administered with RA (a dose of 0.05 μg/μl) and FGF2 (a dose of 0.01 μg/kg), respectively. Groups 5 and 6 respectively received the EV plus the RA (0.05 μg/μl) and FGF2 (0.01 μg/kg). Group 7 received the RA and FGF2 at doses corresponding to healthy mice, and Group 8 received the EV plus the RA + FGF2 (similar to previous doses). RA and FGF2 were injected three times per week for four weeks. Finally, histological and immunohistological parameters of the ovary were evaluated., Results: The study revealed that both single and combined injection of fibroblast growth factor-2 (FGF2) and retinoic acid (RA) in groups 5, 6, and 8 significantly reduced follicular diameters compared to group 2. Measurements confirmed that simultaneous injection of RA and FGF2 into polycystic mice significantly increased antral follicles, corpus luteum (CL), epithelial thickness, and oocyte diameter as well as decreased cystic follicles. Positive TUNEL cells that were considerably increased in the antral follicle of group 2 significantly decreased in the RA and FGF2 recipient groups, either alone or in combination. Besides, the injection of FGF2 increased preantral follicles and CL., Conclusion: The findings of the present investigation reveal that injection of RA and FGF2 has both protective and ameliorative effects that can promise new therapies for women with PCOS., Competing Interests: Declaration of Competing Interest There is no conflict of interest, and all authors have read and approved the manuscript being submitted., (Copyright © 2020. Published by Elsevier B.V.)

The objective of this study was to determine the effects of bovine nerve growth factor-β (NGF) on pre-ovulatory follicle vascular area, LH release, ovulation, and luteal function when administered systemically to heifers. Post-pubertal Holstein heifers (n = 12) received an intravaginal progesterone-releasing device (CIDR) and GnRH agonist (100 μg IM). The CIDR was removed 5 d later, and heifers were given dinoprost (25 mg IM) at CIDR removal and 24 h later, followed by a second dose of GnRH agonist 48 h later. Heifers were randomly assigned to treatments using a cross-over design. For example, heifers assigned to NGF (250 μg reconstituted in 12 mL PBS IM) in replicate 1 were assigned to control (12 mL PBS IM) in replicate 2. Transrectal ultrasonography was performed before treatment and repeated every 4 h up to 32 h to determine the pre-ovulatory follicle diameter, vascular area, and ovulation. Serum samples were obtained to assess LH concentrations during the periovulatory period and every 2 d post-ovulation for measuring progesterone concentrations. A subset of heifers had luteal biopsies performed on days 9 (n = 6 per treatment) and 14 (n = 6 per treatment) post-ovulation to count luteal cell numbers and measure relative mRNA abundance for steroidogenic and angiogenic enzymes and LH receptor. Treatment with NGF increased pre-ovulatory follicle diameter (P = 0.02) and serum LH concentrations (P = 0.03) but did not affect time to ovulation (P = 0.42). Heifers treated with NGF had increased serum progesterone concentrations in the subsequent luteal phase (P = 0.03), but no change in vascular area of the follicle (P = 0.16) or CL (P = 0.20). Heifers treated with NGF had a greater number of small luteal cells (P < 0.01) and a tendency for increased LH receptor (LHR) mRNA abundance in the CL (P = 0.10). There was also increased steroidogenic acute regulatory protein (STAR; P = 0.05) and a tendency for increased cytochrome P450 family 11 (CYP11A1; P = 0.10) mRNA abundance in the CL of NGF-treated heifers. There was decreased prostaglandin E 2 synthase (PGES; P = 0.03) and its receptor (PGER; P = 0.05) mRNA abundance and a tendency for decreased cytochrome P450 family 17 subfamily A member 1 (CYP17A1; P = 0.08) and hydroxysteroid 17-beta dehydrogenase (HSD17B; P = 0.06) mRNA abundance in the CL of NGF-treated heifers. Administration of NGF improved CL function in heifers potentially as a result of increased LH release., Competing Interests: Declaration of competing interest None., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The aim of the study was to determine whether two postpartum intramuscular treatments with 200 mg of beta-(β-)carotene (Carofertin; Alvetra u. Werfft, Vienna, Austria) in a 14-day interval increases β-carotene concentrations in blood, particularly around the time of the first artificial insemination (AI), and to test the effect of the treatment on fertility parameters, luteal size, and progesterone blood levels of dairy cows. A total of 297 Holstein dairy cows were enrolled in the study. Between 28 and 34 days postpartum (dpp) β-carotene concentrations were measured in blood samples using an on-site test (iCheck carotene; BioAnalyt, Teltow, Germany). Cows with a β-carotene concentration <3.5 mg/L, indicating a deficiency of β-carotene, were allocated either to the β-carotene treatment group BCT (n = 123) or to the control group CON (n = 121). Cows with concentrations ≥3.5 mg/L were assigned to an optimally supplied reference group (REF; n = 53). Cows in the BCT group received 200 mg of β-carotene intramuscularly at 28-34 dpp and at 42-48 dpp. Further blood samples were collected at 35-41 dpp, 42-48 dpp, 49-55 dpp, and in the week after the first AI and their β-carotene concentrations were analyzed. Between day 10 and 14 after the first AI, the blood progesterone concentration was measured and the size of the corpus luteum (CL) was determined by ultrasound. Blood β-carotene concentrations increased in the BCT cows in the week after the treatment with a peak at 49-55 dpp and were significantly higher than in the CON group at each time point after the first treatment. Logistic regression models, however, revealed that the treatment with β-carotene had no effect on first service conception rate, days to first service, time to pregnancy, or percentage of pregnant cows within 150 dpp. Furthermore, there was no effect on progesterone concentration or the size of the CL between the groups. In conclusion, two treatments with Carofertin postpartum increased β-carotene blood concentrations but had no effect on the fertility parameters in this study., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The objective was to compare pregnancy per AI (P/AI) between two shortened timed-AI (TAI) protocols in beef cattle. This study also determined whether administration of eCG in heifers and timing of AI in cows would affect P/AI. Cattle were submitted at random to either a modified 5-d Co-synch protocol (Day 0 = progesterone releasing device (CIDR); Day 5 = CIDR removal and 500 μg of cloprostenol (PGF); Day 8 = 100 μg GnRH concurrent with AI) or J-synch protocol (Day 0 = CIDR insertion and 2 mg of estradiol benzoate i.m.; Day 6 = CIDR removal and 500 μg PGF; Day 9 = 100 μg GnRH concurrent with AI). In Experiment 1, 1135 heifers (13-15 mo of age) received an estrus detection patch (Estrotect™) on Day 5 and 579 were selected at random to receive 300 IU of equine chorionic gonadotropin (eCG) at the time of CIDR removal. Patches were scored from 0 to 3 based on color change between initial application and AI; 0 = unchanged, 1 = ≤ 50% change, 2 = > 50% change, 3 = missing. Estrus was defined to have occurred when the patch was scored 2 or 3. In Experiment 2, 399 cyclic, non-lactating beef cows from 1 location were submitted to either the modified 5-d Co-synch or J-synch protocol and within each protocol cows were TAI at either 66 ± 1 (n = 199) or 72 ± 1 h (n = 200) following CIDR removal. Transrectal ultrasonography was used in both experiments to determine presence of a corpus luteum (CL) on Day 0, and to diagnose pregnancy 35 d after TAI. In Experiment 1, eCG increased estrus rate only in heifers without a CL on day 0 that were submitted to the modified 5-d Co-synch protocol (41.9 vs. 69.6%). Heifers submitted to the J-synch protocol had greater (P = 0.03) P/AI compared with those in the modified 5-d Co-synch (48.7 vs. 41.1%) and heifers that expressed estrus before AI had increased (P < 0.0001) P/AI compared to those that did not (53.6 vs. 36.5%). Administration of eCG and presence of a CL tended to affect P/AI (P = 0.13). In Experiment 2, cows submitted to the J-synch protocol tended (P = 0.07) to have greater P/AI compared to those in the modified 5-d Co-synch (74.1 vs. 66.5%). There was no association between P/AI and timing of AI. In summary, the J-synch protocol resulted in greater P/AI than the modified 5-day Co-synch protocol in heifers and cows. Administration of eCG increased estrus rate in heifers without a CL at the start of the protocol and tended to improve P/AI in all heifers. Timing of AI (66 vs. 72 h) had no effect on P/AI in cows subjected to either TAI protocol., (Crown Copyright © 2019. Published by Elsevier Inc. All rights reserved.)

Research Question: What are the safety and feasibility of repeated subcutaneous doses of gonadotrophin-releasing hormone (GnRH) agonist for luteal support in IVF cycles triggered by a GnRH agonist?, Design: In this prospective trial, patients exhibiting oestradiol concentrations of over 2500 pg/ml after use of a GnRH agonist for triggering ovulation were initially randomized to GnRH agonist luteal support (0.1 mg subcutaneously every other day, starting on day 3 after embryo transfer) or to a control group supported by 80 µg of recombinant human chorionic gonadotrophin (HCG) on day 3 after embryo transfer. All patients underwent a day 5 blastocyst transfer. Randomization to the HCG luteal support was stopped owing to two cases of ovarian hyperstimulation syndrome (OHSS) and the study was continued solely with GnRH agonist luteal support., Results: The study included 39 women in the repeated GnRH agonist luteal support group and seven in the HCG micro dose group. There were no cases of OHSS among patients supported by a GnRH agonist, and no other adverse events were recorded. There were no cases of bleeding before the pregnancy test, and hence no cases of an insufficient luteal phase. A clinical pregnancy rate of 43.6% was achieved with GnRH agonist luteal support. Hormone dynamics during the stimulation cycle reflected rising LH and progesterone concentrations after the introduction of GnRH agonist support., Conclusions: Repeated doses of GnRH agonist every other day as a method of luteal support provided safe and effective luteal support for women who underwent GnRH agonist triggering in a GnRH antagonist IVF cycle., (Copyright © 2019 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.)

The use of tail chalk and estrus/heat expression scores (HEATSC) evaluation is instrumental in identifying cows with greater estrus expression and greater artificial insemination pregnancy rates (P/AI) in cows submitted to timed artificial insemination (TAI), and cows with low or no estrus expression present lower P/AI. It was intended in this study to improve the pregnancy rates in TAI for Bos indicus beef cows, and gonadotrophin-releasing hormone (GnRH) injection was hypothesized to increase pregnancy rates in a TAI program for cows submitted to progesterone-estradiol-based protocols with low or no estrus expression, evaluated by HEATSC. Cows (n= 2284) received a progesterone device and 2 mg estradiol benzoate, after 8 days the device was removed and 1 mg estradiol cypionate, 150 μg of d-cloprostenol and 300 IU equine chorionic gonadotropin was administered. All cows were marked with chalk and HEATSC evaluated (scales 1 to 3) at TAI performed on day 10. Animals with HEATSC1 and HEATSC2 (n= 937) received 100 μg de gonadorelin (GNRH group; n= 470), or 1 ml saline (Control group; n= 467), and cows with HEATSC3 (named HEAT group; n= 1347) received no additional treatment. The larger dominant follicle, evaluated on day 8and at TAI (day 10), was greater in HEAT group (P= 0.0145 and P <0.001, respectively). Corpus luteum (CL) area and progesterone concentration was evaluated on day 17, and CL area was larger in HEAT group, intermediary in Control and lower in GnRH group (Control= 2.68 cm2, GnRH= 2.37 cm2, HEAT group= 3.07 cm2, P <0.001). Greater progesterone concentrations were found in HEAT group than in Control and GnRH groups (Control= 4.74 ng/ml, GnRH= 4.29 ng/ml, HEAT group= 6.08 ng/ml, P<0.001). There was a difference in ovulation rate, greater in HEAT group than GnRH and Control groups (Control= 72.5%; GnRH= 81.25%; HEAT group= 90.71%; P= 0.0024). Artificial insemination pregnancy rates was greater in HEAT group (57.09% (769/1347) than in Control and GNRH groups, with positive effect of GnRH injection at the time of TAI in P/AI (Control= 36.18% (169/467), GnRH= 45.95% (216/470); P<0.0001). In conclusion, GnRH application in cows with low HEATSC (1 and 2) is a simple strategy, requiring no changes in TAI management to increase pregnancy rates in postpartum beef cows submitted to progesterone-estradiol-based TAI protocols, without reaching, however, the pregnancy rates of cows that demonstrate high estrus expression at the TAI.

