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The aim in this study was to investigate corpus luteum function and embryonic loss in buffaloes mated by artificial inseminations (AI) during the transitional period from breeding to non-breeding season. The study was carried out using 288 multiparous Italian Mediterranean Buffalo cows at 110 ± 4 days in milk. The buffaloes were mated by AI after synchronization of ovulation by the Ovsynch-TAI protocol 25 days after AI buffaloes underwent trans-rectal ultrasonography to assess embryonic development. Pregnancy diagnosis was confirmed on Days 45 and 70 after AI by rectal palpation. Buffaloes pregnant on Day 25 but not on Day 45 were considered to have undergone late embryonic mortality (LEM), whilst buffaloes pregnant on Day 45 but not on Day 70 were considered to have undergone foetal mortality (FM). Corpus luteum size and blood flow were determined by real-time B-mode/colour-Doppler on day 10 after AI in 122 buffaloes. The resistive index (RI) and pulsatility index (PI) were recorded at the time. Milk samples were collected on Days 10, 20 and 25 after AI in all inseminated buffaloes for the assay of whey P4 concentrations. Data were analysed by . Pregnancy rate on Day 25 after AI was 48.6% (140/288) and declined to 35.4% (102/288) and 30.6% (88/288) by Day 45 and Day 70 respectively. The incidences of LEM and FM were respectively 27.1% (38/140) and 13.7% (14/102). Pregnant buffaloes had greater (p < 0.01) whey concentrations of P4 from Day 20 onwards than buffaloes which showed LEM, whilst P4 in buffaloes that showed FM did not differ from the other two groups on Day 10 and Day 20. Corpus luteum blood flow on Day 10 after AI showed higher RI (p < 0.05) and PI (p = 0.07) values in buffaloes that subsequently were not pregnant on Day 25 compared with pregnant buffaloes. Buffaloes that were not pregnant on Day 45 also had a higher (p = 0.02) RI value on Day 10 than pregnant buffaloes, whilst PI values on Day 10 did not differ for the two groups of buffaloes. It was concluded that blood flow to the corpus luteum on Day 10 after AI influences corpus luteum function as judged by P4 secretion and also embryonic development and attachment in buffaloes. [ABSTRACT FROM AUTHOR]

Most genetic variants associated with fertility in mammals fall in non-coding regions of the genome and it is unclear how these variants affect fertility. Here we use genome-wide association summary statistics for Heifer puberty (pubertal or not at 600 days) from 27,707 Bos indicus, Bos taurus and crossbred cattle; multi-trait GWAS signals from 2119 indicine cattle for four fertility traits, including days to calving, age at first calving, pregnancy status, and foetus age in weeks (assessed by rectal palpation of the foetus); and expression quantitative trait locus for whole blood from 489 indicine cattle, to identify 87 putatively functional genes affecting cattle fertility. Our analysis reveals a significant overlap between the set of cattle and previously reported human fertility-related genes, impling the existence of a shared pool of genes that regulate fertility in mammals. These findings are crucial for developing approaches to improve fertility in cattle and potentially other mammals., (© 2024. The Author(s).)

The pleomorphic adenoma gene1 (PLAG1) encodes a DNA-binding, C2H2 zinc-finger protein which acts as a transcription factor that regulates the expression of diverse genes across different organs and tissues; hence, the name pleomorphic. Rearrangements of the PLAG1 gene, and/or overexpression, are associated with benign tumors and cancers in a variety of tissues. This is best described for pleomorphic adenoma of the salivary glands in humans. The most notable expression of PLAG1 occurs during embryonic and fetal development, with lesser expression after birth. Evidence has accumulated of a role for PLAG1 protein in normal early embryonic development and placentation in mammals. PLAG1 protein influences the expression of the ike growth factor 2 (IGF2) gene and production of IGF2 protein. IGF2 is an important mitogen in ovarian follicles/oocytes, embryos, and fetuses. The PLAG1-IGF2 axis, therefore, provides one pathway whereby PLAG1 protein can influence embryonic survival and pregnancy. PLAG1 also influences over 1,000 other genes in embryos including those associated with ribosomal assembly and proteins. Brahman (Bos indicus) heifers homozygous for the PLAG1 variant, rs109815800 (G > T), show greater fertility than contemporary heifers with either one, or no copy, of the variant. Greater fertility in heifers homozygous for rs109815800 could be the result of early puberty and/or greater embryonic survival. The present review first looks at the broader roles of the PLAG1 gene and PLAG1 protein and then focuses on the emerging role of PLAG1/PLAG1 in embryonic development and pregnancy. A deeper understanding of factors which influence embryonic development is required for the next transformational increase in embryonic survival and successful pregnancy for both in vivo and in vitro derived embryos in cattle., (© The Author(s) 2024. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Background: Several studies show that natural foods are a source of compounds with anticancer properties that affect the gut microbiota and its metabolites. In the present study, we investigate the effect of a delactosed buffalo milk whey by-product (DMW) on colorectal carcinogenesis., Methods: The effect of DMW on colorectal carcinoma (CRC) was investigated in the established mouse model of azoxymethane (AOM)-induced colon carcinoma, which closely resembles the human clinical condition of CRC. The effect of DMW on CRC immortalized cell lines was also evaluated to further identify the antineoplastic mechanism of action., Results: Pretreatment of AOM-treated mice with DMW significantly (P < 0.05) reduced the percentage of mice bearing both aberrant crypt foci with more than four crypts (which are early precancerous lesions that progress to CRC) and tumors. In addition, DMW completely counteracted the effect of AOM on protein expression of caspase-9, cleaved caspase-3 and poly ADP-ribose polymerase in colonic tissue. Administration of DMW alone (i.e. without AOM) resulted in changes in the composition of the gut microbiota, leading to enrichment or depletion of genera associated with health and disease, respectively. DMW was also able to restore AOM-induced changes in specific genera of the gut microbiota. Specifically, DMW reduced the genera Atopobiaceae, Ruminococcus 1 and Lachnospiraceae XPB1014 and increased the genera Parabacteroides and Candidatus Saccharimonas, which were increased and reduced, respectively, by AOM. Blood levels of butyric acid and cancer diagnostic markers (5-methylcytidine and glycerophosphocholine), which were increased by AOM treatment, were reduced by DMW. Furthermore, DMW exerted cytotoxic effects on two human CRC cell lines (HCT116 and HT29) and these effects were associated with the induction of apoptotic signaling., Conclusions: Our results suggest that DMW exerts chemopreventive effects and restores the gut microbiota in AOM-induced CRC, and induces cytotoxic effect on CRC cells. DMW could be an important dietary supplement to support a healthy gut microbiota and reduce the prevalence of CRC in humans. Video Abstract., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Testicular ultrasonography and steroid concentrations (cortisol, dehydroepiandrosterone sulfate (DHEA-S), cortisol/DHEA-S ratio, testosterone) in hair were examined for their utility in the bull breeding soundness evaluation (BBSE). Beef and dairy bulls ( n = 16; 2.7 ± 0.4 years old; body condition score 3.2 ± 0.1) of five breeds were maintained under the same conditions at an accredited semen collection center. Bulls underwent routine semen collection twice weekly for 12 weeks and semen was processed and cryopreserved. Ultrasonography and hair sampling were undertaken at the last semen collection. Bulls with homogeneous testicular parenchyma ( n = 8) had a higher ( p < 0.05) percentage of motile sperm post-thawing compared with bulls with heterogeneous parenchyma ( n = 8). There were no differences ( p > 0.05) in the hair concentrations of cortisol, DHEA-S, and testosterone between bulls with homogeneous and heterogeneous parenchyma. In bulls with homogeneous parenchyma, hair DHEA-S concentration was positively correlated with percentage motile sperm (R 2 = 0.76), progressively motile sperm (R 2 = 0.70), and motility yield (R 2 = 0.71). The findings indicate that the integration of testicular ultrasonography and hair DHEA-S status in the BBSE could provide a more comprehensive assessment of indicative fertility in bulls. Additionally, ultrasonography can be used in the BBSE when the evaluation of semen parameters is not available.

