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Topic:Reproduction
Reproduction in horses encompasses the biological processes and mechanisms involved in the breeding and development of equine offspring. This includes the study of reproductive anatomy, physiology, and endocrinology in both mares and stallions. Key areas of interest include the estrous cycle, ovulation, conception, gestation, and parturition. Researchers also examine factors influencing fertility, reproductive technologies such as artificial insemination and embryo transfer, and management practices that impact reproductive success. This page compiles peer-reviewed research studies and scholarly articles that explore the physiological, genetic, and environmental aspects of equine reproduction.
Disorders of sexual development in the domestic horse, Equus caballus. Abnormalities of sexual development causing infertility in horses have been investigated since the early 1970's. Conventional cytogenetic analysis by karyotyping has been the primary tool used to investigate these horses. Abnormalities have a broad range, from a phenotypically normal mare with gonadal dysgenesis to a horse with ambiguous external genitalia and internal male and female organs. Cytogenetic analysis can determine genetic sex but cannot identify mutations or deletions of genes involved in the sex determination pathway. Molecular technologies have been developed to confirm cytogene...
Influence of the uterine environment on the development of in vitro-produced equine embryos. The necessity for early interaction between the embryo and the oviductal and/or uterine environment in the horse is reflected by several striking differences between equine embryos that develop in vivo and those produced in vitro. Better understanding of the salient interactions may help to improve the efficiency of in vitro equine embryo production. In an initial experiment, cleavage-stage in vitro-produced (IVP) equine embryos were transferred into the uterus of recipient mares that had ovulated recently to determine whether premature placement in this in vivo environment would improve subse...
Consequences of butylated hydroxytoluene in the freezing extender on post-thaw characteristics of stallion spermatozoa in vitro. Ejaculates from six pure Spanish stallions were split, and one subsample frozen in a commercial extender supplemented with the lipid soluble antioxidant butylated hydroxytoluene (BHT), while the other subsample served as control. After at least 4 weeks of storage, samples were thawed and post-thaw sperm quality analysed: sperm motility and kinematics using a CASA system, membrane and acrosome integrity and mitochondrial membrane potential using flow cytometry. The outcome of cryopreservation varied significantly among stallions. However, the supplementation with 1 mm BHT had no significant eff...
GTG mutation in the start codon of the androgen receptor gene in a family of horses with 64,XY disorder of sex development. Genetic sex in mammals is determined by the sex chromosomal composition of the zygote. The X and Y chromosomes are responsible for numerous factors that must work in close concert for the proper development of a healthy sexual phenotype. The role of androgens in case of XY chromosomal constitution is crucial for normal male sex differentiation. The intracellular androgenic action is mediated by the androgen receptor (AR), and its impaired function leads to a myriad of syndromes with severe clinical consequences, most notably androgen insensitivity syndrome and prostate cancer. In this paper, w...
Cloning of equine herpesvirus type 1 438/77 strain genome as a bacterial artificial chromosome. Equine herpesvirus type 1 (EHV-1) is a major cause of respiratory and reproductive diseases in horses worldwide. The genome of EHV-1 strain 438/77 (isolated from an aborted equine fetus) was cloned as a bacterial artificial chromosome (BAC) in E. coli without any gene deletions. The mini-F plasmid sequence was inserted in the middle of ORF19 and 20 via homologous recombination following co-transfection of viral DNA and plasmid pE19_20/HA into RK13 cells. Circular viral DNA was extracted from RK13 cells infected with purified recombinant virus expressing green fluorescent protein (GFP) and elec...
Inbreeding in the Thoroughbred horse. Changes in the inbreeding coefficient, F, in the Thoroughbred horse over the past 45 years have been investigated by genotyping 467 Thoroughbred horses (born between 1961 and 2006) using the Illumina Equine SNP50 bead chip, which comprises 54,602 SNPs uniformly distributed across the equine genome. The Spearman rank correlation coefficient, r, between the year of birth and F was estimated. The results indicate that inbreeding in Thoroughbreds has increased over the past 40 years, with r = 0.24, P < 0.001 demonstrating that there is a highly significant, though relatively weak correlation be...
Harnessing the genetic toolbox for the benefit of the racing Thoroughbred. The understanding and application of genetics have grown extremely quickly since it has become possible to sequence the whole genome of an organism. The human genome sequence was completed in 2001 and that of the horse in 2007. The significance of this is that it makes it more feasible to explain how both genetically simple and complex traits are transmitted from one generation to the next and, therefore, to make informed breeding decisions, modify how horses are managed and trained to minimise the risk of disease and injury, and improve methods of prevention, diagnosis and treatment of many c...