Long-acting injectable progesterone (iP4) supplementation during early diestrus is a strategy to increase conception rates in cow-calf beef operations. However, iP4 treatment causes early functional and structural regression of the corpus luteum (CL) in a proportion of iP4-treated animals, resulting in pregnancy loss. The hypothesis evaluated was that iP4 accelerates downregulation of sex-steroid receptors (PGR, ESR1, ESR2) during early to mid-diestrus and the upregulation of genes controlling PGF 2 α secretion (OXTR, PTGS2, AKR1B1) during late diestrus in the endometrium. Ovulations of cyclic, multiparous Nelore (Bos indicus) cows were synchronized, and cows were divided to receive placebo or 300 mg iP4 3 d postovulation (D3). Growth and vascularization of luteal tissue were evaluated by ultrasonography. Blood samples were collected from 3 d postovulation to 3 d after luteolysis, and P4 plasma concentrations were measured by radioimmunoassay. On days 3, 5, 7, 9, 11, 13, and 16 luminal endometrial samples were taken using a cytologic brush. Transcript abundance was measured by qPCR. Structural luteolysis occurred 3 d earlier in cows receiving iP4 compared to the control group. Analyzing only cows that received iP4, those that presented early luteolysis (ie, ≤ D16) showed a decrease in CL area and P4 concentration after D5, compared to the control group. Cows that presented early luteolysis showed a reduced abundance of transcripts on D5 for the ESR2 gene and a greater abundance of transcripts for OXTR and ESR1 on D16, compared to cows that did not present early luteolysis. The iP4-induced early luteolysis can be explained by two nonexclusive possibilities: the activation of uterine mechanisms that trigger early secretion of endometrial PGF pulses and the formation of a subfunctional CL that is prone to early regression., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The objective of this study was to use estrous behavior alone to determine the appropriate time for beginning an oxytocin treatment protocol for estrus suppression. We hypothesized that administration of oxytocin beginning 8 days after the onset of estrus will prolong the luteal phase in mares. Twenty-three light breed mares (aged 4-20 years) were exposed to a stallion and observed for signs of sexual receptivity. Mares not displaying signs received 250 μg of cloprostenol intramuscularly (IM) and were teased again 3-4 days later. On the day that estrous behavior was observed (Day 0), mares were randomly divided into two groups: oxytocin (n = 11): oxytocin (60 IU, IM) was administered once daily from Day 8-17; control (n = 12): did not receive treatment. Blood was collected from all mares every 4 days throughout Day 17, and every 7 days thereafter until Day 45. Serum progesterone concentrations >1.0 ng/mL were indicative of a functioning corpus luteum. Interestrus interval was defined as the period between Day 0 and the day when progesterone next reached <1.0 ng/mL. The average interestrus interval was higher for treated mares compared with control mares (32.4 ± 4.2 vs. 21.8 ± 1.5 days, respectively, P = .01). In the oxytocin group, the interestrus interval was longer than 31 days in 6 of 11 (54.5%) mares and up to 45 days in 5 of 11 mares (45.5%). We conclude that luteal maintenance beyond 30 days was attained by once-daily oxytocin administration beginning 8 days following behavioral estrus in a majority of mares., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Heifers have two or three major follicular waves per interovulatory interval (IOI). In mares and women, the ovulatory wave is the only major wave in most (75%) IOI. The beginning of diameter deviation during follicle selection of the future dominant follicle (DF) is followed by continued growth of DF and decreasing growth of the future subordinate follicles. Diameter deviation in Bos taurus heifers, mares, and women begins when the future DF is a mean of 8.5, 22.5, and 10.5 mm, respectively. Selection of the ovulatory follicle occurs more frequently from right ovary (RO) in heifers and women and from left ovary (LO) in nulliparous mares with no difference between ovaries in parous mares. The RO predilection for ovulation is preceded by a predilection for more follicles in RO before the beginning of deviation as indicated by (1) in heifers and women, there were more predeviation follicles in RO than LO and ovulation occurred more frequently from RO whereas in mares there was no difference between ovaries in number of predeviation follicles and ovulation occurred with similar frequency between ovaries and (2) in heifers, the number of ovulatory waves with DF in the ovary that had more predeviation follicles was greater than the number of waves with DF in the ovary that had fewer follicles. In heifers, ovulation from RO occurs more frequently when the regressed CL is also in RO and is attributable to a positive intraovarian effect of the CL on predeviation follicles that were adjacent to the CL. The positive two-way effect between CL and future DF when adjacent is expressed by greater dimensions and vascular perfusion of CL and DF. This phenomenon awaits study in mares and women. An exception to more frequent RO ovulation in heifers occurs in wave 3 owing to a switch during predeviation in future dominance to a smaller follicle when the largest follicle is adjacent to the regressing CL. A preovulatory contralateral relationship (DF and CL in opposite ovaries) during the last wave of an IOI in heifers usually (eg, 88%) converts to an ipsilateral relationship during wave 1 of the next IOI in association with continuity in vascular perfusion and number of predeviation follicles per ovary. Alternating relationships between consecutive ovulations were not found in mares and is controversial in women. Applied potential of ovarian asymmetry is indicated by greater blastocyst rate for RO oocytes in cattle and greater pregnancy rate for RO ovulation in women., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The aim of this study was to determine the expression of fibroblast growth factor 22 (FGF22) in the bovine corpus luteum (CL) and to investigate the effects of in vivo total or partial cloprostenol-induced luteolysis on the mRNA abundance of FGF22 and its receptor, FGFR1B. Corpora lutea at different stages of development were then dissected from abattoir ovaries (n = 10/stage); a portion of the tissue samples was fixed in paraformaldehyde and the remaining samples were homogenized and subjected to total RNA extraction. To assess mRNA abundance of target genes during induced luteolysis, nineteen cows were synchronized and then randomly assigned to a Latin square design as follows: Control; 2 administrations of prostaglandin F 2α (PGF 2α , total luteolysis; 2 × 250 μg of cloprostenol sodium) and 1/6PGF 2α (partial luteolysis; 83.33 μg of cloprostenol sodium). FGF22 and FGFR1B expression levels were measured by RT-qPCR, and FGF22 protein expression was detected by immunohistochemistry. In summary, FGF22 mRNA was detected at all stages of CL development, and FGF22 protein was also detected in luteal tissue. FGF22 mRNA expression was lower at stage IV than at stage III (P < 0.05), and the same pattern was observed in luteal immunoreactivity. Furthermore, cloprostenol-induced luteolysis, both total and partial, increased FGFR1B mRNA abundance in luteal tissue (P < 0.05), but did not affect FGF22 mRNA abundance. In conclusion, these data suggest a potential role for the FGF22-FGFR1B system during development and regression of bovine CL., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Donkey jenny's corpus luteum (CL) response to PGF2α analogues has not been investigated in depth. Aim of this study was to evaluate the donkey jenny's corpus luteum (CL) ultrasonographic characteristics (diameter, area and vascularized area) by B-Mode, Color Doppler and serum progesterone concentration ([P4]) after treatment with the prostaglandins F2α analogue alfaprostol at day 3 or day 6 after ovulation (groups PG3 and PG6, respectively). [P4] was positively correlated (P < 0.0001) with CL diameter: r 2  = 0.17; area: r 2  = 0.21 and vascularized area: r 2  = 0.54. The interovulatory interval was significantly reduced in the PG6 group (15 ± 1.8 days), compared to the control group (24.5 ± 2.9 days; P < 0,05), while there were no significant differences in interovulatory interval between PG3 (21.7 ± 7.9 days) and control or PG6 group. [P4], in the 6 jennies of the PG6 group, dropped under 1 ng/mL within 2 days after treatment, remaining under this concentration until [P4] raised again to levels comparable with those of the control group until spontaneous luteolysis. After alfaprostol administration, one of the 2 remaining PG3 group jennies showed a complete luteolysis, and the other one underwent a partial luteolysis and ovulated in diestrus., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Two experiments were performed to evaluate the reproductive performance of zebu beef cows treated with different doses of eCG at the end of a progesterone (P4)/estrogen based protocol for timed artificial insemination (TAI). In Experiment 1, suckling Bos indicus Nelore cows (n = 261) received, on day 0, a progesterone (P4) intravaginal device (PD) and an injection of 1 mg estradiol benzoate (EB). On day 8, the PD was removed, 500 μg of cloprostenol was injected, and cows were assigned to one of the following groups: Control (no treatment), 300 (300 IU of eCG), 600 (600 IU of eCG), and 900 (900 IU of eCG). On day 9, all cows received 1 mg EB and TAI performed 54-56 h after cloprostenol injection. A pregnancy diagnosis was done by ultrasound scanning 40 days after TAI, and the number of fetuses and calves was recorded at pregnancy diagnosis and at birth. More cows treated with eCG displayed estrus within 48 h after removal of the PD (42.3% vs. 11.6%, P < 0.01), and ovulated more than one follicle (42%, 58/138 vs. 1.8%, 1/54; P < 0.01). This effect on ovulation rate was dose dependent (P < 0.05). The pregnancy rate was affected only by cow parity (primiparous, 25.3% vs. multiparous, 48.9%; P < 0.01). Twin pregnancy was higher (P < 0.01) in cows treated with eCG (42%, 58/138) than controls (0%, 0/54). However, few cows (33.3%) were able to keep both fetuses intact until birth. For evaluation of ovarian characteristics by B-mode and Doppler ultrasonography, 43 Nelore cows were submitted In Experiment 2 to the same four groups described in Experiment 1. Although no difference (P > 0.1) was observed for size and blood perfusion in the pre-ovulatory follicles, corpus luteum was larger and with greater blood perfusion (P < 0.05) in eCG-treated cows. In conclusion, eCG increased the number of double/multiple ovulations in a dose-dependent manner, induced larger and more vascularized corpora lutea, but did not affect the fertility of cyclic or anestrous cows. Although eCG results in twin pregnancies, most of cows underwet embryo/fetus loss and birth a single calf., (Copyright © 2018 Elsevier Inc. All rights reserved.)