This study evaluated the effects of supplementing with natural functional feed on the plasma fatty acid profile of lactating Italian Holstein-Friesian dairy cows. Thirty cows in mid-lactation received the natural olive extract PHENOFEED DRY (500 mg/cow/day) which mainly comprises hydroxytyrosol, tyrosol and verbascoside. The total content of polyphenols and the antioxidant power of standard feed, enriched feed and pure extract was evaluated respectively by Folin-Ciocalteu and DPPH assay, and a characterization in HPLC-UV (High-Performance Liquid Chromatography-Ultraviolet) of bioactive molecules present in the extract PHENOFEED DRY was performed. PHENOFEED DRY was provided for 60 days, and the plasma profile of fatty acids was determined by Gas Chromatography. The administration of enriched feed resulted in an increase in the ratio of Omega-6 to Omega-3 polyunsaturated fatty acids from 3:1 to 4:1 (p<0.001). This was not influenced by the calving order. The addition of polyphenols helped to keep monounsaturated (MUFA) and saturated (SFA) levels constant and results in a significant increase in polyunsaturated (PUFA) fatty acid after 15 days of administration. The Omega-6/Omega-3 ratio was in the optimal range. The findings show that inclusion of natural functional food such as plant polyphenols helps to maintain a healthy blood fatty acid profile in lactating dairy cows., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors. Published by Elsevier Ltd.)

1 H NMR-based metabolomics was used to study the effect of abrupt weaning on the blood metabolome of beef calves. Twenty Angus calves (258 ± 5 kg BW; 5 to 6 months old) were randomly assigned to a non-weaned (NW) group that remained grazing with their dam or a weaned (W) group that underwent abrupt separation from their dam to a separate paddock on d 0 of the study. Body weight, behaviour, and blood samples for cortisol and metabolomics were measured at d 0, 1, 2, 7, and 14 of the study. On d 1 and 2, W calves spent less time grazing and ruminating, and more time vocalising and walking, had a greater concentration of cortisol, NEFA, 3-hydroxybutyrate, betaine, creatine, and phenylalanine, and lesser abundance of tyrosine (P < 0.05) compared to NW calves. Compared to NW calves at d 14, W calves had greater (P < 0.01) relative abundance of acetate, glucose, allantoin, creatinine, creatine, creatine phosphate, glutamate, 3-hydroxybutyrate, 3-hydroxyisobutyrate, and seven AA (alanine, glutamate, leucine, lysine, phenylalanine, threonine and valine) but lesser (P < 0.05) relative abundance of low density and very low-density lipids, and unsaturated lipids. Both PCA and OPLS-DA showed no clustering or discrimination between groups at d 0 and increasing divergence to d 14. Blood metabolomics is a useful tool to quantify the acute effects of stress in calves during the first 2 days after abrupt weaning, and longer-term changes in carbohydrate, lipid and protein metabolism due to nutritional changes from cessation of milk intake and greater reliance on forage intake., (© 2023. The Author(s).)

The domestic buffalo (Bubalus bubalis), also known as water buffalo, comprises two sub-species the River buffalo (B. bubalis ssp. bubalis; 50 chromosomes) and the Swamp buffalo (ssp. carabanensis; 48 chromosomes). Domestic buffaloes are a globally significant livestock species. In South Asia, the River buffalo is a primary source of milk and meat and has a very important role in food security. The River buffalo also supports high-value, differentiated food production in Europe and the Americas. The Swamp buffalo is an important draft animal and a source of food in Southeast Asia and East Asia. The growing importance of buffaloes requires that they undergo an accelerated rate of genetic gain for efficiency of production, product quality, and sustainability. This will involve the increased use of assisted reproduction. The initial application of reproductive technology in buffaloes had variable success as it relied on the adoption of procedures developed for cattle. This included artificial insemination (AI), sperm cryopreservation, and embryo technologies such as cloning and in vitro embryo production (IVEP). Reproductive technology has been progressively refined in buffaloes, and today, the success of AI and IVEP is comparable to cattle. Ovarian follicular superstimulation (superovulation) combined with in vivo embryo production results in low embryo recovery in buffaloes and has limited practical application. The contribution of elite female buffaloes to future genetic improvement will therefore rely mainly on oocyte pickup and IVEP. This will include IVEP from females before puberty to reduce generation intervals. This review provides for the first time a clear chronology on the development, adoption, and impact, of assisted reproduction in domestic buffaloes., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Genomic selection combined with in vitro embryo production (IVEP) with oocytes from heifer calves provides a powerful technology platform to reduce generation interval and significantly increase the rate of genetic gain in cattle. The ability to obtain oocytes with developmental competence from calves has been recognised for more than 60years. There is renewed interest in the potential of this reproductive technology as the beef and dairy industries seek to identify and multiply animals that satisfy consumer demand for efficient utilisation of natural resources, with minimal environmental impact and high product quality. Heifer calves show a good follicular response to FSH stimulation and oocytes can be recovered by ovum pick-up (OPU). However, the efficiency of OPU/IVEP remains lower for calves compared with peripubertal heifers and cows, in both indicus (Zebu, Bos indicus ) and taurus (Bos taurus ) breeds. Indicus cattle generally have more follicles, yield a greater number of oocytes, and have a better IVEP outcome, compared with taurus cattle. The treatment of prepubertal heifers with bovine somatotrophin (bST) and propylene glycol before OPU/IVEP has yet to show a major improvement in embryo production. Holstein (taurus) dairy heifers derived from different reproductive technologies (AI, MOET, OPU/IVEP) showed a similar age at puberty and first-lactation milk production. OPU/IVEP Holstein embryos transferred to beef or dairy cows likewise yielded heifers with the same performance. The gains in productivity that can be achieved with strategic use of OPU/IVEP in heifer calves make this a relevant and highly important reproductive technology in cattle breeding. Ongoing optimisation of the technology is needed for the potential of OPU/IVEP in young donors to be fully realised.

