Establishment Of Pregnancy In Cattle Research Articles

Loci and pathways associated with uterine capacity for pregnancy and fertility in beef cattle.

Infertility and subfertility negatively impact the economics and reproductive performance of cattle. Of note, significant pregnancy loss occurs in cattle during the first month of pregnancy, yet little is known about the genetic loci influencing pregnancy success and loss in cattle. To identify quantitative trait loci (QTL) with large effects associated with early pregnancy loss, Angus crossbred heifers were classified based on day 28 pregnancy outcomes to serial embryo transfer. A genome wide association analysis (GWAA) was conducted comparing 30 high fertility heifers with 100% success in establishing pregnancy to 55 subfertile heifers with 25% or less success. A gene set enrichment analysis SNP (GSEA-SNP) was performed to identify gene sets and leading edge genes influencing pregnancy loss. The GWAA identified 22 QTL (p < 1 x 10−5), and GSEA-SNP identified 9 gene sets (normalized enrichment score > 3.0) with 253 leading edge genes. Network analysis identified TNF (tumor necrosis factor), estrogen, and TP53 (tumor protein 53) as the top of 671 upstream regulators (p < 0.001), whereas the SOX2 (SRY [sex determining region Y]-box 2) and OCT4 (octamer-binding transcription factor 4) complex was the top master regulator out of 773 master regulators associated with fertility (p < 0.001). Identification of QTL and genes in pathways that improve early pregnancy success provides critical information for genomic selection to increase fertility in cattle. The identified genes and regulators also provide insight into the complex biological mechanisms underlying pregnancy establishment in cattle.

Basic and practical aspects of pregnancy establishment in cattle

Basic and practical aspects of pregnancy establishment in cattle

Embryo maternal immune interactions in cattle

Mammalian embryo implantation requires the priming of the maternal immune system, but, not the provocation. There are many examples of conditions where a disturbed or aberrant immune profile during embryo implantation leads to pregnancy loss. However, these studies are primarily associated with human and mouse species; data is generally limited for cattle and livestock. Most available information centres on the endometrial response to interferon tau (IFNT), a type I antiviral cytokine, which is the maternal recognition factor for cattle and sheep. Interferon tau secretion by the embryo and detection by the dam is critical to corpus luteum (CL) maintenance and pregnancy retention. However, the large volume of bovine endometrial and conceptual transcriptomic data highlights a broader more integral role of the maternal immune system in the establishment of pregnancy in cattle. When taken together with available immunohistochemistry and flow cytometry data from livestock, mouse, and human, a profile of immune cell involvement from ovulation to conception and placentation emerges. The key events of pregnancy establishment in cattle and the involvement of the maternal immune system will be discussed.

Effect of preovulatory follicle maturity on pregnancy establishment in cattle: the role ofoocyte competence and the maternal environment

Effect of preovulatory follicle maturity on pregnancy establishment in cattle: the role ofoocyte competence and the maternal environment

The contribution of the maternal immune system to the establishment of pregnancy in cattle.

Immune cells play an integral role in affecting successful reproductive function. Indeed, disturbed or aberrant immune function has been identified as primary mechanisms behind infertility. In contrast to the extensive body of literature that exists for human and mouse, studies detailing the immunological interaction between the embryo and the maternal endometrium are quite few in cattle. Nevertheless, by reviewing the existing studies and extrapolating from sheep, pig, mouse, and human data, we can draw a reasonably comprehensive picture. Key contributions of immune cell populations include granulocyte involvement in follicle differentiation and gamete transfer, monocyte invasion of the peri-ovulatory follicle and their subsequent role in corpus luteum formation and the pivotal roles of maternal macrophage and dendritic cells in key steps of the establishment of pregnancy, particularly, the maternal immune response to the embryo. These contributions are reviewed in detail below and key findings are discussed.