This study investigated molecular regulation in the canine corpus luteums/corpora lutea (CL) following multiple low-dose prostaglandin F 2 alpha (PGF 2 α) injections in non-pregnant bitches around 30-35 days after ovulation. The CL were obtained by ovariohysterectomy 1 h after the last PGF 2 α injection. The subjects were divided into the following groups: control (no PGF 2 α injection, n = 4), one PGF 2 α injection (injection at 0 h, 1PGF, n = 4), two PGF 2 α injection (injection at 0 and 8 h, 2PGF, n = 4), and three PGF 2 α injection (injection at 0, 8 and 24 h, 3PGF, n = 4). In the 1PGF group, the steady-state mRNA levels of an immediate early gene (NR4A1) and immune system-related genes (MCP-1 and IL-8) increased. NR4A1 was localized in luteal and endothelial cells. In contrast, MCP-1 was localized in the luteal tissue between the luteal and endothelial cells. LHCGR, CYP11A1, and StAR mRNA expression decreased after the second PGF 2 α injection. FASLG increased only after the third PGF 2 α injection. The mRNA levels of PTGFR, PGT, and PTGS2 decreased as the number of PGF 2 α injections increased. Immunohistochemistry showed a decrease in StAR protein density as the number of PGF 2 α injections increased. BAX and CASP3 mRNA expression levels were similar among the groups. Serum progesterone (P 4 ) levels decreased dramatically after the PGF 2 α injections but were still higher than the basal level at the end of the study. In conclusion, repeated low-dose PGF 2 α injections could induce luteolytic mechanisms in the CL of non-pregnant bitches. Furthermore, it can be concluded that, in non-pregnant bitches, some aspects of the molecular regulation of luteolysis in the CL are similar to some aspects of such regulation in other domestic animals., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The direct effect of melatonin on porcine luteal function during the pregnancy remains unknown. The objective of the study was to analyse the molecular mechanism(s) by which melatonin directly affects progesterone (P4) production in the corpus luteum (CL) of pregnant sows. We evaluated the localization of melatonin membrane receptors (MT1 and MT2) in CL, and investigated the effect of melatonin on P4 secretion along with the expression of P4 synthesis intermediates in luteal cells. Immunohistochemistry analysis showed that MT1 and MT2 were predominantly localized in luteal cells in pregnant luteal tissues. The results of our in vitro experiments showed that melatonin from 5 to 625 pg/mL was able to significantly increase P4 release (P < 0.05) in a dose-dependent manner. And at the dose of 125 pg/mL treatment, the time-dependent effect on P4 secretion was observed. Furthermore, melatonin from 5 to 625 pg/mL up-regulated both P450scc and StAR expression (P < 0.05) in a dose-dependent manner, and the effect was also time-dependent. No difference of 3β hydroxysteroid dehydrogenase (3β-HSD) expression was observed between control and treatment groups. In addition, melatonin induced a dose- and time-dependent promotion on cell viability. Additionally, the stimulatory effects of melatonin were blocked by luzindole, a non-selective MT1 and MT2 receptor antagonist, or partially blocked by a selective MT2 ligand, 4-phenyl-2-propionamidotetralin (4P-PDOT). The data support the presence of MT1 and MT2 in porcine CL and a regulatory role for melatonin in luteal function through MT1 and MT2-mediated signal transduction pathways., (Copyright © 2017 Elsevier Inc. All rights reserved.)

This study evaluated the effect of initial GnRH and timing of AI in a 5-d Co-synch plus CIDR (device containing 1.38 g of progesterone) protocol on pregnancy per AI (P/AI) and pregnancy loss in beef heifers. A secondary objective was to determine if the effect of initial GnRH on reproductive performance was influenced by cyclicity. Crossbred beef heifers (n = 1068; 301-514 kg of body weight, and 13-15 mo of age) at three locations were assigned to either a 5-d Co-synch plus CIDR protocol with (CIDR5G) or without (CIDR5NG) an initial injection of 100 μg of GnRH at CIDR insertion (Day 0). All heifers received a single dose of 500 μg of cloprostenol at CIDR removal (Day 5) and were divided into two groups to receive GnRH and TAI at either 66 or 72 h (Day 8) after CIDR removal. All heifers were inseminated by one technician with frozen-thawed semen from 1 of 4 sires available commercially. Transrectal ultrasonography was performed on Day 0 to determine cyclicity (presence of CL) and normalcy of the reproductive track, and 27 d after TAI to determine pregnancy status. Non-pregnant heifers (n = 470) were assigned to either a CIDR5G or a CIDR5NG protocol with TAI at 72 h after CIDR removal. Twelve days after second AI, heifers were exposure to bulls for 20 d and pregnancy diagnoses were performed approximately 30 d after second TAI and 60 d after bulls were removed to diagnose bull pregnancies and determine pregnancy loss rate. The percentage of acyclic heifers was 20.3%. Overall P/AI after first TAI was 55.6% (594/1068) and did not differ between CIDR5G and CIDR5NG (56.1 vs. 55.1%), or between TAI66 and TAI72 (55.8 vs. 55.4%). However, cyclic heifers were more likely to become pregnant than acyclic ones (59.3 vs. 41.2%; P < 0.01). Moreover, acyclic heifers subjected to the CIDR5NG had fewer P/AI than those subjected to CIDR5G (P < 0.01). Overall P/AI after resynchronization was 55.1% and did not differ between CIDR5G and CIDR5NG (51.3 vs. 59.0%). Overall pregnancy loss after first and second TAI were 3.0% (18/594) and 3.9% (8/205), respectively. When pregnancy loss data were combined, synchronization protocol (4.1 vs. 2.3% for CIDR5NG and CIDR5G; P = 0.01), cyclicity (5.8 vs. 2.9% for acyclic and cyclic; P = 0.03) and the interaction between synchronization protocol and cyclicity (P = 0.04) were significant. The overall cumulative pregnancy at the end of the breeding season was 94.2% (1006/1068); acyclic heifers were less likely to be pregnant at the end of the breeding season (88.4 vs. 95.8%; P < 0.01). In summary, the initial GnRH administration in a 5-d Co-synch plus CIDR protocol that includes a single PGF treatment is necessary in acyclic beef heifers to optimize P/AI, but not in cyclic heifers. Moreover, omission of initial GnRH was associated to greater pregnancy losses, particularly in acyclic heifers. Timing of AI did not affect P/AI., (Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.)