The present study was conducted to examine whether pretreatment with melatonin would enhance ovarian follicular functions and increase response to estrous synchronization and fixed-time AI (TAI) during the nonbreeding season in lactating dairy buffalo. In Experiment 1, buffalo cows without a detectable corpus luteum (CL) were assigned on Day -20 (D-20) to three groups: control (n = 12); melatonin (n = 13); progesterone (P 4 ) (n = 15). Cows in the melatonin group were implanted with melatonin on D-20. From D0 to D9, there was imposing of an estrous synchronization treatment regimen using either a standard Ovsynch protocol (control, melatonin) or a P 4 -based Ovsynch treatment regimen (P 4 ). There were no differences (P > 0.05) among groups for the presence of a CL at D0, size of the largest follicle at D0, ovulation to GnRH injection at D0 and D9, or the time to ovulation after injection of GnRH at D9. In Experiment 2, there was imposing of the same treatment regimens as in Experiment 1, with inclusion of TAI. Females of the P 4 group had a greater (P = 0.001) pregnancy/AI percentage (60 %) than those in the control (17 %) and melatonin (23 %) groups. Females of the P 4 group also had a larger (P = 0.005) CL at D20 compared with those in the control and melatonin groups. Findings indicate treatment with melatonin for 20 days did not affect ovarian functions or the response to an estrous synchronization treatment regimen and TAI during the nonbreeding season in lactating dairy buffalo., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Contraceptive vaccines are designed to elicit immune responses against major components of animal reproductive systems. These vaccines, which are most commonly administered via injection, typically target gonadotropin-releasing hormone (GnRH). However, the need to restrain animals for treatment limits the field applications of injectable vaccines. Oral administration would broaden vaccine applicability. We explored contraceptive vaccine candidates composed of GnRH peptide hormone, universal T helper PADRE (P), and a poly(methylacrylate) (PMA)-based delivery system. When self-assembled into nanoparticles, PMA-P-GnRH induced the production of high IgG titers after subcutaneous and oral administration in mice. PADRE was then replaced with pig T helper derived from the swine flu virus, and the vaccine was tested in pigs. High levels of systemic antibodies were produced in pigs after both injection and oral administration of the vaccine. In conclusion, we developed a simple peptide-polymer conjugate that shows promise as an effective, adjuvant-free, oral GnRH-based contraceptive vaccine.

Early embryo development, implantation and pregnancy involve a complex dialogue between the embryo and mother. In cattle this dialogue starts as early as days 3-4 when the embryo is still in the oviduct, and it continues to implantation. Immunological processes involving cytokines, mast cells and macrophages form an important part of this dialogue. Amongst the cytokines, interleukin-6 (Il-6) and leukemia inhibitory factor (LIF) are secreted by both the embryo and uterine endometrium and form part of an ongoing and reciprocating dialogue. Mast cells and macrophages populate the uterine endometrium during embryo development and are involved in achieving the correct balance between inflammatory and anti-inflammatory reactions at the uterus that are associated with embryo attachment and implantation. Embryo loss is the major cause of reproductive wastage in cattle, and livestock generally. A deeper understanding of immunological processes during early embryo development will help to achieve the next step change in the efficiency of natural and assisted breeding., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The effect of green feed on health-promoting biomolecules in milk was examined in dairy buffaloes. Buffaloes received a total mixed ration (TMR) (Control, C; n = 40) or TMR + alfalfa green feed (30% of diet) (Treated, T; n = 40). Biomolecules and functional activity were measured in milk obtained twice-monthly. Treated buffaloes had higher milk l-carnitine, acetyl-l-carnitine, propionyl-l-carnitine and δ-valerobetaine (P < 0.01). They also had higher antioxidant activity (P < 0.01). Compared with C buffaloes, milk of T buffaloes improved the viability of endothelial cells exposed to high-glucose (P < 0.01), and reduced intracellular lipid peroxidation, reactive oxygen species (ROS), and cytokine release (P < 0.01). Milk of T buffaloes inhibited with greater potency the viability of human HCT116 and Cal 27 cancer cells (P < 0.001). The findings show that including green feed in the diet of dairy buffaloes enhances health-promoting biomolecules and the antioxidant and antineoplastic properties of milk., (Copyright © 2020. Published by Elsevier Ltd.)

The present study examined whether priming with estradiol benzoate (EB) for 12 h increased both the peak and duration of LH release in response to kisspeptin (KISS1, KP) in cows. In a Latin square design, ovariectomized Nelore cows ( n = 8) received: Control, i.m. 4 mL of 0.9% saline; KP, i.m. 4 mg murine KISS1-10; EBKP, i.m. 4 mg KISS1-10 + i.m. 2 mg EB simultaneously; EB12KP, i.m. 4 mg KISS1-10 + i.m. 2 mg EB 12 h before KISS1-10. Concentrations of LH were determined in blood samples obtained at time 0 (treatment), 20, 40, 60, 90, 120, 150, 180, 210 and 270 min. Concentrations of LH were analyzed by Proc GLIMMIX for repeated measures. In case of significance, the adjusted Tukey test was used to test for differences among treatments. GraphPad 8.0 PRISM ® was used to determine the area under the LH-response curve (AUC) after injection of KISS1-10. Plasma LH remained relatively constant throughout sampling after treatment with saline. The peak in LH after injection of KISS1-10 occurred at 20 min in Groups KP and EBKP and at 40 min in Group EB12KP. The peak LH response (∆LH, ng/mL) was greater ( p < 0.01) in Group EB12KP (5.6 ± 0.9) than in Groups KP (2.4 ± 0.9) and EBKP (3.5 ± 0.9), which did not differ. AUC (LH ng/mL*min) was greater ( p = 0.02) in Group EB12KP (439 ± 73) than in Groups KP (176 ± 73) and EBKP (241 ± 73), with the latter two groups not differing. The findings indicated that 12 h priming with EB increased both the peak and duration of the LH response to treatment with KISS1. The incorporation of EB priming and KISS1 could improve the efficiency of estrus synchronization with fixed-time AI in cows. This would have an important practical application in assisted breeding in beef and dairy cattle.

