Fertility in horses encompasses the physiological processes and factors influencing reproductive success in equine species. It involves the study of reproductive anatomy, endocrinology, and behavior, as well as the management practices that affect breeding outcomes. Key aspects include the estrous cycle, conception rates, and factors impacting stallion and mare fertility. Reproductive technologies such as artificial insemination, embryo transfer, and hormonal therapies are also explored to enhance breeding efficiency. This page compiles peer-reviewed research studies and scholarly articles that examine the biological mechanisms, management strategies, and technological advancements related to equine fertility.
Carneiro JAM, Canisso IF, Bandeira RS, Scheeren VFC, Freitas-Dell'Aqua CP, Alvarenga MA, Papa FO, Dell'Aqua JA.This study aimed to evaluate the antioxidant properties of coenzyme Q10 (CoQ10) during cryopreservation of semen obtained from stallions having good and bad semen freezing ability (GFA vs. BFA, respectively). Forty ejaculates (n = 20 stallions) were split into five centrifugation and five freezing extenders containing different concentrations of CoQ10 (0, 25, 50, 75 and 100 μmols/L). If CoQ10 was added to the centrifugation extender, the freezing extender had no CoQ10 added; similarly, if CoQ10 was added to the freezing extender, the centrifugation extender had no CoQ10. Semen cryoprese...
Ginther OJ, Domingues RR, Dangudubiyyam SV, Araujo ER.Diameter deviation or follicle selection during a follicular wave begins with continuous growth rate of F1 (dominant or selected follicle; DF) and decreasing growth rate of F2 (largest subordinate). Intraovarian patterns based on presence or absence of the DF and CL are DF-CL, DF, CL, and devoid (neither DF nor CL). The DF and CL relationships in a pair of ovaries are ipsilateral (DF-CL pattern and devoid pattern) and contralateral (DF pattern and CL pattern). The effects of patterns and relationships on F1, F2, and FSH during deviation were determined in 21 mares. Diameters of F1 and F2 at ex...
Consuegra C, Crespo F, Bottrel M, Ortiz I, Dorado J, Diaz-Jimenez M, Pereira B, Hidalgo M.The aim of this study was to assess different concentrations of sucrose-based extenders combined with bovine serum albumin (BSA) as an alternative to stallion sperm cryopreservation with permeable cryoprotectants. Semen samples (n = 16) were collected from six stallions. Sperm was cooled, filled in 0.5 mL straws and frozen in nitrogen vapor. Post-thaw sperm kinetic parameters, plasma and acrosome membrane integrity were statistically compared among treatments. In Experiment 1, extenders containing 1% of BSA and different concentrations of sucrose (mmol/L, M): 0, 50, 100, 250, 350 and 450...
Jeannerat E, Marti E, Berney C, Janett F, Bollwein H, Sieme H, Burger D, Wedekind C.The major histocompatibility complex (MHC) has repeatedly been found to influence mate choice of vertebrates, with MHC-dissimilar mates typically being preferred over MHC-similar mates. We used horses (Equus caballus) to test whether MHC matching also affects male investment into ejaculates after short exposure to a female. Semen characteristics varied much among stallions. Controlling for this variance with a full-factorial within-subject experimental design, we found that a short exposure to an MHC-dissimilar mare enhanced male plasma testosterone and led to ejaculates with elevated sperm nu...
Bromfield JJ.Artificial insemination has been a landmark procedure in improving animal agriculture over the past 150 years. The utility of artificial insemination has facilitated a rapid improvement in animal genetics across agricultural species, leading to improvements of growth, health and productivity in poultry, swine, equine and cattle species. The utility of artificial insemination, as with all assisted reproductive technologies side-steps thousands of years of evolution that has led to the development of physiological systems to ensure the transmission of genetics from generation to generation. The ...
Al-Essawe EM, Johannisson A, Wulf M, Aurich C, Morrell JM.Addition of seminal plasma (SP) prior to cryopreservation may influence stallion sperm cryosurvival. The objective of this study was to investigate the addition of pooled SP from "good" or "bad" freezer stallions to spermatozoa selected by single layer centrifugation (SLC) prior to cryopreservation on post-thaw sperm quality. Semen from 12 stallions was collected; 5 mL was frozen as control (C) and the remainder was processed by SLC to remove SP and was divided into three aliquots: i) SLC sample without SP (SLC); ii) SLC plus pooled SP from "good freezer" stallions (SLC-GF); iii) SLC plus po...