193 PROGESTERONE CONCENTRATIONS AND MESSENGER RNA EXPRESSION OF PROGESTERONE RECEPTORS IN ENDOMETRIAL TISSUE OF BOVINE UTERINE HORN IPSILATERAL AND CONTRALATERAL TO CORPUS LUTEUM

Generally, conception is established in the uterine horn ipsilateral to the corpus luteum (CL) in cattle. When a bovine embryo is transferred into the uterine horn contralateral to CL, conception rate is low. Since progesterone (P4) is essential for the establishment of pregnancy in cattle, locational effects of P4 released from CL at the uterus may cause the differences in fertility. The aim of this study was to determine the endometrial tissue P4 concentrations (EndP4) and the mRNA expression of nuclear progesterone receptor (PGR), and progesterone receptor component-1 (PGRMC-1) and -2 (PGRMC-2) in the endometrial tissues from the ipsi- and contralateral horn. The uteruses of Holstein cows were obtained at a local abattoir. Endometrial tissues were collected from both horns. Based on ovarian morphology, the oestrus cycle of the cow was estimated as follows: early luteal phase (ELP, Day 5–6, Day 0 = oestrus), mid luteal phase (MLP, Day 8–12), late luteal phase (LLP, Day 15–17), and follicular phase (FP, Day 18–20). EndP4 was measured by enzyme immunoassay. Expressions of mRNA were analysed by real-time RT–PCR. Two-way factorial ANOVA and the Steel-Dwass test were applied for a multiple comparison of means. The interrelations between both parameters were expressed by Spearman correlation coefficient. At ELP and MLP, EndP4 in ipsi-horn were higher than that in contra-horn (P &lt; 0.05, see Table 1). Higher mRNA expression of PGRMC-1 in ipsi-horn was observed at ELP compared with contra-horn (P &lt; 0.05). Expressions of mRNA for PGR and PGRMC-2 were similar in both horns. In ipsi-horn at ELP, EndP4 was positively correlated with PGRMC-1 mRNA (r = 0.87, P &lt; 0.05), but was negatively correlated with PGR mRNA (r = –0.76, P &lt; 0.05). However, in contra-horn, EndP4 has no correlation to mRNA expression of P4 receptors. In conclusion, EndP4 was influenced by the location of CL and stage of oestrus cycle. Higher expression of PGRMC-1 mRNA in endometrial tissue of ipsi-horn at ELP might be up-regulated by higher EndP4. These locational effects of CL on uterus may provide an intrauterine environment suitable for embryo development. Table 1.End P4 and expression of mRNA for P4 receptors in bovine uterine horn1

107 PRODUCTION OF EPIDERMAL GROWTH FACTOR BY BOVINE ENDOMETRIAL CELLS AND ITS EFFECTS ON EMBRYONIC INTERFERONT AND PROSTAGLANDIN