Nucleolar organizer regions (NORs) distribution and behavior in spermatozoa and meiotic cells of the horse (Equus caballus). Nucleolar organizing regions (NORs) containing rDNA gene clusters have been assigned to the equine autosomes ECA1, ECA28, and ECA31. Active NORs (Ag-NORs) are associated with argyrophilic proteins, which allow them to be readily identified using silver staining techniques. Fluorescence in situ hybridization (FISH) for rDNA can also be used to visualize all NOR clusters in the nucleus, regardless of whether they are active or inactive. The present study analyzed the distribution and behavior of equine Ag-NOR and NOR clusters in horse spermatozoa and during male meiosis by FISH and silver staini...
In vitro acute exposure to DEHP affects oocyte meiotic maturation, energy and oxidative stress parameters in a large animal model. Phthalates are ubiquitous environmental contaminants because of their use in plastics and other common consumer products. Di-(2-ethylhexyl) phthalate (DEHP) is the most abundant phthalate and it impairs fertility by acting as an endocrine disruptor. The aim of the present study was to analyze the effects of in vitro acute exposure to DEHP on oocyte maturation, energy and oxidative status in the horse, a large animal model. Cumulus cell (CC) apoptosis and oxidative status were also investigated. Cumulus-oocyte complexes from the ovaries of slaughtered mares were cultured in vitro in presence of...
Relationship between dose of cloprostenol and age of corpus luteum on the luteolytic response of early dioestrous mares: a field study. The objective of this study was to establish and characterize the relationship between the dose of cloprostenol (37.5, 250, 500 and 750 μg) and the age of the early corpus luteum (CL) (80, 88, 96, 104 and 112 h) on the luteolytic response of mares. Behavioural oestrus and ultrasonographic signs of return to oestrus were considered as the occurrence of full luteolysis. A total of 298 mares were divided into groups according to dose of cloprostenol and CL age. There was an effect of dose of cloprostenol (p < 0.001) and age of the CL at the time of treatment (p 0.05); and that of 500 similar...
Unilateral and bilateral laparoscopic ovariectomy in 157 mares: a retrospective multicenter study. To evaluate surgical techniques, perioperative complications and outcome of laparoscopic surgery for unilateral and bilateral ovariectomy in mares. Methods: Retrospective multicenter case series. Methods: Mares (n = 157). Methods: Indications for surgery were neutering, fertility problems, elimination of hormone-related abnormal behavior, ovarian tumors as well as nonregressive ovarian hematomas or cysts. Ovariectomy (n = 206) were performed by a standard laparoscopic procedure in 157 standing sedated mares. Dissection and hemostasis were achieved by using either (1) a linear stapling device, ...
Equine pre-implantation conceptuses express neuraminidase 2–a potential mechanism for desialylation of the equine capsule. During the second and third week of pregnancy, the equine conceptus is covered by an acellular glycoprotein capsule. This capsule contains glycoproteins resembling those of the mucin family with sialic acid making up a high proportion of the carbohydrate. Coinciding with conceptus fixation, a marked decline in sialic acid content of the capsule occurs, which has been proposed to contribute to cessation of conceptus mobility. Herein, we describe the expression of neuraminidase 2 (NEU2) by pre-implantation stages of equine conceptus development. NEU2 transcript abundance was examined in conceptu...
Molecular and cytogenetic studies in a case of XX SRY-negative sex reversal in an Arabian horse. An 18-month-old Arabian foal characterized by a stallion-like appearance was submitted for cytogenetic and molecular genetics examinations due to abnormalities of external genitalia and the presence of ovotestis-like structures in the abdominal cavity. By RB-banding the animal showed the normal female equine karyotype (2n = 64,XX). Molecular analysis revealed the absence of the SRY and ZFY genes and the presence of ZFX, a typical female equine condition. The entire RSPO1 coding region was examined to exclude its involvement. Although a SNP was found in exon 3, it was not responsible for an ami...
Replacing egg yolk with soybean lecithin in the cryopreservation of stallion semen. The objective of this study was to determine whether replacing the egg yolk with soybean lecithin in the Botu-Crio® cryodiluent would maintain the fertility of cryopreserved stallion sperm. Two experiments were performed to evaluate cell freezability. In experiment 1, sperm from 15 stallions were frozen in Botu-Crio® (BC) or Botu-Crio® which contained 45g/L soybean lecithin (BCLS45) in place of the egg yolk. In experiment 2, we compared different concentrations of soybean lecithin: 0, 10.0, 12.5, 15.0, 17.5 and 20.0g/L (BC, BCLS10, BCLS12.5, BCLS17.5 and BCLS20, respectively). In experiment...