The objectives of this study were 1) to monitor corpus luteum (CL) dynamics after two different protocols of ovulation induction in prepubertal Nellore heifers, and 2) to determine differences in luteal function. Fifty-seven heifers (weight 289.61 ± 32.28 kg, BCS 5.66 ± 0.65, age 17.47 ± 0.81 months) were divided into two groups: GP4+GnRH received a progesterone (P4) device of 3rd use for 10 days, followed by the administration of 0.02 mg buserelin acetate (GnRH) 48 h after removal of the device, and GGnRH received only GnRH. The CLs formed were monitored by ultrasonography every 2 days until their functional regression (decrease in the color Doppler signal and serum P4 concentration < 1 ng/mL), determining their diameter and area, numerical pixel value (NPV), pixel heterogeneity, and vascularization percentage. The peak systolic velocity, end diastolic velocity, resistivity index and pulsatility index (PI) of the ovarian artery and serum P4 concentration were also measured. A lifespan of the CL of more than 16 days was classified as normal-function and of less than 16 days as premature regression. The variables were compared between treatments, CL categories (normal-functional, prematurely regressed or non-functional), days of evaluation, and their interactions using the MIXED procedure of the SAS program (p ≤ 0.05). Three animals of each group (6/57 = 11%) did not respond to treatment, corresponding to an ovulation rate of 89%. There was a higher percentage of normal-function CLs in GP4+GnRH (81%) and a higher percentage of non-functional CLs in GGnRH (52%; P4 concentration < 1 ng/mL in all assessments). Normal-function CLs exhibited a greater area, vascularization percentage and P4 concentration than prematurely regressed and non-functional CLs. Lower diameter, area, NPV and P4 concentration were observed for non-functional CLs, but there was no difference in vascularization percentage compared to prematurely regressed CLs. Progesterone concentration was efficient in diagnosing CL function and was positively correlated with CL area (r = 0.62; p < 0.001) and vascularization percentage (r = 0.38; p < 0.001). Diameter and PI were important for the early diagnosis of non-functional and prematurely regressed CLs, respectively. In conclusion, luteal function differed for the first CL that develops after ovulation induction in prepubertal heifers. Ultrasonographic parameters (diameter, area, NPV, vascularization percentage, and PI) can be used to predict CL function., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The objective of the current study was to determine if Nerve Growth Factor-Beta (NGF), purified from bovine seminal plasma, would improve corpus luteum function and enhance conceptus development when administered to cows at artificial insemination. Angus cows (n = 60) were synchronized using a GnRH-prostaglandin and intravaginal progesterone protocol (7-day CO-Synch/CIDR) and randomly allocated to 1 of 2 treatment groups: (1) CONT- 12 mL PBS; or (2) NGF- 296 μg purified NGF in 12 mL PBS administered intramuscularly at insemination (day 0). Blood collections were performed to measure plasma concentrations of progesterone (days 0, 3, 7, 10, 14, 19) and pregnancy-specific protein B (day 24) using immunoassays. Expression of interferon-stimulated genes (ISG15, MX1, MX2, RTP4) were assessed in peripheral blood leukocytes on day 19. Transrectal ultrasound was performed for measuring corpus luteum size (days 0, 3, 7, 10, 14, 19) and pregnancy diagnosis (days 28, 45, 66). Statistical analysis was performed using analysis of variance with repeated measures (SAS 9.4, Cary NC). Corpus luteum volume and diameter increased over time (p < 0.001), but did not differ between treatment groups (p = 0.46). Cows treated with NGF had increased plasma progesterone over CONT cows from days 10-19 (p = 0.04). Pregnancy rates at day 28 were 75% in NGF cows versus 59% in CONT cows (p = 0.13). In pregnant cows, pregnancy-specific protein B concentrations at day 24 were greater in NGF than CONT cows (p < 0.05). Additionally, fold-change expression of ISG15 and MX2 at day 19 were greater in pregnant NGF cows than in pregnant CONT cows (p < 0.05), but no differences for MX1 and RTP4 were present. Here we demonstrate that NGF administration to cows at insemination improved corpus luteum function, which translated to improved early conceptus development as determined by upregulation of interferon-stimulated genes and increased pregnancy-specific protein B concentrations. These results suggest that seminal plasma NGF could play a role in conceptus development and may be important to improve fertility in cattle., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Two experiments were conducted with the aim of determining the effect of equine chorionic gonadotropin (eCG) administration on day 14 after insemination on ovarian response and pregnancy establishment in postpartum anestrous beef cows. In both experiments, cows were subjected to a progesterone- and estradiol-based treatment for fixed-time artificial insemination (FTAI) and were randomly allocated into 4 groups to receive or not receive eCG (400 IU) at the time of device removal and/or at 14 d after FTAI. In experiment 1, from day 14 to 22, daily ultrasonographic determinations were performed to monitor ovarian dynamics, and blood was collected to determine hormone concentrations in 60 cows. In experiment 2, confirmation of pregnancy was performed at 30 and 60 d after FTAI in 1,060 anestrous cows assigned to the same experimental design. Cows that received eCG on day 14 after FTAI showed increases in corpus luteum area (P < 0.01), follicle diameter (P < 0.05), serum progesterone concentrations (P < 0.01), and estradiol-17β concentrations (P < 0.01), compared with cows that did not receive eCG on day 14. Pregnancy rate on day 30 was greater in those cows that received both eCG treatments (ie, at device removal and 14 d after insemination) than in those that did not receive eCG treatment (P < 0.05). In conclusion, eCG administered on day 14 after FTAI increases serum progesterone concentrations during the critical period of pregnancy in anestrous cows, and this second eCG treatment seems to have a positive effect on achieving pregnancy., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The corpus luteum (CL) is an ovarian structure which is critical for the maintenance of reproductive cyclicity and pregnancy support. Diet and/or diet components may affect some luteal functions. FSH is widely used to induce multiple follicle development and superovulation. We hypothesized that FSH would affect luteal function in ewes fed different nutritional planes. Therefore, the aim of this study was to determine if FSH-treatment affects (1) ovulation rate; (2) CL weight; (3) cell proliferation; (4) vascularity; (5) expression of endothelial nitric oxide (eNOS) and soluble guanylate cyclase (sGC) proteins; and (6) luteal and serum progesterone (P4) concentration in control (C), overfed (O), and underfed (U) ewes at the early- and mid-luteal phases. In addition, data generated from this study were compared to data obtained from nonsuperovulated sheep and described by Bass et al. Ewes were categorized by weight and randomly assigned into nutrition groups: C (2.14 Mcal/kg; n = 11), O (2xC; n = 12), and U (0.6xC; n = 11). Nutritional treatment was initiated 60 d prior to day 0 of the estrous cycle. Ewes were injected with FSH on day 13-15 of the first estrous cycle, and blood samples and ovaries were collected at early- and mid-luteal phases of the second estrous cycle. The number of CL/ewe was determined, and CL was dissected and weighed. CL was fixed for evaluation of expression of Ki67 (a proliferating cell marker), CD31 (an endothelial cell marker), and eNOS and sGC proteins using immunohistochemistry and image analysis. From day 0 until tissue collection, C maintained, O gained, and U lost body weight. The CL number was greater (P < 0.03) in C and O than U. Weights of CL, cell proliferation, vascularity, and eNOS but not sGC expression were greater (P < 0.001), and serum, but not luteal tissue, P4 concentrations tended to be greater (P = 0.09) at the early- than mid-luteal phase. Comparisons of CL measurements demonstrated greater (P < 0.01) cell proliferation and serum P4 concentration, but less vascularity at the early and mid-luteal phases, and less CL weight at the mid-luteal phase in superovulated than nonsuperovulated ewes; however, concentration of P4 in luteal tissues was similar in both groups. Thus, in superovulated ewes, luteal cell proliferation and vascularity, expression of eNOS, and serum P4 concentration depend on the stage of luteal development, but not diet. Comparison to control ewes demonstrated several differences and some similarities in luteal functions after FSH-induced superovulation., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The objectives of the study were to determine the effect of seminal plasma β-NGF on Corpus Luteum morphology and function and level of mRNA expression of steroidogenic enzymes. Llamas were assigned (n = 12/per group) to receive an intramuscular dose of: (a) 1 ml phosphate buffered saline (PBS), (b) 5 μg gonadorelin acetate (GnRH), or (c) 1.0 mg of purified llama spβ-NGF. Ovaries were examined by transrectal B-mode ultrasonography from treatment to ovulation (Day 0 = treatment). B mode/Power Doppler ultrasonography and blood samples collection were performed at Days 4, 8 and 10 (n = 3 llamas per treatment group/per time point) to determine CL diameter, vascularization and plasma progesterone concentration respectively. Plasma progesterone concentration was analyzed in all llamas at Day 0. Then females were submitted to ovariectomy at Days 4, 8 and 10 (n = 3 llamas/treatment/time), CL was removed to determine vascular area, proportion of luteal cells and CYP11A1/P450scc and STAR expression by RT-PCR. Ovulation was similar between llamas treated with GnRH or spβ-NGF and CL diameter did not differ between GnRH or spβ-NGF groups by Day 4, 8 or 10. Vascularization area of the CL was higher (P < 0.01) in llamas from the spβ-NGF than GnRH-treated group by Day 4 and 8. Plasma progesterone concentration was higher (P < 0.05) in llamas from the spβ-NGF compared to females of GnRH group by Day 4 and 8. The proportion of small and large luteal cells did not differ between GnRH or spβ-NGF groups by Day 8. CYP11A1/P450scc was upregulated 3 folds at day 4 and 10 by spβ-NGF compared to GnRH. STAR transcription was 3 folds higher at day 4 in females treated with spβ-NGF. In conclusion, the luteotrophic effect of spβ-NGF could be related to an increase of vascularization and up regulation of CYP11A1/P450scc and STAR transcripts enhancing progesterone secretion., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Dietary supplementation of n-3 PUFA decreases the luteolytic PGF 2α and improves the pregnancy rate in the dairy cow. However, its effect in the goat is not known. Accordingly, we studied the effect of supplementation of n-3 PUFA rich Fish oil (FO) on different reproductive events in the goat. Cycling goats (n = 30) were divided into two equal groups and fed an isocaloric and isonitrogenous diet supplemented with either FO (TRT; n = 15) or palm oil (PO) (CON; n = 15) @ 0.6 mL/kg body weight for 72 days during the breeding season. Estrus synchronization was done on day 25 and 36 of supplementation using two PG regimen and the goats in estrus were bred. Mean interval from PGF 2α administration to the onset of estrus was 12 h longer (P < 0.05) in the TRT group than that of CON. The number of preovulatory follicles (POF) and ovulation rate was significantly higher in FO supplemented goats (P < 0.05) by 39.64 and 41.35%, respectively. Though the corpus luteum diameter was significantly higher (P < 0.05) on day 5, 8 and 11 post-breeding in the TRT group, mean serum progesterone (P 4 ) did not differ significantly (P > 0.05). Mean concentration of serum estradiol (E 2 ) was significantly (P < 0.01) lower in the FO supplemented group during day 0-60 post-breeding which could be due to significantly low serum cholesterol (P < 0.01). Though the serum concentration of PGF 2α metabolite (PGFM) and PGE 2 metabolite (PGEM) in the pregnant goats was significantly (P < 0.05) lower in the TRT group on day 16 and 17 post-breeding, the ratio of PGEM to PGFM remained unaffected suggesting a favourable effect of FO supplementation on the early pregnancy. The number of embryos, twinning rate and kidding rate were high in FO supplemented group though it was non-significant. However, gestation length, birth weight of kids and neonatal behaviour were comparable between the groups (P > 0.05). In conclusion, supplementation of n-3 PUFA rich FO significantly increased the number of POF and ovulation rate with numerical increase in the kidding rate. Further, it decreased the serum E 2 and PGFM during the critical window of pregnancy recognition in the doe., (Copyright © 2017 Elsevier Inc. All rights reserved.)

This study examined the effects of fish meal supplementation on spatial distribution of lipid microdomains and lateral mobility of prostaglandin F 2α (FP) receptors on cell plasma membranes of the bovine corpus luteum (CL). Beef cows were stratified by BW and randomly assigned to receive a corn gluten meal supplement (n = 4) or fish meal supplement (n = 4) for 60 d to allow incorporation of fish meal-derived omega-3 fatty acids into luteal tissue. Ovaries bearing the CL were surgically removed between days 10 to 12 after estrus corresponding to approximately day 60 of supplementation. A 200-mg sample of luteal tissue was analyzed for fatty acid content using gas-liquid chromatography (GLC). The remaining tissue was enzymatically digested with collagenase to dissociate individual cells from the tissue. Cells were cultured to determine the effects of dietary supplementation on lipid microdomains and lateral mobility of FP receptors. Luteal tissue collected from fish meal-supplemented cows had increased omega-3 fatty acids content (P < 0.05). Lipid microdomain total fluorescent intensity was decreased in dissociated luteal cells from fish meal-supplemented cows (P < 0.05). Micro and macro diffusion coefficients of FP receptors were greater for cells obtained from fish meal-supplemented cows (P < 0.05). In addition, compartment diameter of domains was larger, whereas resident time was shorter for receptors from cells obtained from fish meal-supplemented cows (P < 0.05). Data indicate that dietary supplementation with fish meal increases omega-3 fatty acid content in luteal tissue causing disruption of lipid microdomains. This disruption leads to increased lateral mobility of the FP receptor, increased compartment sizes, and decreased resident time, which may influence prostaglandin signaling in the bovine CL., (Copyright © 2017 Elsevier Inc. All rights reserved.)

In previous work, an EP2 prostanoid receptor (EP2R) agonist in vivo increased mRNA expression of luteal LH receptors (LHR), unoccupied and occupied luteal; LHR, and circulating progesterone, while an EP3R or FPR agonist decreased; mRNA expression of luteal LHR, unoccupied and occupied luteal LHR, and; circulating progesterone. An EP4R and lysophosphatidic acid (LPA) LPA2R and LPA3R agonists were reported to inhibit luteal function and sirtuins have been proposed to increase prostaglandin synthesis. The objectives were to determine; whether an EP4R, LPA2R, or LPA3R agonist affect ovine luteal function in vivo or; in vitro. In addition, whether sirtuin (SIRT)-1, 2, or 3; LPA2R or LPA3R; or EP1R, EP2R, EP3R, or EP4R agonists affect caruncular endometrial PGF 2α or PGE (PGE1+PGE2) secretion in vitro. Day-10 nonpregnant ewes received a single injection of Vehicle (N = 5); an LPA2R (N = 5); LPA3R (N = 6); or EP4R (N = 5) agonist given into the interstitial tissue of the ovarian vascular pedicle adjacent to the luteal-containing ovary to determine effects on circulating progesterone, mRNA expression of luteal LHR, and luteal unoccupied and occupied LHR. In addition, agonists for LPA2R, LPA3R, EP1R, EP2R, EP3R, or EP4R or SIRT-1, SIRT-2, or SIRT-3 activators were incubated with caruncular endometrial slices in vitro to determine their effect on caruncular endometrial PGF 2α , or PGE secretion. LPA2R, LPA3R, or an EP4R agonist in vivo did not affect (P ≥ 0.05) luteal weight, circulating progesterone, or occupied luteal LHR. However, an LPA2R or EP4R agonist, but; not LPA3R agonist, in vivo increased (P ≤ 0.05) mRNA expression of luteal LHR. An; LPA2R, LPA3R, or EP4R agonist increased (P ≤ 0.05) luteal unoccupied LHR, but; not occupied LHR. An LPA2R, LPA3R, or an EP4R agonist did not affect (P ≥ 0.05); luteal progesterone secretion in vitro. An LPA2R or LPA3R agonist did not affect (P ≥ 0.05) luteal PGF 2α , or PGE secretion in vitro. However, an EP4R agonist tended to decrease (P < 0.066) luteal PGF 2α secretion and increased (P ≤ 0.05) luteal PGE; secretion in vitro. EP1R, EP2R, EP3R, or an EP4R agonist did not affect (P ≥ 0.05); caruncular endometrial PGF 2α secretion in vitro. However, EP1R, EP3R, or an EP4R agonist increased caruncular endometrial PGE secretion in vitro, while two different EP2R agonists did not affect (P ≥ 0.05) caruncular endometrial PGE; secretion. A SIRT-1 activator, but not SIRT-2 or SIRT-3 activators, increased (P ≤ 0.05) caruncular endometrial PGE secretion, while sirtuin 1, 2, or 3 activators did not affect (P ≥ 0.05) caruncular endometrial PGF 2α secretion. In conclusion, receptors for EP4, LPA2, and LPA3 do not appear to be involved; in luteolysis, but EP4R and LPA2R might participate in preventing luteolysis by maintaining luteal mRNA expression for LHR and preventing loss of unoccupied luteal LHR. In addition, SIRT-1, EP1R, EP3R, and EP4R might be involved in; regulating caruncular endometrial PGE secretion, but not PGF 2α secretion., (Copyright © 2017 Elsevier Inc. All rights reserved.)