A dynamic interaction between bioactive products of the embryo (blastocyst) and the endometrium is crucial for the successful establishment of pregnancy. In ruminants, the principal signal for maternal recognition of pregnancy is interferon-τ (IFNT) secreted by the trophoectoderm between Days 8 and 20 post-fertilization. Epidermal growth factor (EGF) produced by the endometrium acting through EGF receptors (EGFR) present in the blastocyst seems to regulate embryonic production of IFNT. Epidermal growth factor and IFNT have been shown to play crucial roles in controlling luteolytic prostaglandin (PG) F2α (PGF) and luteotropic PGE2 production by bovine endometrium. However, it is unknown how these bioactive molecules regulate uterine function during maternal recognition of pregnancy. To clarify the main source of EGF in bovine endometrium and the mechanisms regulating the interaction between the hatched blastocyst and maternal uterine environment, the production of EGF by cultured endometrial epithelial and stromal cells and the effects of EGF on embryonic IFNT and PG were investigated. In addition, the effects of EGF on PGE2 and PGF production by cultured epithelial or stromal cells were examined. Endometrial epithelial and stromal cells were enzymatically isolated on the day of ovulation, seeded at a density of 100 000 viable cells mL–1, and cultured at 38°C in a humidified atmosphere of 5% CO2 in air. After the cells reached 90% confluence, they were cultured in the presence or absence of EGF (0.1, 1.0, 10, and 100 ng mL–1) for 24 h. Cells cultured in the absence of EGF and their cultured media were collected separately for protein analysis. Hatched bovine blastocysts (Days 8–10) were also cultured and exposed to EGF (1, 10, and 100 ng mL–1) for 24 h. Protein concentrations of EGF and IFNT in the cultured media were determined by commercial enzyme immunoassay kit. Hormonal concentrations were analysed by ANOVA followed by Fisher's protected least-significant difference procedure (PLSD) as a multiple comparison test by StatView (Abacus Concepts Inc., Berkeley, CA, USA). The concentration of EGF in the culture media of epithelial cells cultured in the absence of EGF was significantly (P &lt; 0.05) higher than in the cultured media of endometrial stromal cells. Epidermal growth factor (10 and 100 ng mL–1) increased embryonic production of IFNT and luteotropic PGE2 production but not luteolytic PGF by hatched blastocyst. EGF (100 ng mL–1) increased both PGE2 and PGF production (P &lt; 0.05) by cultured endometrial epithelial and stromal cells. The overall results suggest that endometrial epithelial cells rather than stromal cells are the main source of EGF. Epidermal growth factor produced by epithelial cells stimulates the production of IFNT by bovine trophoblasts. The capacity of conceptus to increase IFNT and luteotropic PGE2 production rather than luteolytic PGF in response to EGF stimulation may be essential for the establishment of pregnancy in cattle.

FOXL2 Is Regulated During the Bovine Estrous Cycle and Its Expression in the Endometrium Is Independent of Conceptus-Derived Interferon Tau1

FOXL2, a winged-helix/forkhead domain transcription factor, is a key gene involved in the differentiation and biological functions of the ovary. In a recent transcriptomic analysis, we found that FOXL2 expression in bovine caruncular endometrium was different from that in intercaruncular endometrium. In order to gain new insights into FOXL2 in this tissue, we determined the expression of this transcription factor during the estrous cycle and the establishment of pregnancy in cattle. The endometrial expression of FOXL2 did not vary during maternal recognition of pregnancy (Days 16-20). Using an in vivo bovine model and primary cell cultures, we showed that FOXL2 was not an interferon-tau target gene. Both FOXL2 transcript and protein were expressed from Day 5 to Day 20 of the estrous cycle, and their levels showed a significant increase during the luteolytic phase. A 2-day progesterone supplementation in heifers led to a clear down-regulation of FOXL2 protein levels, suggesting the negative impact of progesterone on FOXL2 expression. Immunohistochemistry data revealed the localization of FOXL2 in endometrial stromal and glandular cells. FOXL2 subcellular distribution was shown to be nuclear in endometrial samples collected during the luteolytic period, while it was not detected in nuclei during the luteal phase and at implantation. Collectively, our findings provide the first evidence that FOXL2 is involved in the regulation of endometrial tissue physiology.

Transcriptomic Analysis of the Bovine Endometrium: What is Required to Establish Uterine Receptivity to Implantation in Cattle ?

In cattle, the majority of pregnancy loss can be attributed to early embryonic loss which occurs prior maternal recognition of pregnancy on Day 16 (Day 0 = ovulation). During this time, carefully orchestrated spatio-temporal alterations in the transcriptomic profile of the endometrium are required to drive conceptus elongation, via secretions from the endometrium (termed histotroph) and establish uterine receptivity to implantation. The two main modulators of these processed are progesterone (P4) and the pregnancy recognition signal interferon tau (IFNT). Altered concentrations of P4 in circulation mediate its effects via the endometrium and have been associated with different rates of conceptus elongation in cattle. Transcriptomic analysis of the endometrium has shown that modulation of circulating P4 alters endometrial expression of genes that can contribute to histotroph composition, which is beneficial (when P4 is supplemented) or detrimental (when P4 is reduced) to the developing conceptus. In addition, down-regulation of the progesterone receptor, required to establish uterine receptivity, is altered in the endometrium of heifers with altered P4 concentrations. IFNT, a type 1 interferon, also significantly impacts on the endometrial transcriptome. It induces the expression of a large number of classical interferon stimulated genes as early as Day 15 of pregnancy. In summary, the successful establishment of pregnancy in cattle requires a sequence of key events to ensure appropriate maternally derived secretions, establish uterine receptivity to implantation as well as an adequate endometrial response to IFNT production.