Cytogenetic and molecular characterization of Y isochromosome in a 63XO/64Xi(Yq) mosaic karyotype of an intersex horse. Sex chromosome aberrations commonly lead to abnormal sexual development. Here we cytogenetically and molecularly characterized Y isochromosome in an intersex horse. Blood lymphocyte analysis showed a mosaic karyotype with 96% 63,XO and 4% 64,Xi(Y) cells. Molecular analysis of the isochromosome was carried out by fluorescence in situ hybridization and polymerase chain reaction with male-specific and pseudoautosomal markers from the horse Y chromosome. We found that the isochromosome was monocentric, composed of 2 long arms, carrying 2 sets of genes of the pseudoautosomal region (PAR) and the ma...
Manual reduction of multiple embryos in the mare: the effect on subsequent pregnancy outcome. To determine whether manually reduced multiple pregnancies (MPs) are at a greater risk of pregnancy loss than single pregnancy (SP) in mares, and to examine if a difference exists in the timing of pregnancy loss between manually reduced MPs and SPs, 1916 Thoroughbred mares were ultrasonically monitored every 2 days during oestrus to confirm ovulation, and up to Day 40 post-ovulation to confirm pregnancy. Ultimate pregnancy outcome was ascertained from the General Thoroughbred Stud book and classified as live foal (LF), early abortion (EA; Days 40-150), slipped foal (SF; Days 150-term), and bar...
Equine fetal sex determination using circulating cell-free fetal DNA (ccffDNA). In this study, polymerase chain reaction (PCR) reamplification of the first PCR product (2nd-PCR) and a qPCR assay were used to detect the sex determining region Y (SRY) gene from circulating cell-free fetal DNA (ccffDNA) in blood plasma of pregnant mares to determine fetal sex. The ccffDNA was isolated from plasma of 20 Thoroughbred mares (5-13 y old) in the final 3 mo of pregnancy (fetal sex was verified after foaling). For controls, plasma from two non-pregnant mares and two virgin mares were used, in addition to the non-template control. The 182 bp nucleotide sequence corresponding to the ...
Hormone concentration changes temporally associated with the hour of transition from preluteolysis to luteolysis in mares. The temporal associations of cortisol, estradiol-17β, and oxytocin with pulses of PGFM at the common hour of transition between preluteolysis and luteolysis was studied in plasma from hourly blood samples in mares (n=8). The transitional hour was determined from progesterone concentrations and occurred between 2PM and 2AM in all mares. Pulses of PGFM were grouped into those occurring at the last pulse of preluteolysis (preluteolytic pulse), at the hour of transition (transitional), and during luteolysis (luteolytic). The preluteolytic PGFM pulse (45±16pg/ml at peak) and transitional pulse (4...
Dose-response effects of estrogenic mycotoxins (zearalenone, alpha- and beta-zearalenol) on motility, hyperactivation and the acrosome reaction of stallion sperm. The aim of this study was to investigate the in vitro effects of the Fusarium fungus-derived mycotoxin, zearalenone and its derivatives alpha-zearalenol and beta-zearalenol on motility parameters and the acrosome reaction of stallion sperm. Since the toxic effects of zearalenone and its derivatives are thought to result from their structural similarity to 17beta-estradiol, 17beta-estradiol was used as a positive control for 'estrogen-like' effects. Methods: Stallion spermatozoa were exposed in vitro to zearalenone, alpha-zearalenol, beta-zearalenol or 17beta-estradiol at concentrations ranging...
The comparison of equine articular cartilage progenitor cells and bone marrow-derived stromal cells as potential cell sources for cartilage repair in the horse. A chondrocyte progenitor population isolated from the surface zone of articular cartilage presents a promising cell source for cell-based cartilage repair. In this study, equine articular cartilage progenitor cells (ACPCs) and equine bone marrow-derived stromal cells (BMSCs) were compared as potential cell sources for repair. Clonally derived BMSCs and ACPCs demonstrated expression of the cell fate selector gene, Notch-1, and the putative stem cell markers STRO-1, CD90 and CD166. Chondrogenic induction revealed positive labelling for collagen type II and aggrecan. Collagen type X was not detec...
Ultrasound characteristics of experimentally induced luteinized unruptured follicles (LUF) and naturally occurring hemorrhagic anovulatory follicles (HAF) in the mare. The development of hemorrhagic anovulatory follicles (HAF) involves luteinization and hemorrhage of the follicle. This is observed on ultrasound as an increase in the echogenicity of the granulosa layer and formation of echoic particles in the antrum. The inhibition of prostaglandin synthesis with flunixin meglumine (FM) during the periovulatory period induces ovulatory failure with development of luteinized unruptured follicles (LUF). These two types of anovulatory follicles appear to share similar ultrasound features but they have not been compared critically. The following endpoints: follic...
