Reproduction : the official journal of the Society for the Study of Fertility.
Publisher:
Journals of Reproduction and Fertility, Ltd.. Bristol, UK : Published for the Society for Reproduction and Fertility by BioScientifica (2004)
Frequency: Monthly
Country: England
Language: English
Author(s):
Society for the Study of Fertility., Society for Reproduction and Fertility.
Start Year:2001 -
ISSN:
1470-1626 (Print)
1741-7899 (Electronic)
1470-1626 (Linking)
1741-7899 (Electronic)
1470-1626 (Linking)
Impact Factor
3.8
2022
| NLM ID: | 100966036 |
| (OCoLC): | 45662873 |
| Coden: | RCUKBS |
| LCCN: | 2001245725 |
| Classification: | W1 RE213KD |
Expression of steroidogenic enzymes during equine testicular development. In the mammalian testis, Leydig cells are primarily responsible for steroidogenesis. In adult stallions, the major endocrine products of Leydig cells include testosterone and estrogens. 3β-hydroxysteroid dehydrogenase/Δ(5)-Δ(4)-isomerase (3βHSD) and 17α-hydroxylase/17,20-lyase (P450c17) are two key steroidogenic enzymes that regulate testosterone synthesis. Androgens produced by P450c17 serve as substrate for estrogen synthesis. The aim of this study was to investigate localization of the steroidogenic enzymes P450c17, 3βHSD, and P450arom and to determine changes in expression during dev...
Trophoblast stem cell marker gene expression in inner cell mass-derived cells from parthenogenetic equine embryos. Although putative horse embryonic stem (ES)-like cell lines have been obtained recently from in vivo-derived embryos, it is currently not known whether it is possible to obtain ES cell (ESC) lines from somatic cell nuclear transfer (SCNT) and parthenogenetic (PA) embryos. Our aim is to establish culture conditions for the derivation of autologous ESC lines for cell therapy studies in an equine model. Our results indicate that both the use of early-stage blastocysts with a clearly visible inner cell mass (ICM) and the use of pronase to dissect the ICM allow the derivation of a higher proportion...
Viability of equine embryos after puncture of the capsule and biopsy for preimplantation genetic diagnosis. The equine embryo possesses a capsule that is considered essential for its survival. We assessed viability after breaching the capsule of early (Day 6) and expanded (Day 7 and 8) equine blastocysts by micromanipulation. The capsule was penetrated using a Piezo drill, and trophoblast biopsy samples were obtained for genetic analysis. Pregnancy rates for Day-6 embryos, which had intact zonae pellucidae at the time of recovery, were 3/3 for those biopsied immediately after recovery and 2/3 for those biopsied after being shipped overnight under warm (∼28 °C) conditions. The pregnancy rates for ...
Successful transfer of day 10 horse embryos: influence of donor-recipient asynchrony on embryo development. A total of 78 day 10 horse embryos were transferred non-surgically to recipient mares that had ovulated 9, 7, 6, 5, 4, 3, 2 or 1 day after (negative asynchrony), on the same day (synchronous), or 2 or 4 days before (positive asynchrony) the donor (n=6 or 8 mares per group). Pregnancy rates between 100% (6/6) and 63% (5/8) were seen in recipient mares that were between +2 and -6 days asynchronous. Embryo survival to the heartbeat stage declined in recipients that were -7 days asynchronous and no embryos survived in recipients that were -9 days asynchronous. Irrespective of uterine asynchrony, c...
Estrogen metabolism by the equine embryo proper during the fourth week of pregnancy. Estrogen production by the trophoblast is considered important in early equine pregnancy and leads to high concentrations in yolk-sac (Y-S) fluid. The embryo proper is a potential site for their action. We examined estrogen metabolism in the embryo proper because some actions of estrogens are derived from locally formed metabolites. The embryo proper, as well as separated extraembryonic tissues, of conceptuses collected about day 25 of pregnancy, were incubated with (3)[H]-estrone (E(1)) and (3)[H]-estradiol (E(2)). Steroids were recovered from media by solid-phase extraction and eluted separa...