Recently, several concerns have been expressed on red meat quality and consumption. The aims of this study were to evaluate the influence of different breeding techniques and a prolonged post dry aging (PDA) maturation process on biomolecules level in raw buffalo meat. In the first experiment, two groups of animals were maintained with different space availability (15 vs. 10 m 2 /animal) for 90 days and biomolecules content was evaluated. In experiment 2, two diets (with or without ryegrass green forage) were used to assess the concentration of these biomolecules. Finally, in experiment 3, the meat of the animals that showed the highest content of biomolecules was chosen to assess the influence of the PDA maturation process. Buffaloes reared at 15 m 2 showed a significantly ( p < 0.05) higher content of the considered biomolecules compared with their counterparts. Similarly, buffaloes fed green forage showed higher content of biomolecules ( p < 0.05) compared with the control group. The meat of the animals bred at 15 m 2 and fed green forage showed a significant ( p < 0.01) increase of biomolecules content during the PDA maturation process up to 60 days without influence microbiological profile in terms of total aerobic bacterial counts, yeasts, and molds. In conclusion, breeding techniques and PDA maturation system could enhance biomolecules levels in terms of quality, without affect health standards.

Kisspeptin (KISS1) is encoded by the KISS1 gene and was initially found to be a repressor of metastasis. Natural mutations in the KISS1 receptor gene (KISS1R) were subsequently shown to be associated with idiopathic hypothalamic hypogonadism and impaired puberty. This led to interest in the role of KISS1 in reproduction. It was established that KISS1 had a fundamental role in the control of gonadotropin releasing hormone (GnRH) secretion. KISS1 neurons have receptors for leptin and estrogen receptor α (ERα), which places KISS1 at the gateway of metabolic (leptin) and gonadal (ERα) regulation of GnRH secretion. More recently, KISS1 has been shown to act at peripheral reproductive tissues. KISS1 and KISS1R genes are expressed in follicles (granulosa, theca, oocyte), trophoblast, and uterus. KISS1 and KISS1R proteins are found in the same tissues. KISS1 appears to have autocrine and paracrine actions in follicle and oocyte maturation, trophoblast development, and implantation and placentation. In some studies, KISS1 was beneficial to in vitro oocyte maturation and blastocyst development. The next phase of KISS1 research will explore potential benefits on embryo survival and pregnancy. This will likely involve longer-term KISS1 treatments during proestrus, early embryo development, trophoblast attachment, and implantation and pregnancy. A deeper understanding of the direct action of KISS1 at reproductive tissues could help to achieve the next step change in embryo survival and improvement in the efficiency of assisted reproductive technology., (© The Author(s) 2020. Published by Oxford University Press on behalf of Society for the Study of Reproduction.)

Wagyu crossbred steers (n = 167) were used to (1) compare the metabolome of individual animals at two distant time-points (days 196 and 432) in a feedlot (this corresponded to 272 and 36 days before slaughter); and (2) determine relationships between the metabolome and marbling, and the effect of days in the feedlot (time-points) on these relationships. 1 H NMR spectroscopy followed by standard recoupling of variables analysis produced 290 features or 'peaks' from which 38 metabolites were identified. There was a positive correlation between the relative concentration (RC) at days 196 and 432 for 35 of 38 metabolites (P > 0.05). The RC of 21 metabolites mostly involved in muscle energy and glucose metabolism increased (P < 0.05) from day 196 to 432, and the RC of 13 metabolites mostly involved in lipid metabolism decreased (P < 0.05). There were 14 metabolites correlated with marbling including metabolites involved in energy and fat metabolism (glucose, propionate, 3-hydroxybutyrate, lipids). The relationship between marbling and the RC of metabolites was affected by time-point, being positive for 3-hydroxybutyrate and acetate (P < 0.05) at day 432 but not at day 196. The findings indicate that the blood metabolome in Wagyu crossbred steers changes with time in a feedlot. Notwithstanding, the metabolome has potential to predict marbling in Wagyu. The ability to predict marbling from the blood metabolome appears to be influenced by days in a feedlot and presumably the stage of development towards a mature body conformation.