Ultrasonographic features of the mule embryo, fetus and fetal-placental unit. The aim of this study was to establish baseline ultrasound data concerning the mule conceptus during gestation. Ten multiparous Trotter mares were artificially inseminated with chilled semen from an Amiatino jack donkey. Daily transrectal ultrasonography was carried out from the day of ovulation until Day 50 of gestation to determine the following: first detection of the embryonic vesicle (EV), mobility phase, EV diameter, day of EV fixation, changes in EV shape, date of yolk sac regression and embryo crown-rump length. Monthly ultrasonic assessments from Day 50 of gestation to term were carri...
Effects of nutritional cues on the duration of the winter anovulatory phase and on associated hormone levels in adult female Welsh pony horses (Equus caballus). Mares have an annual reproductive rhythm, with a phase of inactivity in midwinter. The aim of this study was to determine the impact of food restriction on physiological and metabolic hallmarks of this rhythm. Methods: Over three successive years, 3 groups of 10 mares were kept under natural photoperiod. A 'well-fed' group was fed to maintain the mares in good body condition; a 'restricted' group received a diet calculated to keep the mares thin and a 'variable' group was fed during some periods like the 'restricted' group and during some other periods like the 'well-fed' group, with the aim o...
Uterine artery blood flow remains unchanged in pregnant mares in response to short-term administration of pentoxifylline. The objective of this study was to use Doppler ultrasound technology to determine whether pentoxifylline administration increased uterine blood flow in normal pregnant pony mares. Thirteen pregnant pony mares between 18 and 190 d of gestation (mean ± SEM, 101 ± 55) were utilized for the study during two trial periods. In each trial, pentoxifylline (17 mg/kg by mouth every 12h, diluted in syrup) was administered to half of the mares for 3 d, while the other mares were treated with syrup only. Doppler measurements were obtained from the right and left uterine arteries from each mare for 2 d pr...
Distribution of mast cells in vaginal, cervical and uterine tissue of non-pregnant mares: investigations on correlations with ovarian steroids. Mast cells constitute part of the cellular immune system of the genital tract. They play a potential role in cervical remodelling during parturition and are subject to the influence of ovarian steroids. In this study, the influence of oestradiol-17β and progesterone on the distribution of mast cells in equine vaginal, cervical and uterine tissue was evaluated. Genital tracts were retrieved from healthy mares at a local slaughterhouse. The cervix was divided along the longitudinal axis into five equivalent regions from cranial to caudal (R1 to R5). Toluidine blue staining was used for the dete...
The effects of an advanced uterine environment on embryonic survival in the mare. During embryo transfer (ET) the equine embryo can tolerate a wide degree of negative asynchrony but positive asynchrony of >2 days usually results in embryonic death. There is still confusion over whether this is due to the inability of the embryo to induce luteostasis or to an inappropriate uterine environment. Objective: To assess embryo survival and development in an advanced uterine environment. Objective: Embryo-uterine asynchrony, not the embryo's inability to induce luteostasis, is responsible for embryonic death in recipient mares with a >2 days chronologically advanced uterus. Methods...
A pilot comparison of laser-assisted vs piezo drill ICSI for the in vitro production of horse embryos. Intracytoplasmic sperm injection (ICSI) is the method of choice for the in vitro production (IVP) of equine embryos. However, conventional ICSI has been associated with mechanical damage to the oocyte caused by the deformation of the zona pellucida (ZP) and exposure of the oolemma to negative pressure during injection. Introduction of the less traumatic and more efficient piezo drill-assisted ICSI (PDAI) yielded higher cleavage rates and more consistent results. Nevertheless, PDAI is also associated with disadvantages such as the use of mercury and possible DNA damage. This led us to explore a...
Comparison of transvaginal natural orifice transluminal endoscopic surgery (NOTES®) and laparoscopy for elective bilateral ovariectomy in standing mares. To compare surgical trauma, perioperative pain, surgical time, and complication rate in mares undergoing standing bilateral ovariectomy by natural orifice transluminal endoscopic surgery (NOTES) or laparoscopy. Methods: Experimental study. Methods: Healthy mares (n = 12). Methods: Ovariectomy was performed with a vessel-sealing device by transvaginal NOTES (n = 6) using a flexible endoscope and specialized instruments or by bilateral flank laparoscopy (n = 6). Preoperative and postoperative complete blood count (CBC), plasma fibrinogen concentration, serum amyloid A concentration, peritoneal n...
