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Topic:Embryo
Equine embryo research focuses on the early developmental stages of horses, encompassing the formation, growth, and differentiation of the embryo. This area of study is significant for understanding reproductive biology, improving breeding programs, and advancing assisted reproductive technologies in equines. Key aspects include the processes of fertilization, embryonic development, and implantation. Researchers investigate factors influencing embryonic viability, such as genetic and environmental influences, to enhance reproductive success rates. This page aggregates peer-reviewed research studies and scholarly articles that explore the biological mechanisms, technological advancements, and applied methodologies related to equine embryos.
Long term exposure to T-2 Fusarium mycotoxin fails to alter luteal function, follicular activity and embryo recovery in mares. The effect of long term administration of T-2 toxin was studied in 6 Trotter mares during the summer and early autumn. After one complete oestrous cycle (Cycle 1) each mare was given 7 mg purified T-2 toxin per os daily (1 mg/ml in ethyl alcohol) beginning on Day 10 after ovulation in Cycle 2. Exposure to toxin was continued for 32-40 days, until Day 7 of Cycle 4. During this period all the animals remained in good physical condition, but skin lesions were observed around the mouth in 3 cases. Toxin administration had no effect on the length of the interovulatory interval or on the lengths of ...
The use of early pregnant mares as embryo recipients. Fourteen normal, cyclic mares, treated to synchronise oestrus and ovulation and inseminated artificially with fresh semen, were assigned to a donor or a recipient group after ovulation, with the aim of obtaining a degree of synchrony of > or =2 days. Ten embryos, collected on Day 6 or 7 after ovulation (Day 0), were transferred nonsurgically to inseminated recipient mares (IRM) that had ovulated up to 5 days after the respective donors, or to pregnant recipient mares (PRM) that had ovulated 2-7 days before the donors. Embryonic size and development, as determined by ultrasound examination, wer...
Distribution of putative primordial germ cells in equine embryos. Eighteen equine embryos, 3 each on Days 20, 22, 24, 26, 28 and 30 post ovulation, were collected transcervically by uterine lavage, fixed in 4% paraformaldehyde and embedded in paraffin wax. Ten micron serial sections were stained to determine alkaline phosphatase (AP) activity in the cells. Positive cells were counted and their approximate location determined. The cells were approximately 8 microm in diameter and the entire cell, except the nucleus, stained strongly with many small round areas of intense staining in the cytoplasm. The cells varied from round to elongated in shape and pseudopo...
Treatment of equine oocytes with A23187 after intracytoplasmic sperm injection. In vitro matured horse oocytes with a first polar body (n = 68) were each injected with a single spermatozoon and divided into 2 groups: Group 1 oocytes were treated with 10 microM calcium ionophore A23187 for 5 min while Group 2 oocytes received no activation treatment. After culture in vitro for 2 days, significantly more oocytes treated with A23187 (5/24, 21%) cleaved than oocytes without activation treatment (2/44, 5%, P<0.05). All 7 cleaved zygotes from both treatment groups were transferred to recipient mares but no pregnancies resulted.
The effect of sucrose in the thawing solution on the morphology and mobility of frozen equine embryos. Seventy-five embryos were collected 6 days after ovulation. Sixty embryos were frozen in straws using glycerol as the cryoprotectant in an automatic freezer. In Experiment 1 the freezing and thawing media were supplemented with 1.3 g/l PVP; in Experiment 2 the supplement was 5% FCS. The embryos were thawed for 30 s at +37 degrees C in a waterbath. In Experiment 1 glycerol was removed from 10 embryos in 6 steps. In 10 other embryos, glycerol and sucrose were both removed from the medium in 6 steps. After glycerol and sucrose removal, the embryos were stained with 4',6'-diamidino-2-phenylindole ...
Cryopreservation procedures for Day 7-8 equine embryos. Larger grade 1 or 2 (1 = excellent,.... 4 = degenerate) equine embryos that ranged in diameter from 300 to 680 microm and were recovered from mares on Day 7 or 8 after ovulation, were randomly assigned to 3 widely divergent cryopreservation treatments. Treatment 1 consisted of cooling from -6 degrees C to -35 degrees C at 0.5 degrees C per min followed by plunging into liquid nitrogen, with a one-step addition and a 4-step removal of 1.0 M glycerol. Treatment 2 (step-down equilibration) consisted of a 2-step addition of glycerol to 4.0 M followed by a decrease to 2.0 M prior to freezing, with ...