The aim of this study was to determine if treatment with estradiol cypionate (EC) at the time of P4 withdrawal induced ovulation in a synchronization/timed-AI (TAI) protocol in buffalo. In Experiment 1, 56 buffaloes received an intravaginal P4 device (1.0 g) plus estradiol benzoate (EB, 2.0 mg im) on Day 0 (D0). On Day 9, the P4 device was removed and buffaloes were given PGF 2α (0.53 mg im sodium cloprostenol) plus eCG (400 IU im). Buffaloes were then randomly allocated to one of two groups: Group GEC (n = 29), treated with EC (1.0 mg im) at P4 device removal; Group GEB (n = 27), treated with EB (1.0 mg im) 24 h after P4 device removal. Ovarian ultrasound was undertaken on: D0, to ascertain general ovarian status; D9 to D11 (every 24 h), to measure diameter of the largest follicle (LF) and follicular growth rate; D11 to D13 (every 12 h for 72 h), to determine the time of ovulation and ovulation rate. Following P4 device removal, Groups GEC and GEB had a similar follicular growth rate (0.9 ± 0.1 and 1.1 ± 0.1 mm/day, respectively; P = 0.15) and similar LF diameter on D11 (11.4 ± 0.6 and 12.5 ± 0.5 mm; P = 0.12). Groups GEC and GEB also had a similar diameter of the ovulatory follicle (13.0 ± 0.5 and 13.4 ± 0.6 mm; P = 0.52), interval from P4 device removal to ovulation (68.2 ± 2.8 and 71.1 ± 1.4 h; P = 0.41) and ovulation rate (62.1% and 70.4%; P = 0.44). In Experiment 2, 199 buffaloes were assigned to the two treatments in Experiment 1 (GEC, n = 100; GEB, n = 99). All animals underwent TAI 56 h after P4 device removal and pregnancy diagnosis was preformed on D41. The pregnancy rate was similar for Groups GEC and GEB (50.0 and 45.5%, respectively; P = 0.45). The findings indicate that treatment with EC at the time of P4 withdrawal induces ovulation and achieves the same pregnancy rate to TAI as treatment with EB 24 h after P4 removal. The use of EC requires one less handling which is highly important in facilitating practical adoption of TAI in assisted breeding and genetic improvement in buffalo., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Seasonal breeding in buffalo is influenced by exogenous (photoperiod, climate, nutrition, management) and endogenous (hormones, genotype) factors. Buffalo are negatively photoperiodic and show a natural increase in fertility during decreasing day length. The hormone melatonin is produced by the pineal gland and has a fundamental role in photoperiodic time measurement within the brain. This drives annual cycles of gonadotropin secretion and gonadal function in buffaloes. Some melatonin is released into the systemic circulation and, together with peripherally produced melatonin, acts at somatic tissues. In the ovaries and testes of buffalo, melatonin acts as an antioxidant and scavenges oxygen free radicals to reduce both oxidative stress and apoptosis. This has beneficial effects on gametogenesis and steroidogenesis. Female buffalo treated with melatonin show an improved response to estrus synchronization protocols in out-of-season breeding. Melatonin acts through melatonin receptors MT 1 and MT 2 and the gene for MT 1 (MTNR1A) is polymorphic in buffaloes. Single nucleotide polymorphisms (SNPs) in gene MTNR1A have been associated with fertility in female buffalo. The knowledge and tools are available to lift the reproductive performance of buffalo. This is highly important as the global demand for nutritious buffalo food products has undergone a sharp rise, and continues to grow. Buffalo can make an important contribution to affordable, nutritious animal protein. This will help address global nutritional security., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Artificial insemination (AI) is important for genetic improvement and to control the period of breeding in buffalo and has increased significantly over the past 20 years. AI is more difficult in buffalo compared with cattle due to variable estrous cycles, reduced estrous behavior, and reproductive seasonality. The latter is associated with a higher incidence of anestrus and increased embryonic mortality during the nonbreeding season. Protocols to control the stage of the estrous cycle have undergone recent development in buffalo. These protocols are based on the control of both the luteal phase of the cycle, mainly by prostaglandins and progesterone, and follicle development and ovulation, by prostaglandins, progesterone, GnRH, hCG, eCG and estradiol. Protocols that synchronize the time of ovulation enable fixed timed AI, avoiding estrous detection. Factors to consider when selecting an AI protocol include animal category (heifers, primiparous or pluriparous), reproductive status (cyclic or anestrus), and season. This review looks at the current status of estrus synchronization and AI in buffalo and provides some practical suggestions for application of AI in the field., (Crown Copyright © 2020. Published by Elsevier Inc. All rights reserved.)

The aim of the present study was to determine the recovery of embryonic structures (ova/embryos) and fertilization rate in superovulated buffaloes treated with PGF 2α during the periovulatory period. On day 0 (D0), buffaloes at random stages of the estrous cycle were treated with an intravaginal progesterone device (P4; 1.0 g) and estradiol benzoate (EB, 2.0 mg i.m.). From D4 to D7, all buffaloes received i.m. FSH (200 mg total) twice-daily over 4 days in decreasing doses. On D6 and D7, the animals were given PGF 2α analogue (0.53 mg i.m. sodium cloprostenol) and the P4 device was removed on D7. On D8, all buffaloes received GnRH (20 μg i.m. buserelin acetate). Buffaloes were then randomly allocated to one of three groups: control (Group C, n = 18), no further treatment; PGF 2α analogue injection (Group IM-PGF; n = 18), four injections (0.53 mg i.m. sodium cloprostenol) 12 h apart, from D8 to D10; PGF 2α analogue osmotic pump (Group OP-PGF; n = 18), s.c. osmotic mini-pump (2.12 mg sodium cloprostenol) from D8 to D10. The study had a crossover design (three treatments x three replicates). All animals underwent timed AI, 12 and 24 h after treatment with GnRH. Embryonic structures were recovered on D14. Ovarian ultrasonography was used on D8 and D14 to record follicular superstimulation and superovulatory responses. Blood samples were obtained on Days 7, 8, 9 and 10 to measure circulating concentrations of P4, E2 and PGFM. Data were analyzed by GLIMMIX procedure of SAS®. There was no effect (P = 0.58) of treatment on the total number of embryonic structures (Group C, 2.1 ± 0.8; Group IM-PGF, 2.1 ± 0.6; Group OP-PGF, 1.4 ± 0.4). There was also no effect (P = 0.93) of treatment on the recovery rate of embryonic structures (oocytes and embryos D14/CL D14). The fertilization rate was higher (P = 0.04) in Groups IM-PGF (84.6%) and OP-PGF (88.0%), which did not differ, than Group C (63.2%). The viable embryos rate was greater (P < 0.01) for Groups IM-PGF (82.0%) and OP-PGF (88.0%) than Group C (52.6%). There was no interaction between treatment and time and treatment effects for P4, E2 and PGFM concentrations. The findings showed that treatment with PGF 2α during the periovulatory period has potential to increase fertilization rate and embryo production in superovulated buffaloes., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Survival of the embryo and establishment of a pregnancy is a critical period in the reproductive function of female cattle. This review examines how the transforming growth factor-β (TGFB) superfamily (i.e. bone morphogenetic protein (BMP) 15, growth differentiation factor (GDF) 9, anti-Müllerian hormone (AMH)) and interferon-τ (IFNT) affect ovarian function and embryo development. The oocyte in a primary follicle secretes BMP15 and GDF9, which, together, organise the surrounding granulosa and theca cells into the oocyte-cumulus-follicle complex. At the same time, the granulosa secretes AMH, which affects the oocyte. This autocrine-paracrine dialogue between the oocyte and somatic cells continues throughout follicle development and is fundamental in establishing the fertilisation potential and embryo developmental competency of oocytes. The early bovine embryo secretes IFNT, which acts at the uterine endometrium, corpus luteum and blood leucocytes. IFNT is involved in the maternal recognition of pregnancy and immunomodulation to prevent rejection of the embryo, and supports progesterone secretion. Manipulation of BMP15, GDF9, AMH and IFNT in both invivo and invitro studies has confirmed their importance in reproductive function in female cattle. This review makes the case that a deeper understanding of the biology of BMP15, GDF9, AMH and IFNT will lead to new strategies to increase embryo survival and improve fertility in cattle. The enhancement of oocyte quality, early embryo development and implantation is considered necessary for the next step change in the efficiency of natural and assisted reproduction in cattle.