The uterine environment modulates trophectodermal POU5F1 levels in equine blastocysts. The reported patterns of trophectodermal expression of POU5F1 protein in blastocysts vary among species, and are possibly related to the differences in placental growth and function. This study investigated the pattern of embryonic POU5F1 expression in the horse, a species with delayed placental formation. Immature equine oocytes expressed POU5F1 protein in the cytoplasm and nucleus. Staining for POU5F1 protein in in vitro-produced (IVP) embryos decreased to day 5 of culture, then the nuclear staining increased to day 7. IVP day-7 to -11 blastocysts showed POU5F1 staining in nuclei throughout ...
The extracellular calcium-sensing receptor is expressed in the cumulus-oocyte complex in mammals and modulates oocyte meiotic maturation. The extracellular calcium-sensing receptor (CASR) plays an important role in cells involved in calcium (Ca2+) homeostasis by directly sensing changes in the extracellular Ca2+ ion concentration. We previously reported the localization and quantitative expression of CASR protein in human oocytes. In this study, we examined the expression and the functional role of CASR during oocyte meiotic maturation in a large mammal animal model, the horse. As in humans, CASR protein was found to be expressed in equine oocytes and cumulus cells. Western-blot analysis revealed a single 130 kDa band in denuded...
Lipid peroxidation, assessed with BODIPY-C11, increases after cryopreservation of stallion spermatozoa, is stallion-dependent and is related to apoptotic-like changes. Lipid peroxidation (LPO) of stallion spermatozoa was assessed in fresh semen and in samples of the same ejaculates after freezing and thawing. Particular attention was paid to individual differences in the susceptibility to LPO and its possible relationship with freezability. Innate levels of LPO were very low in fresh spermatozoa but increased after thawing, a change that was largely stallion-dependent. The level of LPO in fresh spermatozoa was not correlated with that of the thawed spermatozoa. Negative correlations existed between LPO and intact membranes post-thaw (r=-0.789, P<0.001), a...
Uterine influences on embryogenesis and early placentation in the horse revealed by transfer of day 10 embryos to day 3 recipient mares. Eight day 10 horse embryos were transferred non-surgically to recipient mares that had ovulated 7 days after the donors. The embryonic vesicle was seen ultrasonographically in all eight recipients, and three out of eight (38%) of the vesicles developed an embryo proper with a beating heart. Conceptus expansion was initially slower than that in control mares but continued until day 22 (recipient day 15). Time of fixation of the vesicle was related to its diameter, rather than uterine stage. Although the embryo proper first appeared ultrasonographically on day 22, as normal, it grew more slowly ...
Protective effects of the cumulus-corona radiata complex during vitrification of horse oocytes. Vitrifying oocytes is a potentially valuable means of preserving the female germ line, but significantly compromises oocyte developmental competence. This study examined the hypothesis that the cumulus complex protects the oocyte during vitrification. Vitrified-warmed immature cumulus oocyte complexes (COCs) were labelled with a plasma membrane impermeant DNA marker (ethidium homodimer-1) to examine the percentage and location of dead cumulus cells, and to investigate the effect of the proportion of dead cells (+1,+2 or +3) on the success of in vitro maturation (IVM). Further, oocytes were lab...
Seasonal effects on the response of ovarian follicles to IGF1 in mares. The response of follicles to IGF1 was compared between the transition into the ovulatory season (transitional period) and the ovulatory season (ovulatory period) in eight mares using a cross-over experimental design within periods. Granulosa cells were collected from follicles 15-24 or 25-34 mm and expression of IGF1R, IGF2R, FSHR, LHCGR and PAPPA was determined by qPCR. In addition, 10 mg IGF1 or vehicle were injected into the largest follicle (transitional period) or the second largest follicle (ovulatory period) of a follicular wave before the beginning of diameter deviation between the two...