The effects of endometritis on the establishment of pregnancy in cattle

Endometritis is common in post partum dairy cows and is associated with impaired reproductive performance reflected in reduced first service conception, reduced hazard of pregnancy over the breeding period and increased risk of reproductive culling. The observed effects may be mediated directly by bacterial products, such as lipopolysaccharide (LPS, endotoxin), or indirectly by inflammatory mediators, such as cytokines, eicosanoids, nitric oxide and oxidative stress affecting sperm, ovarian, uterine and embryonic function. An inflammatory milieu in the uterus has been associated with changes in sperm motility and function as well as increased sperm phagocytosis. Zygotes resulting from fertilisation of oocytes with sperm subjected to oxidative stress are less likely to develop to the blastocyst stage. In addition, LPS and tumour necrosis factor-α (TNFα) impair follicular steroidogenesis, growth and ovulation. Oocytes exposed to LPS or prostaglandin (PG) F(2α) during maturation are less likely to develop to blastocyst stage after fertilisation. Embryos exposed to inflammatory mediators during development have fewer trophoectoderm cells. Nitric oxide impairs development of preimplantation embryos and TNFα increases blastomere apoptosis. Endometritis in women has been associated with higher rates of implantation failure. Extragenital inflammation (e.g. mastitis) is also associated with an increased rate of embryonic loss in cattle. These observations make it clear that direct and indirect effects of endometritis, and inflammation in general, can interrupt successful reproduction at several crucial stages.

Regulation of non-classical major histocompatability complex class I mRNA expression in bovine embryos

Regulation of expression of the class I major histocompatability complex (MHC class I) at the maternal fetal interface may play a critical role in embryo survival and the establishment of pregnancy in cattle. However, information concerning immunoregulation of implantation in cattle remains quite limited. Therefore, our current research is concerned with characterizing the expression and regulatory effect of a number of immune factors in the developing bovine embryo. We have analysed the effect of embryo culture in vitro (IVC) in medium supplemented with progesterone (P4): leukemia inhibitory factor (LIF), interferon gamma (IFNG), interleukin (IL)-1B, IL3, IL4, IL10 and granulocyte-colony stimulating factor (G-CSF) on in vitro embryo development and expression of the bovine non-classical MHC class I genes NC2, NC3 and N4 in blastocysts. Cytokine supplementation during IVC did not affect cleavage rate or blastocyst development. However, embryo mRNA expression of NC2, NC3 and NC4 was significantly (p≤0.05) modified in a gene- and cytokine-specific manner. Sequence analysis of the promoter regions of these genes confirmed the presence of appropriate binding sites through which the cytokine signalling could be mediated. In contrast to the lack of effect on in vitro blastocyst development, the non-classical MHC-I expression data suggests a preferential immunomodulatory role of these cytokines during preimplantation embryo development.

Expression of enzymes involved in the synthesis of prostaglandin E2 in bovine in vitro-produced embryos