Cell-cell adhesion molecules have critically important roles in the early events of reproduction including gamete transport, sperm-oocyte interaction, embryonic development, and implantation. Major adhesion molecules involved in reproduction include cadherins, integrins, and disintegrin and metalloprotease domain-containing (ADAM) proteins. ADAMs on the surface of sperm adhere to integrins on the oocyte in the initial stages of sperm-oocyte interaction and fusion. Cadherins act in early embryos to organize the inner cell mass and trophectoderm. The trophoblast and uterine endometrial epithelium variously express cadherins, integrins, trophinin, and selectin, which achieve apposition and attachment between the elongating conceptus and uterine epithelium before implantation. An overview of the major cell-cell adhesion molecules is presented and this is followed by examples of how adhesion molecules help shape early reproductive events. The argument is made that a deeper understanding of adhesion molecules and reproduction will inform new strategies that improve embryo survival and increase the efficiency of natural mating and assisted breeding in cattle., (© 2020 Wiley-Liss, Inc., A Wiley Company.)

Current diagnosis methods for Bovine Respiratory Disease (BRD) in feedlots have a low diagnostic accuracy. The current study aimed to search for blood biomarkers of BRD using 1 H NMR metabolomics and determine their accuracy in diagnosing BRD. Animals with visual signs of BRD (n = 149) and visually healthy (non-BRD; n = 148) were sampled for blood metabolomics analysis. Lung lesions indicative of BRD were scored at slaughter. Non-targeted 1 H NMR metabolomics was used to develop predictive algorithms for disease classification using classification and regression trees. In the absence of a gold standard for BRD diagnosis, six reference diagnosis methods were used to define an animal as BRD or non-BRD. Sensitivity (Se) and specificity (Sp) were used to estimate diagnostic accuracy (Acc). Blood metabolomics demonstrated a high accuracy at diagnosing BRD when using visual signs of BRD (Acc = 0.85), however was less accurate at diagnosing BRD using rectal temperature (Acc = 0.65), lung auscultation score (Acc = 0.61) and lung lesions at slaughter as reference diagnosis methods (Acc = 0.71). Phenylalanine, lactate, hydroxybutyrate, tyrosine, citrate and leucine were identified as metabolites of importance in classifying animals as BRD or non-BRD. The blood metabolome classified BRD and non-BRD animals with high accuracy and shows potential for use as a BRD diagnosis tool.