Significance of aquaporins and sodium potassium ATPase subunits for expansion of the early equine conceptus. Expansion of the equine conceptus can be divided into blastocoel and yolk sac phases. The endodermal layer transforming the blastocoel into the yolk sac is completed around day 8 of pregnancy. From that time, the size of the spherical conceptus increases tremendously due mainly to the accumulation of fluid rather than cell multiplication. In this study, we have investigated the abundance and localisation of Na(+)/K(+)-ATPases and aquaporins (AQP) in the equine conceptus on days 8, 10, 12, 14 and 16 by multiplex reverse transcriptase PCR, Western blot and immunohistochemistry. During conceptus ...
Uterine blood flow and perfusion in mares with uterine cysts: effect of the size of the cystic area and age. Transrectal color and power Doppler ultrasonography was used to study uterine blood flow and perfusion in mares with and without uterine cysts. Vascular perfusion of the uterus and blood flow velocities, vascular perfusion, diameter, circumference, and area of a cross section of the mesometrial attachment were evaluated. To study the effect of internal cysts, two matched groups (cystic and control, n=21 mares/group) were used. Uterine vascular perfusion in mares with cysts was less (P<0.0001) in the cystic than the noncystic region and less (P<0.0009) than that for controls. Mares with c...
Production of cloned horse foals using roscovitine-treated donor cells and activation with sperm extract and/or ionomycin. We evaluated the effect of different activation treatments on the production of blastocysts and foals by nuclear transfer. Donor cells were prepared using roscovitine treatment, which has previously been associated with increased production of viable offspring. All activation treatments were followed by culture in 6-dimethylaminopurine (6-DMAP) for 4 h. In experiment 1, blastocyst production after activation by injection of sperm extract followed by treatment with ionomycin was significantly higher than that for activation with a serial treatment of ionomycin, 6-DMAP, and ionomycin (12.5 vs 2....
Characterization of NADPH oxidase 5 in equine testis and spermatozoa. Reactive oxygen species (ROS) play an important role in normal sperm function, and spermatozoa possess specific mechanisms for ROS generation via an NAD(P)H-dependent oxidase. The aim of this study was to identify the presence of an NADPH oxidase 5 (NOX5) in equine testis and spermatozoa. The mRNA of NOX5 was expressed in equine testis as detected by northern blot probed with human NOX5 cDNA and by RT-PCR. Immunoblotting with affinity purified alpha-NOX5 revealed one major protein in equine testis and other tissues. Immunolocalization of NOX5 showed labeling over the rostral sperm head with so...
Changes in major proteins in the embryonic capsule during immobilization (fixation) of the conceptus in the third week of pregnancy in the mare. During the third week of pregnancy, the equine conceptus is enclosed within a capsule, the glycan composition of which changes at around day 16 (ovulation = day 0) when the conceptus becomes immobilized (fixed) in the uterine lumen. Our objective was to characterize the process of fixation by identifying changes in major capsule-associated proteins. Individual equine conceptuses (n = 55) were collected transcervically by uterine lavage between days 13.5 and 26.5. Major proteins extracted from capsules were compared with those in fluids from the uterus and yolk sac by SDS-PAGE. Until day 14, a ...
Cryptorchidism in common eutherian mammals. Cryptorchidism is failure of one or both testes to descend into the scrotum. Primary fault lies in the testis. We provide a unifying cross-species interpretation of testis descent and urge the use of precise terminology. After differentiation, a testis is relocated to the scrotum in three sequential phases: abdominal translocation, holding a testis near the internal inguinal ring as the abdominal cavity expands away, along with slight downward migration; transinguinal migration, moving a cauda epididymidis and testis through the abdominal wall; and inguinoscrotal migration, moving a s.c. cauda...