In the dairy industry, excess dietary CP is consistently correlated with decreased conception rates. However, amount of excess CP effects on reproductive function in beef cattle is largely undefined. The objective of this experiment was to determine the effects of excess metabolizable protein (MP) supplementation from a moderately abundant rumen undegradable protein (RUP) source (corn gluten meal: 62% RUP) on ovarian function and circulating amino acid (AA) concentrations in beef cows consuming low quality forage. Non-pregnant, non-lactating beef cows (n=16) were allocated by age, BW and body condition score (BCS) to 1 of 2 isocaloric supplements designed to maintain BW for 60 days. Cows had ad libitum access to corn stalks and were individually offered a corn gluten meal-based supplement daily at 125% (MP125) or 150% (MP150) of National Research Council (NRC) MP requirements. After a 20-day supplement adaptation period, cows were synchronized for ovulation. After 10 days of synchronization, follicular growth was reset with gonadotropin releasing hormone. Daily thereafter, transrectal ultrasonography was performed to diagram ovarian follicular waves, and blood samples were collected for hormone, metabolite and AA analyses. After 7 days of observation of estrus, corpus luteum (CL) size was determined via ultrasound. Data were analyzed using the MIXED procedures of SAS. No differences (P⩾0.21) in BW and BCS existed throughout the study; however, plasma urea N at ovulation was greater (P=0.04) in MP150. Preovulatory ovarian follicle size at dominance, duration of dominance, size at spontaneous luteolysis, length of proestrus and wavelength were not different (P⩾0.11) between treatments. However, ovulatory follicles were larger (P=0.04) and average antral follicle count was greater (P=0.01) in MP150 than MP125. Estradiol concentration and ratio of estradiol to ovulatory follicle volume were not different due to treatment (P⩾0.25). While CL volume 7 days post-estrus was greater (P<0.01) in MP150 than MP125, circulating progesterone 7 days post-estrus and ratio of progesterone to CL volume were not different (P⩾0.21). Total AA were not different (P⩾0.76) at study initiation or completion; however, as a percent of total AA, branched-chain AA at ovulation were greater (P=0.02) in MP150. In conclusion, supplementation of CP at 150% of NRC MP requirements from a moderately undegradable protein source may enhance growth of the ovulatory follicle and subsequent CL compared with MP supplementation at 125% of NRC MP requirements.

Research was conducted to determine the effect of altrenogest and exposure to exogenous gonadotropins on ovarian function in prepubertal and mature gilts. Crossbred, presumably sexually mature gilts (n = 51), were fed with altrenogest for 18 consecutive days and the day after the last feeding with altrenogest, gilts were treated with eCG and 72 hours later challenged with hCG. Animals were slaughtered on Days 10 to 13 of their gonadotropins synchronized estrous cycle. Ovaries were examined for the number of CL, number of follicular cysts, and presence of corpora albicantia. Gilts were divided into two groups: those possessing corpora albicantia (group A-mature; n = 36) and those without corpora albicantia (Group W-prepubertal; n = 15) on their ovaries. In addition, each group was divided into two subgroups depending on the presence of follicular cysts (AC and WC) or their absence (AO and WO). There was no difference between the number of CL in group A and group W. Presence of corpora albicantia determined percentage of gilts possessing follicular cysts (13.9% group A vs. 66.7% group W). Gilts without follicular cysts (AO plus WO; n = 36) had higher number of CL (P < 0.01) than gilts bearing cysts (AC plus WC; n = 15). Comparison AO-AC did not show significant difference (P = 0.075) between CL number in mature cyst-free and cysts bearing gilts. A prepubertal gilts not bearing follicular cysts (WO) had higher (P < 0.02) number of CL than gilts bearing cysts. A significant negative correlation between the number of CL and number of follicular cysts was found (r = -0.664; P = 0.007). There were no differences in blood plasma progesterone and estradiol concentration between cyst-free and cyst-bearing gilts. These results indicate: (1) a higher follicular cysts appearance in prepubertal than mature gilts challenged with altrenogest and exposed to exogenous gonadotropins and (2) a negative effect of follicular cysts on the number of CL (ovulations) in prepubertal gilts., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Oxytocin (OXT) has been used to prolong the luteal phase in mares, but its mechanism of action is unknown. The aim of this study was to evaluate the effect of chronic exogenous OXT administration to mid-luteal phase mares on luteal maintenance. Also, endometrial expression of prostaglandin endoperoxide synthase 2 (PTGS2), prostaglandin F 2 α, E 2 and I 2 synthases (AKR1C3, PTGES, and PTGIS), oxytocin receptor (OXTR), progesterone receptor (PGR), and estrogen receptors 1 (ESR1) and 2 (ESR2) were assessed in mares experiencing luteal maintenance 2 weeks after chronic exogenous OXT administration. Control mares (n = 5; C group) received 6 mL of saline im, whereas OXT (60 units/mare) was administered im (n = 6; OXT group), every 12 hours, on days 7 to 14 postovulation. After endometrial biopsy in groups C (Day 10) and OXT (Day 24), luteolysis occurred within 3 or 6 days, respectively. Luteal maintenance took place in 4 of 6 (67%) of OXT-treated mares. Progesterone in C group was the highest on biopsy day (P < 0.05). In OXT mares, PTGS2, ESR1 (P < 0.05), PTGES, PTGIS, PGR, and ESR2 (P < 0.01) gene transcription decreased, whereas OXTR increased (P < 0.05) in comparison with the C group. In OXT-treated mares, endometrial ESR2 protein expression decreased (P < 0.05), but OXTR increased (P < 0.05) compared with control animals. In both experimental groups, PTGS2 was mainly immunolocalized in surface epithelium, whereas AKR1C3, PTGES, PTGIS, and PGR were in surface and glandular epithelia. ESR1 and ESR2 were found in glandular epithelium and OXTR in stromal cells. High immunolabeling for PTGES, PTGIS, PGR, and OXTR and low for ESR2 was detected in endometrium of OXT-group mares with extended diestrus. Prolonged luteal function associated with chronic OXT treatment may be related to different spatial expression of OXTR and PGR in the endometrium. The observed reduction of endometrial ESR2 may be responsible for the maintenance of PGR in luminal and glandular epithelium. Also, ESR2 may attenuate the transcriptional activity of ESR1 in mare endometrium. This study offers new knowledge on the endometrial expression of ovarian steroids and OXT receptors in OXT pharmacologically induced luteal maintenance in the mare., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The porcine corpus luteum (CL) displays delayed sensitivity to PGF-2α (luteolytic sensitivity, [LS]) until days 12 to 13 of cycle. The control of LS is unknown, but it is temporally associated with macrophage (which secrete tumor necrosis factor-α; TNF-α) infiltration into the CL. Other studies showed that TNF-α induces LS in vitro and that prostaglandins (PGs) may be involved in this mechanism. In experiment 1, PGF-2α and PGE secretion by luteal cells (LCs) was measured on days 4 to 14 of the estrous cycle, and the expression of PTGFS/AKR1B1 and PTGES/mPGES-1, determined by Western blot, before (day 7) vs after (day 13) the onset of LS. Results showed that the PGF-2α:PGE ratio increased significantly (P < 0.05) from day 4 to 13-14, and PTGFS/AKR1B1 and PTGES/mPGES-1 were significantly increased (P < 0.05) on day 13 (vs day 7). In experiment 2, LCs were collected from porcine CL at early (∼days 4-6) or mid (∼days 7-12) stages of the estrous cycle and cultured with 0, 0.1, 1, or 10 ng/mL TNF-α. Results showed that TNF-α significantly increased (P < 0.05) messenger RNA (mRNA) expression of cyclooxygenase (COX)-2 and mPGES-1 but not AKR1B1. TNF-α had no significant effects on AKR1B1 or mPGES protein abundance. TNF-α significantly increased (P < 0.05) PGE-2 but had no effect on PGF-2α secretion or on the PGF-2α:PGE2 ratio. In conclusion, although TNF-α increased COX2 and mPGES-1 mRNA, and PGE-2 secretion in vitro, it did not increase the PGF-2α:PGE2 ratio. Studies are currently directed toward exploring other pathways (eg, FP receptor signaling) by which TNF-α induces LS in the porcine CL., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Progestin concentration in plasma during the early luteal phase is crucial for endometrial function and conceptus development. We hypothesized that periovulatory gonadotrophin treatment via support of luteal function affects endometrial gene expression in horses. Effect of age was analyzed as well. Shetland mares (n = 8, age 4-25 years) were assigned to the following treatments during five consecutive cycles in alternating order following a cross-over design: treatment hCG/-: preovulatory injection of hCG, but no gonadotrophin injection at detection of ovulation, treatment -/hCG: no preovulatory gonadodrophin injection, but injection of hCG at detection of ovulation, treatment eCG/-: preovulatory injection of eCG, but no gonadotrophin injection at detection of ovulation, treatment -/eCG: no preovulatory gonadotrophin injection, but injection of eCG at detection of ovulation, treatment control: no treatment. Concentration of progestin was analyzed by ELISA from the day of ovulation until Day 10. On Day 10, endometrial cells were collected transvaginally by cytobrush technique. Expression of mRNA of cyclooxygenase-2 (COX-2), prostaglandin F2α-synthase, prostaglandin E-synthase, progesterone receptor (PR), estradiol receptor (E2R), acyl-CoA-dehydrogenase (ACAD), uteroglobin (UGB), uteroferrin, and uterocalin was analyzed by RT qPCR. Immunohistological staining of endometrial tissue, obtained via biopsy, was performed for COX-2, PR and UGB. The P4 concentration was influenced by day of cycle (P < 0.01), but not by treatment. No effects of age on gene expression were determined. Neither of the periovulatory gonadotrophin treatments nor age influenced mRNA expression of the genes of interest. Treatment did also not affect immunohistological staining of the endometrium. In contrast, age affected the percentage of PR positive stromal cells (e.g. mare 1 (4 years): 65.5 ± 2.6, mare 2 (24 years): 82.7 ± 2.2%, P < 0.05) and COX-2 positive stained ciliated cells (e.g. mare 1: 15.8 ± 2.9, mare 2: 33.4 ± 6.0%, P < 0.05). In conclusion, no effects of periovulatory gonadotrophin treatment and age on endometrial gene expression in luteal phase pony mares were reported. A lack of treatment effects on luteal function and expression of PRs in the endometrium can at least in part be explained by differences in the reproductive physiology between horses and ponies., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Resumption of ovulatory activity and the timely lysis of the first CL postpartum (pp) are important determinants for the reproductive performance of dairy cows. Cystic ovarian follicles (COFs) and persistent CLs preclude normal ovarian cyclicity and increase the calving interval. The objective of this study was to investigate the effect of GnRH on the incidence of COFs and the effect of PGF2α on the incidence of a prolonged luteal phase (PLP) and on fertility parameters in dairy cows. A total of 476 cows were examined ultrasonographically for the presence of a dominant follicle (12-25 mm, without CL >10 mm; n = 237) or a functional CL (≥20 mm; n = 239) between 28 and 35 days pp and were allocated to one of four groups. Cows with a dominant follicle received 10-μg GnRH (buserelin; group F-T; n = 118) or saline (group F-C; n = 119), and cows with a functional CL received 0.5 mg of a PGF2α analogue (cloprostenol; group CL-T; n = 119) or saline (group CL-C; n = 120) on the day of initial examination, defined as Day 0. Cows were reexamined 7 and 21 days (F-T and F-C) and 3 and 24 days (CL-T and CL-C) later, and COFs were treated immediately after diagnosis in all cows. On the basis of the ovarian findings on Days 21 and 24, cows were treated according to a protocol aimed at timely breeding. The incidence of COFs by Days 7 (F-T vs. F-C; 7.6% vs. 16.8%) and 21 (11.0% vs. 21.8%) decreased (P ≤ 0.03) with GnRH; however, this did not lead to a substantial improvement of calving-to-conception interval (means ± standard error of the mean; 107.91 ± 5.70 vs. 117.94 ± 6.63 days), first-service conception rate (42.3% vs. 41.3%), and number of services per conception (2.06 ± 0.12 vs. 2.31 ± 0.15). Treatment with PGF2α decreased (P < 0.0001) the incidence of PLP by Day 24 (CL-T vs. CL-C; 1.7% vs. 17.5%), decreased calving-to-conception interval(91.28 ± 4.77 vs. 101.75 ± 5.03 days), increased first-service conception rate (63.3% vs. 38.7%), and reduced the number of services per conception (1.65 ± 0.10 vs. 2.08 ± 0.12; each P ≤ 0.01). The results indicate that strategic treatment with GnRH or PGF2α in week 5 pp to induce early ovulation and luteolysis reduces the incidence of COFs and PLP, respectively. Initial treatment with PGF2α also enhanced reproductive performance when used in conjunction with a standardized treatment protocol for all cows in week 8 pp (aimed at timely breeding). In contrast, GnRH did not improve fertility parameters of cystic cows in herds where all cows with COFs were treated as expeditiously as possible., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Gonadotrophin releasing hormone agonist (GnRHa) trigger is effective in the induction of oocyte maturation and prevention of ovarian hyperstimulation syndrome during IVF treatment. This trigger concept, however, results in early corpora lutea demise and consequently luteal phase dysfunction and impaired endometrial receptivity. The aim of this strenghths, weaknesses, opportunities and threats analysis was to summarize the progress made over the past 15 years to optimize ongoing pregnancy rates after GnRHa trigger. The advantages and potential drawbacks of this type of triggering are reviewed. The current approach to the management of GnRHa trigger in autologous cycles is based on the peak serum oestradiol level or follicle number and aims at a fresh embryo transfer or a segmentation approach with elective cryopreservation policy. We recommend intensive luteal support with transdermal oestradiol and intramuscular progesterone alone if peak serum oestradiol is 4000 or more pg/ml after GnRHa trigger or dual trigger with GnRHa and HCG 1000 IU if peak serum oestradiol is less than 4000 pg/mL. On the contrary, we recommend HCG 1500 IU 35 h after GnRHa trigger if there are less than 25 follicles, or freeze all oocytes or embryos if there are over 25 follicles., (Copyright © 2015. Published by Elsevier Ltd.)