Cryopreservation of equine embryos with glycerol plus sucrose and glycerol plus 1,2-propanediol. Six or 7-day-old equine embryos were divided into 4 groups; Group 1, n = 15, Day 7 embryos destined for immediate transfer; Group 2, n = 15, Day 6 embryos destined for deep-freezing with glycerol plus sucrose as cryoprotectant; Group 3, n = 10, Day 6 embryos destined for deep-freezing with glycerol plus 1,2-propanediol as cryoprotectant and Group 4, n = 3, fresh embryos destined for ultrastructural analysis. All the frozen/thawed embryos were transferred to recipient mares, except 3 embryos in Group 3 that were subjected to ultrastructural analysis. After thawing the cryoprotectants were remov...
Parentage testing of Day 10 equine embryos by amplified PCR analysis of microsatellites. Paternity analysis was performed on the DNA of 21 equine embryos collected nonsurgically 10 days after ovulation from known mares, but involving 3 possible sires. After extraction, the DNA of each embryo was typed by radioactive PCR amplification using 10 characterised microsatellites; HMS 1, 2, 5, 6, 7 and 8 (Guérin et al. 1994) and HTG 3, 4, 6 and 10 (Marklund et al. 1994). The 21 dams and 3 sires were genotyped using DNA extracted from blood and amplified by PCR. After electrophoresis and autoradiography of the PCR products of the embryo and parents, the alleles of the embryo were compared...
Spontaneous multiple ovulation and development of multiple embryonic vesicles in a mare. A Warmblood mare was observed to ovulate spontaneously 12 follicles within 2 days, none of which exceeded 22 mm in diameter. On Days 13 and 17 after ovulation, 6 embryonic vesicles were identified in the uterus by ultrasonography but by Day 26, 5 of the vesicles had disappeared. Development of the surviving conceptus was monitored until Day 42. Plasma progesterone concentrations rose to 14 ng/ml on Day 7, decreased over the next 8 days and then plateaued to around 4-6 ng/ml until Day 70. The occurrence of multiple spontaneous ovulations was diagnosed repeatedly in this mare. However, the devel...
Effect of anti-freeze protein (AFP) on the cooling and freezing of equine embryos as measured by DAPI-staining. Equine embryos recovered on Day 6 after ovulation were cooled to +4 degrees C, or frozen with AFP alone or together with glycerol. Twenty embryos (140-200 microm in diameter) were randomly assigned to 6 treatment groups. In the first 3 groups, the embryos were cooled from room temperature to +4 degrees C at a rate of 3 degrees C/min and warmed again at a rate of 32 degrees C/min in a programmable freezer. In the second 3 groups, the embryos were frozen using a standard protocol, stored in liquid nitrogen for 5-7 days and then thawed in a 37 degrees C waterbath. After cooling/warming or freezin...
Success rates when attempting to nonsurgically collect equine embryos at 144, 156 or 168 hours after ovulation. The purpose of this study was to evaluate the exact age when the equine embryo reaches the uterus. The time of ovulation was determined by hourly ultrasound examinations starting 32 h after an injection of crude equine pituitary gonadotrophin or human chorionic gonadotrophin, or after the first of 4 injections of buserelin. Nonsurgical uterine flushings were carried out 144 h (Day 6), 156 h (Day 6.5) or 168 h (Day 7) after ovulation. Induction of ovulation was attempted in 101 oestrous cycles and 61 of 101 mares (60.4%) ovulated 32-44 h post injection. Sixty embryo collections were performed w...
Effects of follicular fluid supplementation of in-vitro maturation medium on the fertilization and development of equine oocytes after in-vitro fertilization or intracytoplasmic sperm injection. The aim of this study was to compare the effect of the addition of follicular fluid (FF) collected from preovulatory follicles with that of oestrous mare serum (EMS) (acting as the control) to TCM-199 medium on the in-vitro maturation, fertilization and development of equine cumulus-enclosed oocytes. Oocytes (<30 mm in diameter) were obtained from the ovaries of slaughtered mares. After in-vitro maturation in the presence of the two supplements, their fertilization, cleavage and developmental potential were compared after conventional in-vitro fertilization (IVF) or intracytoplasmic sperm inje...
Evidence for the presence of sodium- and potassium-dependent adenosine triphosphatase alpha1 and beta1 subunit isoforms and their probable role in blastocyst expansion in the preattachment horse conceptus. The unusual hypotonicity of equine blastocyst fluid has prompted us to investigate the role of sodium- and potassium-dependent adenosine triphosphatase (Na+,K+-ATPase) in the process of fluid accumulation in the horse conceptus. Nine mares were used for the experiments. Reverse transcriptase polymerase chain reaction was conducted on two sets of five conceptuses recovered between 12 and 28 days (+/- 1 day) after ovulation. Messenger RNAs encoding the alpha1 and beta1 subunit isoforms of Na+,K+-ATPase were detected in all embryonic tissues examined. Western blot analysis showed that alpha1 and ...