The aim of the present study was to determine if space allocation influenced the concentration of biomolecules in buffalo milk and dairy products. Intensively housed buffaloes (n = 96) were randomly assigned to 2 groups according to days in milk, parity, and milk yield: group S10 had a space allocation of 10 m 2 per buffalo and group S15 had a space allocation of 15 m 2 per buffalo. Individual milk yield was recorded daily. Twice a month, a bulk milk sample was collected for each group, as well as whey, ricotta, and mozzarella cheese, to assess cheese yield and to conduct HPLC-electrospray ionization-tandem mass spectrometry, milk antioxidant activity, and cell viability analyses. We tested milk extracts from the 2 groups in vitro to evaluate their efficacy in counteracting endothelial oxidative damage induced by high glucose. We evaluated reproductive function in 28 buffaloes from each group using the Ovsynch-timed artificial insemination program. We observed no differences in milk quantity or quality in terms of fat, protein, or lactose, and reproductive function did not differ between the 2 groups. Compared with group S10, group S15 had higher concentrations of carnitine (56.7 ± 1.1 vs. 39.8 ± 0.7 mg/L in milk and 40.9 ± 0.8 vs. 31.7 ± 0.7 mg/L in whey), acetyl-l-carnitine (51.9 ± 0.3 vs. 39.7 ± 0.7 mg/L in milk and 41.1 ± 1.7 vs. 28.7 ± 2.6 mg/L in whey), propionyl-l-carnitine (34.8 ± 1.0 vs. 21.0 ± 0.9 mg/L in milk and 26.9 ± 0.8 vs. 17.6 ± 1.2 mg/L in whey), glycine betaine (23.1 ± 1.9 vs. 13.5 ± 1.6 mg/L in milk and 10.7 ± 0.4 vs. 7.9 ± 0.5 mg/L in whey), and δ-valerobetaine (24.2 ± 0.5 vs. 16.7 ± 0.5 mg/L in milk and 22.0 ± 0.9 vs. 15.5 ± 0.7 mg/L in whey). Group S15 also had higher total antioxidant activity than group S10 (56.7 ± 1.9 vs. 46.4 ± 1.13 mM Trolox equivalents). Co-incubation of high-glucose-treated endothelial cells with milk extracts from group S15 improved cell viability compared with cells treated with high glucose only; it also reduced intracellular lipid peroxidation (144.3 ± 0.4 vs. 177.5 ± 1.9%), reactive oxygen species (141.3 ± 0.9 vs. 189.3 ± 4.7 optical density units), and cytokine release (tumor necrosis factor-α, IL-1β, IL-6). Greater space allocation was associated with higher levels of biomolecules in buffalo milk. This could have been the result of improved welfare in buffaloes that were allocated more space., (Copyright © 2019 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Nutrition is of fundamental importance in reproductive function of female beef cattle. Nutrition determines live weight (LW) and body condition score (BCs) and both were found more than 50 years ago to underpin fertility in pubertal heifers and postpartum cows. In heifers, LW at weaning and average daily gain (ADG) after weaning determines age and LW at puberty, and subsequent lifetime fertility. In cows, BCS at parturition is the most important factor that determines the period to re-conception postpartum. Nutrition establishes systemic metabolic homeostasis. Metabolic hormones such as leptin, IGF1 and Ghrelin act as signaling factors that regulate activity of GnRH neurons in the hypothalamus. The release of GnRH and function of the reproductive endocrine system is determined by the balance of positive signals (IGF1, leptin) and negative signals (Ghrelin) at GnRH neurons. Nutritional factors also directly influence ovarian follicles, oocytes and embryos. Saturated fatty acids (FAs) are detrimental to oocytes and embryos whilst unsaturated FAs may be beneficial. The ratio of FAs (saturated, monounsaturated, polyunsaturated) is likely the key to optimal reproductive function. Nutrition controls the levels of metabolic hormones (leptin, IGF1, Ghrelin) and metabolic factors (FAs) and both have major roles in reproduction in female beef cattle., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The aim of this study was to compare estradiol benzoate (EB) and GnRH for the induction of ovulation in a TAI protocol in buffalo during the nonbreeding season. In experiment 1, 141 buffaloes (56 cows and 85 heifers) received an intravaginal P4 device (1.0 g) plus EB (2.0-mg, intramuscular [im]) at random stage of the estrous cycle (Day 0). On Day 9, the P4 device was removed, and buffaloes were given PGF 2α (0.53-mg im sodium cloprostenol) plus eCG (400-IU im). Buffaloes were then randomly allocated to one of three groups and treated as follows: EB24 (n = 47), EB (1.0 mg im) 24 hours after P4 device removal; EB36 (n = 50), EB 36 hours after P4 device removal; GnRH48 (n = 44), GnRH (10 μg im buserelin acetate) 48 hours after P4 device removal. Ultrasound examinations were performed on Day 0 to ascertain ovarian follicular status, Day 9 to measure follicular diameter, and from Day 11 to Day 14 (every 12 hours for 60 hours) to establish the time of ovulation. There were no significant differences between EB24, EB36, and GnRH48 for diameter of the ovulatory follicle (13.1 ± 0.3, 13.7 ± 0.3, and 13.7 ± 0.3 mm; P = 0.26) and ovulation rate (78.7%, 82.0%, and 84.1%; P = 0.93). When compared with heifers, cows had a greater diameter of the dominant follicle on Day 9 (10.3 ± 0.3 and 8.6 ± 0.2 mm; P = 0.0001), diameter of the ovulatory follicle (14.1 ± 0.3 and 13.1 ± 0.2 mm; P = 0.01), ovulation rate (91.1% and 75.3%; P = 0.02), and interval from P4 device removal to ovulation (76.3 ± 1.3 and 72.5 ± 1.4 hours; P = 0.05). In experiment 2, 511 buffaloes (354 cows and 157 heifers) were assigned to the same treatments described in experiment 1 (EB24, n = 168; EB36, n = 172; and GnRH48, n = 171), and all animals were submitted to timed artificial insemination (TAI) 64 hours after P4 device removal. Pregnancy diagnosis was undertaken 30 days after TAI. There were no significant differences between EB24, EB36, and GnRH48 for pregnancy rate (45.2%, 43.0%, and 49.7%; P = 0.46), and the pregnancy rate did not differ (P = 0.31) for cows (47.5%) and heifers (42.7%). The findings from the two experiments indicated that EB (24 or 36 hours) and GnRH (48 hours) induce comparable follicular responses, ovulation, and pregnancy rates in buffalo cows and heifers. Although there were some differences in the follicular responses between cows and heifers, the pregnancy rate to TAI was nonetheless similar., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The aim of this study was to evaluate the relationship between the ovarian follicular response at the start of the OVsynch-fixed-time artificial insemination (TAI) program and pregnancy outcome in the Mediterranean river buffalo. The study involved 116 pluriparous buffaloes at 118 ± 65 days in milk maintained under commercial conditions on a buffalo dairy enterprise in southern Italy. Synchronization of the stage of the estrous cycle in the Ovsynch-TAI program involves an injection of GnRH on Day 0 (GnRH1), PGF 2α on Day 7, and GnRH on Day 9 (GnRH2). Timed artificial insemination is carried out on Day 10. Ovarian ultrasound examinations were undertaken on days 0, 2, 7, 9, 10, and 11 to record total follicles, the dimensions of follicles greater than 0.75 cm, dimensions of the corpus luteum (CL), and the occurrence of ovulation. Blood concentrations of P4 were measured on days 0 and 10 and 10 days after TAI. Pregnancy status was determined on days 27 and 45 after TAI to calculate late embryonic mortality. The data were analyzed by ANOVA and multiple logistic regression. A total of 70 of 116 (60.4%) buffaloes showed ovulation after GnRH1. Buffaloes that ovulated after GnRH1 had a smaller (P < 0.05) area of the ovulatory follicle on Day 0 compared with buffaloes that did not ovulate (1.08 ± 0.1 and 1.29 ± 0.1 cm 2 , respectively). The former buffaloes also had a larger (P < 0.05) CL area (1.81 ± 0.1 and 1.51 ± 0.1 cm 2 , respectively) and greater (P < 0.05) blood concentrations of P4 (1.84 ± 0.2 and 1.34 ± 0.2 ng/mL, respectively) on Day 0. A larger (P < 0.05) proportion of buffaloes that ovulated after GnRH1 showed regression of the CL after the injection of PGF 2α on Day 7 compared with buffaloes that did not ovulate (68 of 69 [98.6%] and 27 of 32 [84.4%], respectively). Buffaloes that ovulated after GnRH1 also had a greater (P < 0.05) ovulation rate after GnRH2 than buffaloes that did not ovulate (65 of 70 [92.8%] and 37 of 46 [80.4%], respectively). Buffaloes that ovulated after GnRH1 had a higher (P < 0.01) pregnancy rate than buffaloes that did not ovulate, both at Day 27 (53 of 70 [75.7%] and 14 of 46 [30.4%], respectively) and Day 45 (46 of 70 [65.7%] and 10 of 46 [21.7%], respectively). The incidence of late embryonic mortality tended to be less for buffaloes that ovulated after GnRH1 than buffaloes that did not ovulate (7 of 53 [13.2%] and 4 of 14 [28.6%], respectively). The present findings have demonstrated a strong direct relationship between the ovarian follicular response at the start of an OVsynch-TAI program and pregnancy outcome in the Mediterranean river buffalo., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The present study investigated the expression and localization of angiopoietin (ANPT) family members in buffalo ovarian follicles of different size. It also looked at the role of ANPTs in estradiol secretion and mRNA expression of phosphoinositide-3-kinase-protein kinase B signaling pathway cellular proliferation (phosphoinositide-dependant kinase and protein kinase B [AKT]) and proapoptotic (BAD) factors with caspase 3 in cultured buffalo granulosa cells (GCs). The mRNA and protein expression of ANPT-1 was greatest (P < 0.05), whereas ANPT-2 was reduced (P < 0.05) in preovulatory follicles as compared to F1 follicle. Tyrosine kinase with immunoglobulin-like and EGF-like domains 1 transcripts and protein expression did not change in all follicular groups, whereas tyrosine kinase with immunoglobulin-like and EGF-like domains 2 mRNA was highest (P < 0.05) in theca interna but not GC layer of preovulatory follicle. All members of ANPT family were localized in GC and theca interna showing a stage specific immunoreactivity. Cultured GCs were treated with ANPT-1 and ANPT-2 separately at doses of 1, 10, and 100 ng/mL and in combination at 100 ng/mL for three incubation periods (24, 48, and 72 hours). Estradiol secretion was highest (P < 0.05) at 100 ng/mL at 72 hours of incubation when GCs were treated with either protein alone. The mRNA expression of phosphoinositide-dependant kinase and AKT was highest (P < 0.05), and BAD with caspase 3 was lowest (P < 0.05) at 100 ng/mL at 72 hours of incubation, when cultured GCs were treated separately with each protein or in combination. The immuoreactivity of AKT, pAKT, and pBAD were maximal, whereas BAD was minimal with 100 ng/mL at 72 hours when cultured GCs treated with either protein alone. The findings indicate that ANPTs are expressed in a regulated manner in buffalo ovarian follicle during different stages of development where they may promote steroidogenesis and GC survival through autocrine and paracrine actions., (Copyright © 2016 Elsevier Inc. All rights reserved.)