Cloning of equine prostaglandin dehydrogenase and its gonadotropin-dependent regulation in theca and mural granulosa cells of equine preovulatory follicles during the ovulatory process. The mammalian ovulatory process is accompanied by a gonadotropin-dependent increase in follicular levels of prostaglandin E2 (PGE2) and PGF2alpha, which are metabolized by 15-hydroxy prostaglandin dehydrogenase (PGDH). Little is known about ovarian PGDH regulation in non-primate species. The objectives of this study were to characterize the structure of equine PGDH and its regulation in follicles during human chorionic gonadotropin (hCG)-induced ovulation. The full-length equine PGDH was obtained by RT-PCR, 5'- and 3'-rapid amplification of cDNA ends (RACE). Its open reading frame encodes a 26...
Molecular characterization of tumor necrosis alpha-induced protein 6 and its human chorionic gonadotropin-dependent induction in theca and mural granulosa cells of equine preovulatory follicles. The preovulatory rise in gonadotropins causes an expansion of the cumulus-oocyte complex, a process requiring the induction of several genes. The objectives of this study were to clone the equine tumor necrosis factor alpha-induced protein 6 (TNFAIP6), and investigate its regulation in equine follicles during human chorionic gonadotropin (hCG)-induced ovulation. The isolation of the equine TNFAIP6 cDNA revealed that it contains an open reading frame of 834 bp (including the stop codon), encoding a predicted 277 amino acid protein that is highly similar (91-93% identity) to known mammalian homo...
Effect of cumulus morphology and maturation stage on the cryopreservability of equine oocytes. Oocyte cryopreservation is a potentially valuable way of preserving the female germ line. However, the developmental competence of cryopreserved oocytes is presently poor. This study investigated whether the morphology of the cumulus complex surrounding an immature equine oocyte and/or the oocyte's stage of maturation affect its cryopreservability. Compact (Cp) and expanded (Ex) cumulus oocyte complexes (COCs) were vitrified either shortly after recovery (germinal vesicle stage, GV) or after maturation in vitro (IVM); cryoprotectant-treated and -untreated non-frozen oocytes served as controls....
Follicle selection in cattle and horses: role of intrafollicular factors. The eminent event in follicle selection during a follicular wave in monovular species is diameter deviation, wherein one follicle continues to grow (developing dominant) and other follicles (subordinates) begin to regress. In cattle, the IGF system, oestradiol and LH receptors are involved in the intrafollicular events initiating deviation as indicated by the following: (1) concentrations of free IGF-I and oestradiol in the follicular fluid and number of LH receptors in the follicular wall increase more dramatically in the future dominant follicle than in the future subordinate follicles befor...
Temporal relationship between proliferating and apoptotic hormone-producing and endothelial cells in the equine corpus luteum. The temporal relationship between endothelial cell death, vascular regression and the death of hormone-producing cells in the mare has not been established. To determine the dynamics of cell proliferation and death throughout the luteal phase, corpora lutea were studied at the early, mid- and late luteal phase, and after treatment with cloprostenol in the mid-luteal phase to induce premature luteolysis. Changes in cell proliferation and apoptosis were investigated utilising specific markers (phosphorylated histone-3 and activated caspase-3 respectively). Histone-3 positive cells were most abun...
Production of horse foals via direct injection of roscovitine-treated donor cells and activation by injection of sperm extract. We evaluated the effects of different donor cell treatments and activation methods on production of blastocysts after equine nuclear transfer. Nuclear transfer was performed by direct injection of donor cells, using a piezo drill, and standard activation was by injection of sperm factor followed by culture with 6-dimethylaminopurine. There was no difference in blastocyst development between embryos produced with roscovitine-treated or confluent donor cells (3.6% for either treatment). Addition of injection of roscovitine or culture with cycloheximide at the time of activation did not affect bl...
Germ cell development in equine testis tissue xenografted into mice. Grafting of testis tissue from immature animals to immunodeficient mice results in complete spermatogenesis, albeit with varying efficiency in different species. The objectives of this study were to investigate if grafting of horse testis tissue would result in spermatogenesis, and to assess the effect of exogenous gonadotropins on xenograft development. Small fragments of testis tissue from 7 colts (2 week to 4 years of age) were grafted under the back skin of castrated male immunodeficient mice. For 2 donor animals, half of the mice were treated with gonadotropins. Xenografts were analyzed a...