Prostaglandin E2 (PGE2) may play a major role in embryo development and the establishment of pregnancy in cattle. The biosynthesis of PGE2 implies the sequential transformation of arachidonic acid to PGH2 by cyclooxygenases (COXs), then the conversion of PGH2 to PGE2 by prostaglandin E synthases (PGESs). Quantitative RT-PCR was used to examine the expression of COX-1, COX-2, microsomal PGES-1 (mPGES-1), microsomal PGES-2 (mPGES-2) and cytosolic PGES (cPGES) mRNAs in day 7 in vitro-produced (IVP) embryos from oocytes collected by ovum pick-up in Holstein heifers. Transcripts for COX-2 and mPGES-1 were detected in all embryos, whereas transcripts for COX-1 and mPGES-2 were not detected and cPGESs were at the limit of detection in 40% of embryos. Levels of COX-2 and mPGES-1 mRNAs were significantly higher in blastocysts and expanded blastocysts than in morulae and early blastocysts. Furthermore, excellent-quality embryos (grade 1) displayed higher levels of both COX-2 and mPGES-1 than did embryos of good and medium qualities (grades 2-3). Our results suggest that bovine IVP embryos at the morula and blastocyst stages use exclusively the COX-2/mPGES-1 pathway for PGE2 biosynthesis, and that PGE2 is potentially involved in blastocyst expansion and developmental competence.

Blood flow in the wall of the preovulatory follicle and its relationship to pregnancy establishment in heifers

Color-Doppler ultrasonography was used to compare the vascularity of the preovulatory follicle between heifers that became pregnant compared with nonpregnant. Heifers ( n = 34) received a luteolytic dose of PGF2α when the diameter of the largest follicle of the second follicular wave reached ≥11 mm (actual diameter 11.63 ± 0.06 mm). An ovulation-inducing injection of GnRH analogue was given 36 h after the PGF2α treatment. Artificial insemination (AI) was performed 26 h after GnRH treatment, and ovulation occurred in all heifers within 3 h after AI. Follicle blood flow was assessed at Hour 0 (GnRH treatment) and at Hour 26 (AI). Follicle diameter at Hours 0 and 26 was greater (main effect of group; P < 0.03) in the group that became pregnant ( n = 25) compared with nonpregnant ( n = 9). Percentage of follicle wall with power Doppler signals of blood flow showed an interaction of group-by-hour ( P < 0.03), reflecting greater blood flow in the pregnant group than in the nonpregnant group at Hour 26 but not at Hour 0. Spectral-Doppler evaluation of the resistance index for a vessel in the follicle wall resulted in a group-by-hour interaction ( P < 0.03) from a lower index in the pregnant group at Hour 26. The lower index indicated greater downstream vascular perfusion in the follicle wall. Results supported the hypothesis of a positive relationship between the extent of blood flow of the preovulatory follicle and successful establishment of pregnancy in cattle.

Bovine Somatotropin Attenuates Phorbol Ester-Induced Prostaglandin F2α Production in Bovine Endometrial Cells

The recent observation that bovine somatotropin (bST) treatment at a timed insemination improves pregnancy rates in lactating dairy cows raises the possibility that growth hormone (GH) may modulate the endocrine and biochemical cross talk between the conceptus and maternal uterus at the time of pregnancy establishment in cattle. The objective of this study was to characterize the cellular and molecular mechanisms by which exogenous GH affects phorbol esterinduced prostaglandin F2α (PGF2α) production in cultured bovine endometrial (BEND) cells. Serum-deprived BEND cells were incubated with or without recombinant bovine GH (rbGH), insulin-like growth factor (IGF)-I, recombinant bovine interferon (rbIFN)-τ or a combination of rbGH + rbIFN-τ for 3h and then treated with phorbol 12,13-dibutyrate (PDBu) for an additional 6h. Exogenous PDBu increased PGF 2α secretion and steady-state levels of COX-2 mRNA within 3h. Priming of BEND cells with rbGH reduced PGF2α response to PDBu, whereas cotreatment with IGF-I amplified PDBu induction of PGF2α Preincubation of cell monolayers with rbIFN-τ suppressed PGF2α and COX-2 mRNA responses to PDBu. Inhibitory effects of rbGH and rbIFN-τ on PDBu-induced PGF2α production were additive. Results provide the first direct evidence that supplemental bST may interact with conceptus-secreted IFN-τ to modulate PGF2α secretion at the critical time of maternal recognition of pregnancy.