A study was performed on the effect of a single dose per mare of 0 (n = 9), 100 (n = 8), or 300 (n = 9) of GnRH on Day 10 (Day 0 = ovulation) on concentrations of LH, FSH, and progesterone (P4) and blood flow to the CL ovary. Hormone concentration and blood flow measurements were performed at hours 0 (hour of treatment), 0.25, 0.5, 1, 2, 3, 4, and 6. Blood flow was assessed by spectral Doppler ultrasonography for resistance to blood flow in an ovarian artery before entry into the CL ovary. The percentage of the CL with color Doppler signals of blood flow was estimated from videotapes of real-time color Doppler imaging by an operator who was unaware of mare identity, hour, or treatment dose. Concentrations of LH and FSH increased (P < 0.05) at hour 0.25 and decreased (P < 0.05) over hours 1 to 6; P4 concentration was not altered by treatment. Blood flow resistance decreased between hours 0 and 1, but the decrease was greater (P < 0.05) for the 100-μg dose than for the 300-μg dose. The percentage of CL with blood flow signals increased (P < 0.05) between hours 0 and 1 with no significant difference between the 100- and 300-μg doses. The results supported the hypothesis that GnRH increases LH concentration, vascular perfusion of the CL ovary, and CL blood flow during the luteal phase; however, P4 concentration was not affected., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Three experiments were done to evaluate the effects of progesterone (P4) supplementation starting during metestrus on formation of the CL and on fertility of lactating dairy cows subjected to fixed-time artificial insemination (FTAI) or embryo transfer (ET). In experiment 1, 42 Holstein cows were randomly allocated to untreated (Control) or to receive an intravaginal implant containing 1.9 g of P4 from Day 3 to 20 after FTAI (controlled internal drug release [CIDR]). Blood samples were collected on Days 3, 4, 7, 11, 14, 17, 20, and 21 after FTAI to evaluate the effect of CIDR supplementation on plasma concentration of P4 using radioimmunoassay. Ultrasound scans were performed at Days 4, 7, 11, 14, and 20 to evaluate CL volume. In experiment 2, the effect on CIDR supplementation on fertility was evaluated in 668 Holstein and crossbred dairy cows that were subjected to FTAI and allocated randomly to untreated (AI-Control) or to receive a CIDR from Day 3 to 17 (AI-CIDR) after FTAI. In experiment 3, 360 Holstein cows were treated with PGF and after heat detection (Day 0), they were allocated to untreated (ET-Control) or to receive a CIDR from Day 4 ± 1 to 8 ± 1 (ET-CIDR-4) or a CIDR from 4 ± 1 to 18 ± 1 (ET-CIDR-14). In vitro-produced embryos were transferred on Day 8 ± 1. Pregnancy diagnoses were performed by ultrasound. In experiment 1, there was interaction between treatment and day in relation to plasma P4 on Days 4 and 7 due to CIDR supplementation. Independent of treatment, pregnant cows had higher plasma P4 from Day 14 to 21 than nonpregnant cows (P ≤ 0.05). Supplementation with CIDR did not alter CL development. In experiment 2, there was no effect of supplementation of P4 on pregnancy per AI on Day 32 (32.0% vs. 31.8%, for AI-Control and AI-CIDR, respectively) or pregnancy loss (15.6% vs. 17.6%, for AI-Control and AI-CIDR, respectively). In experiment 3, P4 supplementation compromised pregnancy per ET (P/ET) on Day 32 in both supplemented groups (39.7% vs. 21.3% vs. 15.2%, for ET-Control, ET-CIDR-4, and ET-CIDR-14, respectively), with no effect on pregnancy loss. Therefore, although CIDR insertion on Day 3 after FTAI did not affect CL function and increased circulating P4, it did not increase pregnancy per AI in lactating dairy cows submitted to FTAI. Moreover, P4 supplementation decreased pregnancy per ET in lactating recipient cows., (Copyright © 2015 Elsevier Inc. All rights reserved.)

To understand the role of ovulation-inducing factor (or nerve growth factor) (OIF [NGF]) in bovine seminal plasma, we (1) used an in vivo llama bioassay to test the hypothesis that bovine seminal plasma induces ovulation and CL development in llamas similar to that of llama seminal plasma when the dose of seminal plasma is adjusted to ovulation-inducing factor content (experiment 1) and (2) determined the effect of bovine seminal plasma on the interval to ovulation and luteal development in heifers (experiment 2). Within species, seminal plasma was pooled (n = 160 bulls, n = 4 llamas), and the volume of seminal plasma used for treatment was adjusted to a total dose of 250 μg of ovulation-inducing factor. In experiment 1, mature female llamas were assigned randomly to four groups and treated intramuscularly with either 10 mL of PBS (negative control, n = 5), 50-μg GnRH (positive control, n = 5), 6-mL of llama seminal plasma (n = 6), or 12 mL of bull seminal plasma (n = 6). Ovulation and CL development were monitored by transrectal ultrasonography. In experiment 2, beef heifers were given a luteolytic dose of prostaglandin followed by 25-mg porcine LH (pLH) 12 hours later to induce ovulation. Heifers were assigned randomly to three groups and given 12 mL bovine seminal plasma intramuscularly 12 hours after pLH treatment (n = 10), within 4 hours after ovulation (n = 9), or no treatment (control, n = 10). Ovulation was monitored by ultrasonography every 4 hours, and the CL development was monitored daily until the next ovulation. In experiment 1, ovulation was detected in 0/5, 4/5, 4/6, 4/6 llamas in the PBS, GnRH, llama seminal plasma, and bovine seminal plasma groups, respectively (P < 0.05). Luteal development was not different among groups. In experiment 2, the interval to ovulation was more synchronous (range: 4 vs. 22 hours; P < 0.0001) in heifers treated with seminal plasma before ovulation compared with the other groups. Luteal development was not different among groups; however, plasma progesterone concentrations tended to be greater in the postovulation treatment group compared with other groups. In summary, results confirmed the presence of bioactive ovulation-inducing factor in bull seminal plasma and supported the hypothesis that bovine and llama seminal plasma have similar ovulatory effects, using a llama bioassay. Treatment with bovine seminal plasma resulted in greater synchrony of ovulation in heifers pretreated with pLH. Plasma progesterone concentration tended to be higher in heifers given bovine seminal plasma within 4 hours after ovulation, suggesting that bovine ovulation-inducing factor is luteotrophic., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Previously, it was reported that intraluteal implants containing prostaglandin E1 or E2 (PGE1 and PGE2) in Angus or Brahman cows prevented luteolysis by preventing loss of mRNA expression for luteal LH receptors and luteal unoccupied and occupied LH receptors. In addition, intraluteal implants containing PGE1 or PGE2 upregulated mRNA expression for FP prostanoid receptors and downregulated mRNA expression for EP2 and EP4 prostanoid receptors. Luteal weight during the estrous cycle of Brahman cows was reported to be lesser than that of Angus cows but not during pregnancy. The objective of this experiment was to determine whether intraluteal implants containing PGE1 or PGE2 alter vascular endothelial growth factor (VEGF), fibroblast growth factor-2 (FGF-2), angiopoietin-1 (ANG-1), and angiopoietin-2 (ANG-2) protein in Brahman or Angus cows. On Day 13 of the estrous cycle, Angus cows received no intraluteal implant and corpora lutea were retrieved, or Angus and Brahman cows received intraluteal silastic implants containing vehicle, PGE1, or PGE2 on Day 13 and corpora lutea were retrieved on Day 19. Corpora lutea slices were analyzed for VEGF, FGF-2, ANG-1, and ANG-2 angiogenic proteins via Western blot. Day-13 Angus cow luteal tissue served as preluteolytic controls. Data for VEGF were not affected (P > 0.05) by day, breed, or treatment. PGE1 or PGE2 increased (P < 0.05) FGF-2 in luteal tissue of Angus cows compared with Day-13 and Day-19 Angus controls but decreased (P < 0.05) FGF-2 in luteal tissue of Brahman cows when compared w Day-13 or Day-19 Angus controls. There was no effect (P > 0.05) of PGE1 or PGE2 on ANG-1 in Angus luteal tissue when compared with Day-13 or Day-19 controls, but ANG-1 was decreased (P < 0.05) by PGE1 or PGE2 in Brahman cows when compared with Day-19 Brahman controls. ANG-2 was increased (P < 0.05) on Day 19 in Angus Vehicle controls when compared with Day-13 Angus controls, which was prevented (P < 0.05) by PGE1 but not by PGE2 in Angus cows. There was no effect (P > 0.05) of PGE1 or PGE2 on ANG-2 in Brahman cows. PGE1 or PGE2 may alter cow luteal FGF-2, ANG-1, or ANG-2 but not VEGF to prevent luteolysis; however, species or breed differences may exist., (Published by Elsevier Inc.)