River buffalo are able to adapt to diverse climatic zones and are widespread globally. The resource use efficiency of buffalo is highly relevant in a resource-constrained world and the increasing requirement to produce more food. Buffalo clearly have an important role in meeting the growing demand for animal protein. In the Mediterranean and higher latitudes, buffalo show annual cycles of ovarian activity, embryonic development, and pregnancy rate. In buffalo, the CL starts to develop early in the cycle, and there is also an early increase in concentrations of progesterone (P4) in circulation. This appears to be necessary for optimal embryonic development. The failure to establish a pregnancy in buffalo can occur before Day 21 (early embryonic mortality), from Day 21 to 45 (late embryonic mortality), and from Day 46 to 90 (fetal mortality) after mating. Treatment with P4, hCG, and GnRH on Day 5 after mating increases P4 in circulation and reduces early embryonic mortality in circumstances where concentrations of P4 are relatively low. The same treatments applied on Day 20 to 25 after mating can lower the occurrence of late embryonic mortality and fetal mortality., (Copyright © 2016 Elsevier Inc. All rights reserved.)

High-producing dairy cows enter a period of negative energy balance during the first weeks of lactation. Energy intake is usually sufficient to cover the increase in energy requirements for fetal growth during the period before calving, but meeting the demand for energy is often difficult during the early stages of lactation. A catabolic state predominates during the transition period, leading to the mobilisation of energy reserves (NEFA and amino acids) that are utilised mainly by the liver and muscle. Increased uptake of mobilised NEFA by the liver, combined with the limited capacity of hepatocytes to either oxidise fatty acids for energy or to incorporate esterified fatty acids into VLDL results in fatty liver syndrome and ketosis. This metabolic disturbance can affect the general health, and it causes economic losses. Different nutritional strategies have been used to restrict negative effects associated with the energy challenge in transition cows. The provision of choline in the form of rumen-protected choline (RPC) can potentially improve liver function by increasing VLDL exportation from the liver. RPC increases gene expression of microsomal TAG transfer protein and APOB100 that are required for VLDL synthesis and secretion. Studies with RPC have looked at gene expression, metabolic hormones, metabolite profiles, milk production and postpartum reproduction. A reduction in liver fat and enhanced milk production has been observed with RPC supplementation. However, the effects of RPC on health and reproduction are equivocal, which could reflect the lack of sufficient dose-response studies.

The aim of the present study was to evaluate small follicle number, oocyte yield, and in vitro embryo production (IVEP) in Zebu beef cows treated long term with a GnRH agonist to suppress ovarian follicular growth. Nelore (Bos indicus) cows (n = 20) showing regular estrous cycles were randomly assigned to one of two groups: control (n = 10, placebo ear implant without a GnRH agonist); GnRH agonist (n = 10, GnRH agonist ear implant containing 9.4-mg deslorelin). All cows underwent an ovum pick-up (OPU) session 14 days (Day 14) before the start of treatments (Day 0) followed by seven OPU-IVEP procedures at 30-day intervals (Days 0, 30, 60, 90, 120, 150, and 180). Semen from a single batch of a previously tested bull was used for all the IVEP. Cows treated with agonist reported a decrease over time in the proportion of animals with a (CL; P ≤ 0.05) and large follicles (>10 mm, P ≤ 0.05). These cows had a lesser number of medium + large follicles (>5 mm; 1.74 ± 0.5 vs. 4.13 ± 0.5; P ≤ 0.05), greater number of small follicles (2-5 mm; 44.3 ± 2.8 vs. 30.8 ± 1.8; P ≤ 0.05), greater yield of cumulus-oocyte complexes (COCs; 21.0 ± 2.3 vs. 15.6 ± 1.9; P ≤ 0.05), greater proportion of COCs cultured (79.2 vs. 73.9%; P ≤ 0.05), COCs cleaved (10.6 ± 1.5 vs. 6.8 ± 1.1, P ≤ 0.05), and cleaved rate (52.8 vs. 44.3%; P ≤ 0.05) compared with control cows. The number (3.4 ± 0.7 vs. 3.0 ± 0.6; P > 0.05) and proportion (16.5 vs. 19.1%; P > 0.05) of blastocysts produced were similar between agonist and control cows, respectively. The study has shown that Zebu beef cows treated long term with a GnRH agonist had follicular growth restricted to small follicles. This did not compromise the ability of oocytes to undergo IVF and embryonic development., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The immunoneutralization of gonadotropin-releasing hormone (GnRH) can be used for the treatment of human hormone-dependent male and female cancers or as immunocontraceptives in animals. Vaccine candidates 1 [Th(K-LP)GnRH], 2 [GnRH(K-LP)Th], 3 [GnRH(K-Th)LP], and 4 [Th(K-GnRH)LP] (for which K=lysine, LP=lipopeptide Ser-Ser-C16 -C16 , and Th=T helper cell epitope influenza HA2), were synthesized by assembling a CD4(+) T helper cell epitope (Th), GnRH, and an adjuvanting lipid moiety (LP) in various spatial arrangements. All compounds were efficiently taken up by antigen-presenting cells with significant immunogenicity without an external adjuvant. Compounds 2, 3, and 4, in which GnRH is conjugated through its C terminus, produced higher GnRH-specific antibody responses than construct 1, in which the GnRH moiety is conjugated through its N terminus. All four constructs induced a significant antiproliferative effect (up to 55 %) on GnRH-receptor-positive LNCaP cells, but showed weaker activity in the GnRH-receptor-negative SKOV-3 cell line. Marked degenerative changes were observed in morphology and follicular development in the ovaries of immunized mice, with approximately 30 % higher degenerative antral and atretic follicles., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)