Relationships between nutrition and fertility in dairy cattle

AbstractThe reduced fertility that is becoming more evident in high yielding dairy cows may be related to many factors including changes in milk production, food intake and fluctuations in body condition. Metabolic and production markers have been studied as a way of predicting success to a particular artificial insemination. Successful conception to a particular service was not associated with milk production, body condition or plasma concentrations of several indicators of metabolic state around the time if insemination. This highlights the importance of time of information collection in fertility management programmes. Increased food intake may reduce systemic progesterone concentrations. This is more evident in sheep than cattle, but a positive relationship between systemic progesterone early post mating and establishment of pregnancy in cattle has been reported. However, progesterone concentrations in the ovarian vein and endometrium are not strongly correlated with systemic progesterone. Thus, the significance of modest changes in systemic progesterone in affecting oocyte and embryo development must be questioned. Blood urea concentrations can be altered by diet, and reduced pregnancy rates have been reported in cows with high urea concentrations. However, in other recent studies, no difference was reported in serum urea in cows that conceived and those that failed to conceive. Pregnancy rate was equally high in heifers when in-vitro produced embryos were transferred to heifers on high and low urea diets. When embryos were produced in sheep on high and low dietary urea, the effects on embryo development appear to occur early in the developmental process, suggesting a substantial effect on the development of the oocyte. The developmental capacity of oocytes and quality of embryos is reduced in cattle maintained on extremely high dietary intakes. Oocyte developmental capacity is reduced in cows of higher genetic merit and embryo quality can be substantially reduced in the early postpartum period. Collectively, these results suggest that high dietary intake or high metabolic load is deleterious to normal oocyte development and establishment of pregnancy. This highlights the importance of further studies on the effect of dietary intake on metabolic state and follicle, oocyte and embryo development. In a practical context, these results highlight the importance of nutritional management and avoiding changes in the amount or type of diet around the time of mating in high-production dairy cows.

Endocrine and physiological events from ovulation to establishment of pregnancy in cattle

AbstractA series of in vitro and in vivo experiments were conducted to characterise the dialogue between embryo and maternal units relative to the mechanisms controlling embryo survival in dairy cattle. Endometrial explants from pregnant cows had an attenuated PGF2αsecretory response following treatment with melittin (stimulator of PLA2) and phorbol 12, 13 dibutyrate (PDBu). Thus previous exposure to the conceptus appears to regulate the endometrial synthetic pathway at a point coincident with or distal to PLA2as well as inhibit PKC or PKC mediated events. Endometrial explants collected from cows receiving intrauterine infusions of rblFN-τ had a reduced secretory response following stimulation with PDBu indicating attenuation in PKC activity. Based upon tyrosine-phosphorylation of STAT-proteins and their translocation to the nucleus after treatment with rbIFN-τ, the JAK-STAT pathway is functional in immortalised bovine endometrial cells (BEND cells). Bend cells, exposed to rblFN-τ, reduced PDBu induction of PGF2αsecretion and also decreased protein expression of Cox-2 and PLA. RblFN-τ clearly reduced PKC mediated events leading to an antiluteolytic response in endometrial cells. Feeding diets containing 2.6, 5.2 and 7.8% Menhaden fish meal to lactating dairy cows reduced uterine secretion of PGF2αfollowing sequential injections of oestradiol and oxytocin. Thus antiluteolytic effects in early pregnancy may be amplified by feeding by-pass fats. Pregnancy rate to a timed insemination at first service post-partum is increased in association with injection of bST(500 mg; sc) given at insemination. Furthermore injection of bST at time of insemination in superovulated donor cows increased the number of blastocysts and reduced number of unfertilised embryos. Prospects of integrating novel strategies to improve embryo development and survival into reproductive management systems appear to be attainable in high producing dairy cows.