The aim of this study was to test for the postulated luteotropic effect of prostaglandin E2 during early diestrus in the dog in an in vivo study. This study was performed on 30 bitches which were randomly assigned to a treatment group (TG) and a control group. Starting on the day of ovulation (Day 0), dogs of the TG were treated for 5, 10, 20, or 30 days with 10 mg firocoxib/kg body weight per day (Previcox, a selective PTGS2 inhibitor) and ovariohysterectomized for collection of corpora lutea on the last day of treatment. Similarly, dogs of the control group were ovariohysterectomized on Days 0, 5, 10, 20, and 30. Blood samples for progesterone measurement were collected every second day; additionally, the area of luteal cell nuclei and the expression of 3β-hydroxysteroid-dehydrogenase at the mRNA and the protein levels were assessed. Mean P4 concentrations were lower in TGs; however, a significant difference was only observed on Day 10. This observation is in line with the finding that treatment with firocoxib reduced expression of 3β-hydroxysteroid-dehydrogenase mRNA and protein (P < 0.05) and the area of luteal cell nuclei (P < 0.05). The results of this study further point to the postulated luteotropic function of prostaglandin E2., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The Iberian lynx (Lynx pardinus) is the most critically endangered felid. A high reproductive success within the Iberian Lynx Conservation Breeding Program is crucial to maintaining the goal of reintroducing captive born offspring to the wild and thus increasing the population. Lynx follow a unique reproductive strategy with a monoestrous cycle and persisting CLs over many years. These persistent CLs constantly produce progesterone (on average 5 ng/mL) and are hypothesized to hinder a polyestrous cyclicity in lynx. The aim of this study was to evaluate whether artificial luteolysis can be achieved with common luteolytic drugs and if luteolysis would induce a second estrus naturally. We observed a functional regression of lynx CLs after artificial luteolysis with 2.5 μg/kg body weight PGF2α analogue (cloprostenol) administered three times every 16 hours. We could see a similar effect when combining cloprostenol with other drugs like an anti-gestagen (aglepristone) or a dopamin-agonist (prolactin-inhibitor, cabergolin) or by prolonging the cloprostenol administration to a total of 5 days. However, the sample size was too small to draw conclusions about which protocol is superior or if combining different drugs would result in a positive synergism. Neither structural regression of CLs nor subsequent spontaneous estrus induction was induced with any of these treatments. We suggest that a dose of 2.5 μg/kg body weight cloprostenol administered once daily over 3 to 5 days is sufficient for functional luteolysis in lynx. The next step would be to compare the success of estrus induction with or without the preceding artificial luteolysis., (Copyright © 2014 Elsevier Inc. All rights reserved.)

It has been reported that early development of wave 3 in three-wave interovulatory intervals occurs during the luteolytic period, and the frequency of the ipsilateral relationship between the preovulatory follicle and CL is lower (e.g., 33%) than for the contralateral relationship (67%). In this study, luteolysis was induced with PGF2α when the largest follicle of wave 2 reached 8.5 mm or more (diameter at expected deviation). A two-way interaction (P < 0.004) of follicle (first follicle to reach 8.5 mm and the next-largest follicle) by group (ipsilateral and contralateral relationship between the 8.5-mm follicle and CL) represented smaller posttreatment diameter difference between the two follicles within the ipsilateral group than within the contralateral group; the 8.5-mm follicle was smaller and the next-largest follicle was larger in the ipsilateral than in the contralateral group. Switching in the destiny of the 8.5-mm follicle from dominant to subordinate occurred in 3 of 8 (41%) and 0 of 5 (0%) heifers in the ipsilateral and contralateral groups, respectively. These novel findings supported the hypothesis that follicle deviation during luteolysis may result in decreased diameter of the largest follicle and increased frequency of switching of the largest follicle from future dominant to subordinate status when the follicle and CL are ipsilateral. Support for the switching hypothesis indicated that the reported lower frequency of an ipsilateral than contralateral relationship between the preovulatory follicle and CL for wave 3 of three-wave interovulatory intervals can be attributed to development of the wave during luteolysis., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The aim of this study was to investigate the effect of dichlorodiphenyltrichloroethane(DDT), dichlorodiphenyldichloroethylene (DDE) and γ-hexachlorocyclohexane (HCH) (10 ng/ml) on myometrial motility and the secretory function of the myometrium and corpus luteum (CL) collected from cows on days 8-12 of the estrous cycle. All of the xenobiotics increased (P<0.05) myometrial contractility. Moreover, the xenobiotics stimulated the secretion of the following prostaglandins (PGs) from myometrial strips: PGF2α, PGE2 and PGI2. DDT and DDE also increased (P<0.05) the release of PGF2α from CL strips, and HCH had the same effect (P<0.05) on the secretion of PGE2 and PGI2. The studied xenobiotics did not affect (P>0.05) PG synthesis, but DDT and DDE increased the mRNA expression levels of leukemia inhibitor factor (LIF), which can stimulate PG production. In summary, the xenobiotics affected PG secretion from cow myometrium and CL, which may contribute to the mechanism of uterine contraction disturbance., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The hypothesis that ovulation-inducing factor/nerve growth factor (OIF/NGF) isolated from llama seminal plasma exerts a luteotrophic effect was tested by examining changes in circulating concentrations of LH and progesterone, and the vascular perfusion of the ovulatory follicle and developing CL. Female llamas with a growing follicle of 8 mm or greater in diameter were assigned randomly to one of three groups (n = 10 llamas per group) and given a single intramuscular dose of PBS (1 mL), GnRH (50 μg), or purified OIF/NGF (1.0 mg). Cineloops of ultrasonographic images of the ovary containing the dominant follicle were recorded in brightness and power Doppler modalities. Llamas were examined every 4 hours from the day of treatment (Day 0) until ovulation, and every other day thereafter to Day 16. Still frames were extracted from cineloops for computer-assisted analysis of the vascular area of the preovulatory follicle from treatment to ovulation and of the growing and regressing phases of subsequent CL development. Blood samples were collected for the measurement of plasma LH and progesterone concentrations. The diameter of the dominant follicle at the time of treatment did not differ among groups (P = 0.48). No ovulations were detected in the PBS group but were detected in all llamas given GnRH or OIF/NGF (0/10, 10/10, and 10/10, respectively; P < 0.0001). No difference was detected between the GnRH and OIF/NGF groups in the interval from treatment to ovulation (32.0 ± 1.9 and 30.4 ± 5.7 hours, respectively; P = 0.41) or in maximum CL diameter (13.1 ± 0.4 and 13.5 ± 0.3 mm, respectively; P = 0.44). The preovulatory follicle of llamas treated with OIF/NGF had a greater vascular area at 4 hours after treatment than that of the GnRH group (P < 0.001). Similarly, the luteal tissue of llamas treated with purified OIF/NGF had a greater vascular area than that of the GnRH group on Day 6 after treatment (P < 0.001). The preovulatory surge in plasma LH concentration began, and peaked 1 to 2 hours later in the OIF/NGF group than in the GnRH group (P < 0.05). Plasma progesterone concentration was higher on Day 6 in the OIF/NGF group than in the GnRH group (P < 0.001). Results support the hypothesis that OIF/NGF exerts a luteotrophic effect by altering the secretion pattern of LH and enhancing tissue vascularization during the periovulatory period and early stages of CL development., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The objective of this study was to determine the effects of timely injections of flunixin meglumine (FM) or vaginal application of prostaglandin E2 (PgE2) on pregnancy, fertility, fecundity, and prolificacy rates in Saanen goats. One hundred and sixty-three nonlactating Saanen does were treated with a flugestone acetate (20 mg)-containing intravaginal sponge for 12 days. They also received eCG (400 IU) and a PGF2α analogue (50 μg) 10 days after progestagen priming. Does detected in estrus were mated and assigned randomly to one of three treatment groups. The PgE2 group (N = 40) received PgE2 (2.5 mg) intravaginally 15 days after mating. The FM group (N = 54) received flunixin meglumine (total dose, 100 mg) intramuscularly 15 days after mating. Flunixin meglumine was administered at 9:00 AM. Animals in the control group (N = 69) received no treatment. Pregnancy was diagnosed using transrectal ultrasonography (B-mode at 8 MHz) 30 days after mating. The pregnancy rate was significantly greater (P < 0.01) after 30 days in goats treated with PgE2 and also in the control group than in those treated with FM (67.5%, 59.4%, and 42.5%, respectively). The pregnancy rate did not differ between the PgE2 and the control group. The pregnancy and fertility rate were lowest in the FM group compared with the other groups. There was no significant difference in the prolificacy rate among experimental groups. In conclusion, our results showed that FM administration during a late luteal phase is detrimental to early pregnancy in goats., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The early corpus luteum (CL) (before Day 6) does not regress after a single PGF2α treatment. We hypothesized that increasing PGF2α dose or number of treatments would allow regression of the early CL (Day 5). Nonlactating Holstein cows (N = 22) were synchronized using the Ovsynch protocol. On Day 5 (Day 0 = second GnRH treatment), cows were assigned to: (1) control (N = 5): no further treatment; (2) 1PGF (N = 6): one dose of 25 mg PGF2α; (3) 2PGF (N = 5): two doses of 25 mg PGF2α (50 mg) given 8 hours apart (second PGF2α on Day 5 at the same time as the other PGF2α treatments); (4) DPGF (N = 6): double dose of 25 mg PGF2α (50 mg) given on Day 5. Blood samples were collected to monitor progesterone (P4) profiles in two periods. In the first period (0 to 24 hours), there were effects of treatment (P = 0.01), time (P < 0.01), and an interaction of treatment and time (P = 0.02). Group 1PGF versus control was different only at 12 hours (P = 0.02). Cows treated with DPGF were different than control at 4 hours (P = 0.04), 12 hours (P < 0.01), and 24 hours (P < 0.01). Only cows treated with 2PGF had lower P4 than control during the entire period and low P4 (0.37 ± 0.17 ng/mL) at 24 hours, usually indicative of luteolysis. In the second period (Day 5 to 15 of the cycle), there were effects of treatment (P < 0.01), time (P < 0.01), and interaction of treatment and time (P = 0.002). Group 1PGF was not different than control from Day 5 to 13 and P4 was greater than control on Day 14 (P = 0.01) and 15 (P < 0.01). Circulating P4 in DPGF cows was lower than control from Day 7 (P = 0.05) through 12 (P < 0.01). Likewise, there were differences between control and 2PGF from Day 7 to 13, but not on Day 14 and 15. On Day 15, all PGF2α-treated groups had circulating P4 consistent with an active CL. Ultrasound evaluation confirmed that no CL from any group completely regressed during the experiment and no new ovulations occurred to account for functional CL later in cycle. In summary, a double dose of PGF2α (twice on Day 5 or 8 hours apart) can dramatically decrease P4, consistent with classical definitions of luteolysis; however, these CL recover and become fully functional. Thus, the Day 5 CL of mature Holstein cows do not regress even to two doses of PGF2α., (Copyright © 2014. Published by Elsevier Inc.)