Martins HS, da Silva GC, Cortes SF, Paes FO, Martins Filho OA, Araujo M, Stahlberg R, Lagares MA.During cryopreservation, sperm was submitted to an increase in reactive oxygen species generation. This work aimed to improve the quality of frozen equine sperm after the addition of antioxidants lactoferrin (Lf) and catalase (Cat) to a freezing extender. Semen from six stallions was frozen with the extenders: F1) control, INRA 82 freezing extender, F2) F1 + 500 μg/ml Lf and F3) F1 + 200 IU/ml Cat. After thawing, sperm motility parameters, membrane functionality and integrity, and acrosome integrity and spontaneous acrosome-reacted sperm were evaluated with a computer-assisted sperm anal...
Ewert M, Lüders I, Böröcz J, Uphaus H, Distl O, Sieme H.The aim of the present study was to analyze the effects of stallion and mare, their ages, and maternal lineage on the gestation length (GL) in Thoroughbreds. In addition, additive genetic effects of the dam, stallion and fetus were analyzed. Data were taken from 1993 through 2009, and included 16,226 pregnancies from 5959 Th oroughbred mares mated with 290 different stallions. All analyses were performed using linear mixed models. The GL ranged from 306 to 390 days, with a mean length of 347.0 ± 14.4 days. Mating of mares with stallions aged 17 years and older resulted in a significantly ...
Pérez-Marín CC, Requena FD, Arando A, Requena L, Requena F, Agüera EI.The aim of this study was to determine the effects of various abiotic factors, such as light, physical stress (pipetting) and thermal shock, on the quality of fresh and cooled equine sperm. In experiment I, four sperm aliquots were subjected to different light exposures: (i) protected control samples (CTRL), (ii) exposed to UV light at 10 cm (UV10), (iii) exposed to UV light at 20 cm (UV20) and (iv) exposed to laboratory lighting (LAB). In experiment II, four semen aliquots were subjected to repeated pipetting for 0, 10, 20 and 30 times (CTRL, P10, P20 and P30, respectively). In experiment I...
Macedo S, Bliebernicht M, Carvalheira J, Costa A, Ribeiro F, Rocha A.Glycerol-based extenders are widely utilized for freezing equine semen, but media combining methylformamide may better preserve sperm motility and mitochondrial function. Semen is cryopreserved utilizing either a Styrofoam box filled with liquid nitrogen or an automatic freezer. The objective of this experiment was to compare the post-thaw characteristics of the same ejaculates cryopreserved in a Styrofoam box or in an automatic freezer, utilizing a glycerol-based extender (Gent) and an extender that combines methylformamide and glycerol (BotuCrio ). For that, one ejaculate from 30 stallions c...
Del Prete C, Tafuri S, Ciani F, Pasolini MP, Ciotola F, Albarella S, Carotenuto D, Peretti V, Cocchia N.Stallion semen is damaged by oxidative stress during cooling and transport. Semen processing and extenders have been tested to improve the fertilizing capacity of semen and to preserve semen during transport. Dietary supplementation with natural antioxidants has been proposed to prevent oxidative damages. In this study, for the first time, the effect of dietary supplementation with Lepidium meyenii (Maca) on the characteristics of fresh and chilled stallion semen was evaluated. Maca is a traditional Andean crop used as a nutraceutical for the fertility-enhancing properties that are linked with...
Herrera M, Herrera JM, Cantatore S, Aguilar J, Felipe A, Fumuso E.Uterine acute post-breeding inflammation is a physiological tissue response to the entry of exogenous elements, with persistent endometritis being the main pathology responsible for subfertility in the mare (Equus ferus caballus; Linnaeus, 1758). Mares can be classified as susceptible or resistant to endometritis according to their ability to remove intrauterine fluid within 48 hr after experimental inoculation. Endometrial biopsy is a technique that is commonly used to establish the degree of lesions that can affect the fertility of the mare. Endometrial histomorphometry is an objective and ...
Canesin HS, Brom-de-Luna JG, Choi YH, Pereira AM, Macedo GG, Hinrichs K.Previous studies have found low rates of blastocyst development (0-11%) after vitrification of germinal vesicle (GV)-stage equine oocytes. In this study, we systematically evaluated a short (non-equilibrating) system for GV-stage oocyte vitrification. In Exp. 1, we assessed oocyte volume in cumulus-oocyte complexes (COCs) exposed to components of a short protocol, using 2% each of ethylene glycol and propylene glycol in the first solution (VS1); 17.5% of each plus 0.3 M trehalose in the second solution (VS2); and fetal bovine serum as the base medium. Based on the time to oocyte minimum volu...