Centrosome changes during meiosis in horse oocytes and first embryonic cell cycle organization following parthenogenesis, fertilization and nuclear transfer. Various types of cell cycle organization occur in mammals. In this study, centrosome changes during meiosis in horse oocytes, and first cell cycle organization following fertilization, parthenogenesis and nuclear transfer, were monitored. Cumulus oocyte complexes harvested from horse ovaries obtained from slaughtered mares were cultured in vitro. Meiotic oocytes of germinal vesicle (GV), germinal vesicle breakdown (GVBD), metaphase I and II (MI and MII) stages were selected at various set times during in vitro maturation. Embryos at the first cell cycle stage were generated by subjecting MII s...
Relationships of changes in B-mode echotexture and colour-Doppler signals in the wall of the preovulatory follicle to changes in systemic oestradiol concentrations and the effects of human chorionic gonadotrophin in mares. A duplex grey-scale and colour-Doppler ultrasound instrument was used to study the changes in the wall of the preovulatory follicle in mares. When the follicle reached > or =35 mm (hour 0), mares were randomized into control (n = 16) and human chorionic gonadotropin (hCG)-treated (n = 16) groups. The hCG treatment was given at hour 0. Scanning was done every 12 h until hour 36, every hour between hours 36 and 48, and every 12 h thereafter until ovulation. Blood was sampled every 12 h for oestradiol assay. During the period 0-24 h post-treatment, oestradiol concentrations decreased in the hC...
Somatic cell nuclear transfer in horses: effect of oocyte morphology, embryo reconstruction method and donor cell type. The objective of the present work was to investigate and clarify the factors affecting the efficiency of somatic cell nuclear transfer (NT) in the horse, including embryo reconstruction, in vitro culture to the blastocyst stage, embryo transfer, pregnancy monitoring and production of offspring. Matured oocytes, with zona pellucida or after zona removal, were fused to cumulus cells, granulosa cells, and fetal and adult fibroblasts, and fused couplets were cultured in vitro. Blastocyst development to Day 8 varied significantly among donor cells (from 1.3% to 16%, P < 0.05). In total, 137 nuclear...
Systemic concentrations of hormones during the development of follicular waves in mares and women: a comparative study. Changes in systemic concentrations of FSH, LH, oestradiol and progesterone during the ovulatory follicular wave were compared between 30 mares and 30 women. Based on a previous study, the emergence of the future ovulatory follicle was defined as occurring at 13.0 mm in mares and 6.0 mm in women, and deviation in diameter between the two largest follicles was expected to begin at 22.7 mm in mares and 10.3 mm in women. Mean FSH concentrations were high in mares during the luteal phase, resulting from statistically identified FSH surges occurring in individuals on different days and in different ...
Temporal and spatial associations of oestrogen receptor alpha and progesterone receptor in the endometrium of cyclic and early pregnant mares. Uterine function is primarily controlled by the combined actions of oestrogen and progesterone working through their cognate nuclear receptors. The mechanism of establishment of pregnancy in the mare is of interest because it involves prolonged pre-attachment and conceptus migration phases, and both invasive and non-invasive placental cell types, and as such has been an important comparative model. This study characterised regulation of oestrogen (ER) and progesterone (PR) receptors in the endometrium of the mare during the oestrous cycle and early pregnancy. Endometrial tissues collected duri...
Effect of cryopreservation on the cellular integrity of equine embryos. Horse embryos are rarely cryopreserved in practice because expanded blastocysts tolerate freezing poorly, and the embryo begins expanding very soon after entering the uterine cavity. This study examined the effects of freezing on cytoskeleton integrity, and investigated whether cell damage could be reduced using trypsin to thin the blastocyst capsule or cytochalasin-B (cyto-B) to stabilise the cytoskeleton. Sixty-nine embryos were recovered 7 days after ovulation and equilibrated in 10% glycerol, with or without pretreatment with 0.2% trypsin or 7.5 microg/ml cyto-B. Forty-two of the embryos w...