To study the ovarian response to the long-term effect of PGF2α, 16 cows were treated with 25 mg tromethamine dinoprost (Pronalgon F; Pfizer, Tokyo, Japan) for 21 days after natural ovulation. Five control cows were treated with sterile physiological saline. The follicle and corpus luteum (CL) development were monitored using a real-time ultrasound instrument. In addition, the plasma concentration of progesterone (P4) was determined. In nine of the 16 Pronalgon-treated cows, the first dominant follicle (1st DF), second dominant follicle (2nd DF), and third dominant follicle ovulated consecutively (group A). In five cows, the 1st and 2nd DFs ovulated consecutively (group B). The developing CL started to regress approximately 5 days after each ovulation without maturation in groups A and B. In the two remaining Pronalgon-treated cows, there was no further ovulation after natural ovulation (group C). In one cow in group C, the 1st DF became atretic and the 2nd DF became cystic with the diameter of the cystic follicle reaching 31.2 mm on Day 30. In another cow, the 1st DF became cystic with a diameter of 30.9 mm on Day 18. Although P4 began to increase after each ovulation in all of the Pronalgon-treated cows, it decreased immediately after each ovulation without a large increase, peaking at approximately 1 ng/mL. Furthermore, the number of days when P4 was >1 ng/mL from natural ovulation to Day 21 was 2.6 ± 0.7 days, which was significantly less than that in the control cows (16.0 ± 0.6 days). These results indicate that the long-term effect of PGF2α has an important role in ovulation of all dominant follicles and might induce cystic ovaries in cows., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Recent experiments using expression, immunolocalization, and cell culture approaches have provided leading insights into regulation of luteal angiogenesis by different growth factor systems and its role in the function of corpus luteum (CL) in buffalo. On the contrary, lymphangiogenesis and its regulation in the CL are still poorly understood. The aim of this study was to evaluate the expression and localization of lymphangiogenic factors (vascular endothelial growth factor [VEGF]-C and VEGFD), their receptor (VEGFR3), and lymphatic endothelial marker (LYVE1) in bubaline CL during different stages of the estrous cycle and to investigate functional role of VEGFC and VEGFD in luteal lymphangeogenesis. The mRNA and protein expression of VEGFC, VEGFD, and VEGFR3 was significantly greater in mid and late luteal phases, which correlated well with the expression of LYVE1. The lymphangiogenic factors were localized in luteal cells, exclusively in the cytoplasm. Immunoreactivity of VEGFC was greater during midluteal phase and that of VEGFD was greater during the mid and late luteal phases. Luteal cells were cultured in vitro and treated for different time duration (24, 48, and 72 hours) with VEGFC and VEGFD each at 50, 100, and 150 ng/mL concentration and VEGFC with VEGFD at 100 ng/mL concentration. The temporal increase in LYVE1 mRNA expression was significant (P < 0.05) in VEGFC and VEGFC with VEGFD treatment and no significant change was seen in VEGFD treatment. Thus, it seems likely that VEGFD itself has little role in lymphangiogenesis but along with VEGFC it might have a synergistic effect on VEGFR3 receptors for inducing lymphangiogenesis. In summary, the present study provided evidence that VEGFC and VEGFD, and their receptor VEGFR3, are expressed in bubaline CL and are localized exclusively in the cell cytoplasm, suggesting that these factors have a functional role in lymphangiogenesis of CL in buffalo., (Copyright © 2014 Elsevier Inc. All rights reserved.)

In this study we compared the effect of dinoprost and cloprostenol on changes of corpus luteum blood flow during luteolysis. Ten nonlactating cyclic ewes were synchronized with double PGF2α injections 11 days apart. At Day 10, the animals were classified into 2 groups and received the third dose of PGF2α after confirmation of the presence of a mature CL. The first group received (12.5 mg/im) dinoprost and the second group received (250 μg/im) cloprostenol. A color Doppler ultrasound scan was performed by the same operator according to the following timeline: 0, 0.5, 1, 2, 4, 6, 12, and 24 hours, then every 24 hours until Day 4). The size, morphology, and blood flow of the CL was evaluated during the regression. The results showed that regression of the CL did not differ between the dinoprost and cloprostenol groups. There was no significant effect on diameter of the CL in both groups, though the size of the CL decreased gradually and slowly. Pretreatment progesterone concentration did not differ between groups. The results showed that the nitric oxide level was significantly increased within half an hour after the dinoprost treatment, and was significantly decreased in the cloprostenol group after half an hour. The blood velocity was increased significantly half an hour after the dinoprost treatment and it was decreased in the cloprostenol-treated group. In conclusion, both cloprostenol and dinoprost affect CL by controlling the nitric oxide level and blood supply of the CL via different mechanisms to induce luteolysis., (Copyright © 2013 Elsevier Inc. All rights reserved.)

The objective was to determine the effects of eCG given on the day of, or 2 days before removal of an intravaginal progestin device, on ovarian follicle diameter, luteal volume, serum progesterone (P4) concentrations, and pregnancy per insemination in a fixed-time AI (FTAI) protocol. Lactating, anestrous, multiparous Bos taurus cross beef cows, 40 to 60 days postpartum, were given estradiol benzoate (2 mg im) and a progestin intravaginal device containing 250 mg of medroxyprogesterone acetate on Day 0 and cloprostenol (0.265 mg) on Day 6. Intravaginal devices were removed on Day 8 and GnRH (100 μg im) was given on Day 9, with timed AI 16 hours later. In experiment 1, cows were randomly assigned to receive 400 IU im eCG on Day 6 (eCG6; N = 8) or Day 8 (eCG8; N = 8), or to not receive eCG (control; N = 8). Dominant follicle diameter on Day 9 in the eCG6 group (10.0 ± 0.5 mm) was larger (P < 0.05) than in the eCG8 (8.6 ± 0.2 mm) or control (8.5 ± 0.4 mm) groups. Corpora lutea (CL) in all cows in the control group underwent premature luteolysis within 10 days after ovulation. Luteal volumes and P4 concentrations 10 and 15 days after ovulation were higher (P < 0.05) in the eCG6 group than in the eCG8 group. In experiment 2, the eCG6 (N = 121) and eCG8 (N = 125) protocols were compared in lactating anestrous cows that underwent FTAI. Pregnancy rate was higher (P < 0.05) in the cows that received eCG on Day 6 (27.3%; 33/121) than on Day 8 (16.0%; 20/125). Furthermore, CL volumes and P4 concentrations were higher (P < 0.05) in the eCG6 group (5784.0 ± 857.3 mm(3) and 8.1 ± 1.3 ng/mL, respectively) than in the eCG8 group (3220.9 ± 505.1 mm(3) and 4.5 ± 0.7 ng/mL, respectively). We concluded that eCG given 2 days before progestin removal in this FTAI protocol for anestrous beef cows increased diameter of the dominant follicle, luteal volume, serum P4 concentrations, and pregnancy rates., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Exogenous eCG for stimulation of a single dominant follicle or for superovulation are common strategies to improve reproductive efficiency by increasing pregnancy rates and embryo production, respectively. Morphofunctional changes in the CL of eCG-treated cattle include increases in CL volume and plasma progesterone concentrations. Therefore, we tested the hypothesis that eCG alters the content of luteal cells and mitochondria related to hormone production. Twelve crossbred beef cows were synchronized and then allocated into three groups (four cows per group) and received no further treatment (control) or were given eCG either before or after follicular deviation (superovulation and stimulation of the dominant follicle, respectively). Six days after ovulation, cows were slaughtered and CL collected for morphohistologic and ultrastructural analysis. Mitochondrial volume per CL was highest in superovulated followed by stimulated and then control cows (18,500 ± 2630, 12,300 ± 2640, and 7670 ± 3400 μm(3); P < 0.001), and the density of spherical mitochondria and the total number of large luteal cells were increased (P < 0.05) in stimulated cows compared with the other two groups (110.32 ± 14.22, 72.26 ± 8.77, and 70.46 ± 9.58 mitochondria per μm(3) and 678 ± 147, 245 ± 199, and 346 ± 38 × 10(6) cells, respectively. However, the largest diameters of the large luteal cells were increased in superovulated and control cows versus stimulated ones (32.32 ± 0.06, 31.59 ± 0.81, and 29.44 ± 0.77 μm; P < 0.0001). In contrast, the total number of small luteal cells was increased in superovulated cows (1456 ± 268, 492 ± 181, and 822 ± 461 × 10(6), P < 0.05). In conclusion, there were indications of cellular changes related to increased hormonal production (stimulatory treatment) and increased CL volume (superovulatory treatment)., (Copyright © 2013 Elsevier Inc. All rights reserved.)

The objectives were to: (1) evaluate the efficacy of varying intervals of oxytocin administration in preventing luteolysis in mares; (2) examine PGF(2α) release in mares experiencing prolonged diestrus secondary to oxytocin treatment; and (3) evaluate the endometrial expression of oxytocin receptor, estrogen receptor α, and prostaglandin synthesis enzymes after oxytocin administration. In experiment I, mares received oxytocin (60 IU, im) daily on Days 8 to 10, 8 to 12, or 8 to 14 postovulation, and control mares received sterile saline. Prolongation of diestrus was defined by elevation of serum progesterone >1.0 ng/mL through Day 30 postovulation. The proportion of mares experiencing prolonged cycles increased (P < 0.01) as the number of days of oxytocin administration increased. Oxytocin administration on Days 8 to 10, 8 to 12, and 8 to 14 prolonged luteal maintenance in 3/7, 4/7, and 6/7 mares respectively, compared with 0/7 control mares. Treated mares with prolonged diestrus had lower (P < 0.05) plasma PGFM concentrations at Day 16 than did mares with normal diestrus periods. In experiment II, endometrial biopsies from mares treated with oxytocin from Days 8 to 14 postovulation (N = 6) had reduced cyclooxygenase-2 expression (P < 0.05) compared with control mares (N = 6) as determined by quantitative reverse transcription polymerase chain reaction and immunohistochemical staining. Oxytocin administration prolonged luteal maintenance in mares, with an increasing number of mares responding to treatment as the number of days of oxytocin administration was increased beyond Day 8 postovulation. Luteal maintenance in mares was also associated with decreased plasma PGFM concentrations and reduced endometrial cyclooxygenase-2 expression., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Concerns regarding the mechanisms of action of emergency contraception (EC) create major barriers to widespread use and could also lead to incorrect use of EC and overestimation of its effectiveness. While the copper intrauterine device (Cu-IUD) is the most effective method available for EC, the hormonal methods are frequently considered to be more convenient and acceptable. Today, the most commonly used method for hormonal EC is levonorgestrel (LNG). More recently, the progesterone receptor modulator ulipristal acetate (UPA) has been shown to be more effective than LNG to prevent an unwanted pregnancy. The main mechanism of action of both LNG and UPA for EC is delaying or inhibiting ovulation. However, UPA appears to have a direct inhibitory effect on follicular rupture which allows it to be effective even when administered shortly before ovulation, a time period when use of LNG is no longer effective. The main mechanism of action of the Cu-IUD is to prevent fertilization through the effect of Cu ions on sperm function. In addition, if fertilization has already occurred, Cu ions influence the female reproductive tract and prevent endometrial receptivity. Based on this review of the published literature, it can be concluded that existing methods used today for EC act mainly through inhibition of ovulation or prevention of fertilization. An additional effect on the endometrium as occurs for the Cu-IUD, but not for the hormonal alternatives, seems to increase the efficacy of the method., (Copyright © 2013 Elsevier Inc. All rights reserved.)