Expression of Antiviral Activity and Induction of 2′,5′-Oligoadenylate Synthetase by Conceptus Secretory Proteins Enriched in Bovine Trophoblast Protein-11

Establishment of pregnancy in cattle has been proposed to depend on production of a conceptus protein, bovine trophoblast protein-1 (bTP-1), which has a high degree of sequence homology with bovine interferon-alpha (bIFN-alpha), especially the alpha II subfamily. A preparation of bovine conceptus secretory proteins enriched for bTP-1 has antiviral and physico-chemical properties similar to other bIFN-alpha. Antiviral activity is initially detectable in uterine flushings on Day 14 of pregnancy, when the conceptus measures 4-5 mm in length, and increases as the conceptus elongates through Day 18. Day 17 conceptuses produce more than 10(6) U antiviral activity during 24 h of culture. All IFNs induce the enzyme 2',5'-oligoadenylate synthetase, which catalyzes production of 2',5'-oligo(A), which in turn is involved in antiviral and growth inhibitory effects of IFNs. This enzyme activity is induced in Madin-Darby bovine kidney cells by the partially purified bTP-1 preparation similarly to IFN-alpha, -beta, and -gamma. Likewise, the partially purified bTP-1 and bIFN-alpha 1 induce 2',5'-oligo(A) synthetase activity in monolayers of endometrial epithelial and stromal cells. Compared to epithelial cells, stromal cells have higher baseline activity of 2'-5'-oligo(A) synthetase activity (p less than 0.01) and show a greater degree of induction in the presence of either the partially purified bTP-1 or bIFN-alpha 1 (p less than 0.01). Also, 2',5'-oligo(A) synthetase of endometrial stromal cells is induced to a greater degree by our enriched bTP-1 preparation than by bIFN-alpha 1 (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Establishment of pregnancy in cattle by reconstituted eggs produced by fusion of an in vitro matured oocyte and a blastomere from a 16-cell embryo produced in vivo

The techniques of in vitro maturation (IVM) and in vitro fertilization (IVF) of bovine oocytes, with development of the resulting embryos to advanced preimplantation stages in vitro, have gained widespread popularity in recent years because of the potential for obtaining information about regulatory mechanisms, and for inexpensively mass-producing embryos for research or for transfer. However, the efficiency of the techniques (blastocyst yield) is unsatisfactory, due to inadequate information about the requirements of bovine embryos for development in culture, and of oocytes for achieving normal maturation. An experiment was designed to determine effects of using different media for bovine oocyte maturation on subsequent embryo development. Five of seven media tested on oocyte maturation resulted in higher levels of fertilization and/or first cleavage division. In addition, the data indicated that culture conditions for oocyte maturation also affect development to the morula/blastocyst stages. A second experiment evaluated culture conditions for embryo development, using two media (HECM-1 and TCM-199) with or without supplementation by serum and/or oviduct cell conditioning, in a 2×2×2 factorial design. Neither serum supplementation nor conditioning nor the two together increased morula/blastocyst formation (35–46% in all treatments). Serum had a biphasic effect, suppressing the first cleavage division but enhancing morula compaction. No block was observed at the 8- to 16-cell stage with TCM-199 + serum, contrary to many other studies. The data support the hypothesis that the reported stimulation of bovine IVM/IVF embryo development by somatic cell conditioning is due to removal of inhibitory influences from the culture environment. The ability to support development of IVM/IVF embryos to the morula and blastocyst stages in defined media (i.e., without serum or somatic cell conditioning) will help to elucidate the metabolic and nutritional requirements of bovine oocytes and embryos, and to formulated improved culture media. These advances will increase the utility of bovine IVM/IVF for both basic and applied resarch, and for commercial embryo production.

Prolongation of luteal lifespan in cows by intrauterine infusion of recombinant bovine alpha-interferon.

The infusion of recombinant bovine interferon-alpha I 1 into the uterus of cyclic cows from day 15.5 to 21 after estrus delayed luteolysis. Interferon-treated cows had a mean inter-estrous interval of 26.8 +/- 1.39 d, a length that was longer (P less than 0.02) than the inter-estrous interval for control cows (22.8 +/- 0.80 d). Measurements of circulating concentrations of progesterone confirmed that the longer inter-estrous intervals were due to an increase in corpus luteum lifespan following treatment with interferon. These results support the hypothesis that an alpha-interferon-like molecule could play a major role in establishment of pregnancy in cattle.