[Spontaneous and manual embryo reduction in mares: a twin prevention programme]. Data from the literature and own data for 67 twin pregnancies were used to establish the factors essential to the decision on how to treat twins at different gestational ages. Spontaneous (natural) reduction was compared with manual embryo reduction. Manual embryo reduction is always indicated when a twin is diagnosed before day 16 after ovulation. Thereafter, the type of fixation is the main determinant. Manual embryo reduction is always first choice for bilateral and unilateral non adjacent embryos and must be applied as early as possible. The change of natural reduction up to day 30, is hig...
Embryological development of the equine heart. The research examines the embryological development of the equine (horse) heart, aiming to understand its formation, function, and performance abilities. The study observes the developing morphology of the equine heart, […]
Intracytoplasmic sperm injection (ICSI) versus conventional IVF on abattoir-derived and in vitro-matured equine oocytes. Conventional IVF as well as several assisted microfertilization techniques have shown limited success in the horse. After recent positive results achieved with intracytoplasmic injection of a single spermatozoon (ICSI) in human IVF, we chose to try the method in the horse. We compared conventional IVF to ICSI by fertilization rates of oocytes with compact and expanded cumuli and by developmental potential of the resulting embryos. Cumulus-oocyte complexes (COCs) were obtained by aspirating the follicular fluid from the ovaries of slaughtered mares. Complexes showing complete cumulus investment...
Biochemical changes in the equine capsule following prostaglandin-induced pregnancy failure. The equine embryonic capsule, an acellular covering that envelops the conceptus during the second and third weeks of pregnancy, is composed of mucin-like glycoproteins. Its structure is consistent with a dual role during early pregnancy: protection of the conceptus, and communication between the embryo and the mother. Loss of sialic acid from the capsular glycoproteins at day 16 correlates with the time of "fixation," or loss of conceptus mobility throughout the uterine horns. This study investigated how the structure of the capsule is linked to the maintenance of pregnancy. Six pregnancies, c...
A developmental switch in expression from blastocyst to endometrial/placental-type cytochrome P450 aromatase genes in the pig and horse. Pig blastocysts exhibit a transient period of estrogen production at periimplantation, with a second, more sustained period of estrogen synthesis occurring in endometrium and placenta at later pregnancy. Previously we reported the isolation of cDNA clones encoding a novel isoform of cytochrome P450 aromatase (the terminal enzyme in the estrogen biosynthetic pathway) from porcine periimplantation blastocysts. The present study investigated pregnancy-associated expression, in blastocysts and maternal reproductive tract tissues of this and an additional mRNA transcript encoding a distinct P450 ar...
The influence of progesterone-induced proteins on glucose metabolism in early equine embryos. The influence of different maternal plasma progesterone concentrations on embryonic glucose metabolism was studied. Uterine flushes were obtained after treating ovariectomized mares (n = 3) with 0 (control), 100 or 200 mg progesterone daily for 7 d. A group of progesterone-induced proteins (PIP) of Mr approximately 20,000 were identified in flushes from progesterone treatments by SDS-PAGE but were not observed in control flushes. Progesterone-induced proteins were removed from half the pooled flush in each treatment group by Sepharose blue CL-6B. In a 3 x 2 factorial (progesterone treatments, ...
Production of free estrogens and estrogen conjugates by the preimplantation equine embryo. In vitro production of free estrogens and estrogen conjugates by intact Day 12.5, 13.5 and 14.5 equine embryos was measured at 2-h intervals over a 24-h culture period. Production of free estrogens was higher for Day 14.5 than Day 12.5 embryos. Differences in production of conjugated estrogens were not significant, but a trend toward increased production with increased age of embryo was apparent. No trend toward increased free and conjugated estrogen production per cell was observed with age. Embryo diameter and number of cells increased with age but varied considerably within groups. The amou...
Trophectoderm projections: a potential means for locomotion, attachment and implantation of bovine, equine and human blastocysts. The behaviour of bovine, equine and human blastocysts was studied in vitro by time-lapse videomicrography and computer imaging. This study revealed that cytoplasmic extensions of the trophectoderm ['trophectoderm projections' (TEP)] were expressed by embryos of all three species, prior to or during zona escape. Bovine and human blastocysts escaped their zonae with a combination of blastocoele expansion, collapse and re-expansion coupled with the penetration of the zona pellucida by TEP. In equine embryos, after several cycles of blastocoele expansion and collapse, trophectoderm ruptured the zo...
Factors related to the time of fixation of the conceptus in mares. The temporal relationships among day of conceptus fixation (cessation of mobility), conceptus diameter, uterine tone, uterine contractility, and myometrial and endometrial thickness of the middle and caudal segments of the uterine horns were assessed in 13 pony mares with fixation in the caudal segment of a uterine horn. The mean day of fixation (14.9 +/- 0.3) was established by 2-h mobility trials. Uterine tone increased (P < 0.0001) gradually over Days 11 to 21, whereas uterine contractility decreased (P < 0.0001) between Days 14 and 18. The diameter of the spherical embryonic vesicle ...
Nucleologenesis and ribonucleic acid synthesis in preimplantation equine embryos. The nucleolus is believed to be the active site of rRNA synthesis in all eukaryotic cells. In preimplantation embryos, the embryonic genome is apparently more or less silent up to a species-specific developmental stage at which a major burst of transcription occurs. Here we report on nucleologenesis and some ultrastructural aspects of the onset of RNA synthesis in equine embryos during in vivo development. The zygotes and embryos up to blastocyst stages were surgically recovered from normally cycling mares. Mares were induced to ovulate by treatment with 3000 IU hCG and inseminated 20 and 34 h...
[Preservation of genetic variation in domestic animals using biotechnical methods]. The conservation of endangered breeds as live animals is at present the main national strategy of the government and breeding organizations to maintain genetic diversity. Fourty-three breeds and some old strains of cattle, pig, sheep, goat and horses are currently involved. Cryopreservation and banks for sperm, embryos or DNA are another type of genetic material which could subsequently be used for breeding and production in agriculture. Present semen banks involve 9 endangered cattle breeds and also a small amount of deep-frozen sperm of some endangered sheep and horse breeds. Only 2 embryo b...
Cryopreservation of equine embryos. Principles and procedures for cryopreservation of equine embryos are described. Embryos less than 250 microM in diameter can be cryopreserved successfully if glycerol is used as the cryoprotectant. Cooling is takes place in such a way that most of the water leaves the cells before intracellular ice forms, and glycerol is removed after thawing without undue osmotic swelling of cells. Vitrification procedures also show promise for small embryos. Satisfactory procedures for cryopreserving embryos of more than 250 microM in diameter are not yet available.
Early embryonic development and evaluation of equine embryo viability. Tremendous progress has been made in the development of assisted reproductive techniques that may enhance the reproductive efficiency of the horse. However, techniques that involve the manipulation of oocytes and/or embryos may themselves be detrimental to embryo viability and subsequent development. Therefore, an objective method of assessing viability of embryos before and/or after oocyte/embryo manipulation is desirable. At this time, morphologic evaluation is the most widely used method of determining the viability of equine embryos. Although morphologic assessment of embryo quality will n...
Maturation and fertilization of equine oocytes. Equine oocytes obtained either by transvaginal ultrasound-guided follicular aspiration or from slaughterhouse ovaries can be matured in vitro. This generally requires culture in TCM-199 containing serum and hormones for 30 to 36 hours. With this protocol, approximately 50% to 60% of the oocytes are at metaphase-II at the end of the culture period. At least some of these oocytes appear viable based on production of fertilized eggs either through in vitro fertilization or fertilization in vivo of a recipient mare. The success of producing equine embryos in vitro is still extremely low. More than...
Recovery rate and quality of embryos from mares inseminated at the first post-partum oestrus. The pregnancy rate is lower in mares inseminated at the first post-partum (p.p.) oestrus (40-50%) compared with pregnancy rates in subsequent oestrous cycles (55-65%). The causes of the lowered pregnancy rate are not fully understood. The aim of the present study was to examine if embryonic defects could be one of the reasons for lowered pregnancy rate. A total of 23 p.p. and 14 non-lactating control mares were flushed 7 days after detection of ovulation. Embryo recovery rate was 48% and 71% in p.p. and control mares, respectively (p = 0.16). Embryos were photographed, measured, graded and sta...
Recovery rate and quality of embryos from mares inseminated at the first post-partum oestrus. The pregnancy rate is lower in mares inseminated at the first post-partum (p.p.) oestrus (40-50%) compared with pregnancy rates in subsequent oestrous cycles (55-65%). The causes of the lowered pregnancy rate are not fully understood. The aim of the present study was to examine if embryonic defects could be one of the reasons for lowered pregnancy rate. A total of 23 p.p. and 14 non-lactating control mares were flushed 7 days after detection of ovulation. Embryo recovery rate was 48% and 71% in p.p. and control mares, respectively (p = 0.16). Embryos were photographed, measured, graded and sta...
Initiation of transcription and nucleologenesis in equine embryos. The time of activation of the embryonic genome (maternal-embryonic transition) in equine embryos was investigated by assessing incorporation of 3H-uridine and nucleolar development. In Experiment 1, embryos were recovered from the oviduct (n = 15) and the uterus (n = 3). Recovered embryos were assessed for morphologic development and quality score. Recovered embryos with less than 8 cells (two cells, n = 4; four cells, n = 5; five cells, n = 2) were incubated with 3H-uridine (560 microCi/ml) for 10 hr, while eight-cell embryos (n = 2), morulae (n = 2), and blastocysts (n = 3) were incubated wi...