Oliveira Neto IV, Canisso IF, Segabinazzi LG, Dell'Aqua CPF, Alvarenga MA, Papa FO, Dell'Aqua JA.This study compared hormone treatments given to mares during anestrus, spring transition, and different stages of the estrous cycle, by assessing uterine features and pregnancy rates after embryo transfer (ET). Embryo recipient mares (n = 160) were equally arranged as follows: G1-spontaneous ovulation (control), G2-anestrus, G3-spring transition, G4-early estrus, G5-estrus, G6-diestrus, G7-early diestrus treated with a dose of dinoprost, and G8-early diestrus treated with two doses of dinoprost. At treatment initiation (Day-4), G2-7 were given dinoprost and estradiol-17β, thereafter, estr...
Diaz-Jimenez M, Dorado J, Ortiz I, Consuegra C, Pereira B, Gonzalez-De Cara CA, Aguilera R, Mari G, Mislei B, Love CC, Hidalgo M.The aim of this study was to evaluate the effect of different concentrations of sucrose combined with bovine serum albumin (BSA), as non-permeable cryoprotectants, on donkey sperm parameters after cryopreservation, in comparison to a control extender containing glycerol. Semen from five Andalusian donkeys (n = 12) were centrifuged and resuspended with a commercial extender for equine sperm (Gent A, Minitube) adding 1% BSA and different concentrations (M, mol/l) of water-diluted sucrose: 0.05, 0.1, 0.25, 0.35 and 0.45. Thereafter, semen (n = 24) were diluted in the same base extender co...
Miyakoshi D, Sato F, Murase H, Inoue Y, Shikichi M, Iwata K, Okai K, Ito K, Nambo Y.The purpose of this study was to evaluate foaling rates of mares that were rebred after the pregnancy loss in same reproductive season and to examine factors influencing them in Hidaka, Japan. The study included 82 Thoroughbred mares that had experienced pregnancy loss. The foaling rate of the mares that were rebred after pregnancy loss was 57.3%. The foaling rate decreased as the period until detection of pregnancy loss increased. Aging and lower body condition score of mares decreased the foaling rate.
Satoh M, Higuchi T, Inoue S, Gotoh T, Murase H, Nambo Y.This is a retrospective study of uterine torsion (UT) in seven mares. In two cases, serum progesterone and estradiol concentrations were also investigated. The mare and foal/fetus survival rate was 57% (4/7). Four cases presented with clockwise torsion, and two cases presented with counterclockwise torsion. The direction was undetermined in one case. The degree of torsion varied. Correction of torsion was performed by ventral midline celiotomy in all cases. In the two cases with measured hormone levels, elevated levels of P were decreased through medication. All mares discharged from the hospi...
Gil MC, Ferrusola CO, Anel-López L, Ortiz-Rodriguez JM, Alvarez M, de Paz P, Anel L, Peña FJ.Spermatozoa undergo apoptotic changes during the cryopreservation process. These changes, recently termed spermptosis, resemble the cryopreservation induced delayed onset of cell death observed after thawing of somatic cells. Due to its importance in cryobiology, methods to easily identify spermptotic cells are warranted. In this study, a well-validated method for identification of spermatozoa with caspase 3 activity was compared with use of the combination of Hoechst 33342 (H-42) and ethidium homodimer (Eth-1). Live, dead and apoptotic spermatozoa assessed with each method were compared using...
Burger D, Thomas S, Aepli H, Dreyer M, Fabre G, Marti E, Sieme H, Robinson MR, Wedekind C.Genes of the major histocompatibility complex (MHC) have been shown to influence social signalling and mate preferences in many species, including humans. First observations suggest that MHC signalling may also affect female fertility. To test this hypothesis, we exposed 191 female horses () to either an MHC-similar or an MHC-dissimilar stimulus male around the time of ovulation and conception. A within-subject experimental design controlled for non-MHC-linked male characteristics, and instrumental insemination with semen of other males ( = 106) controlled for potential confounding effects of ...
Inoue Y, Sekiguchi M.Therapeutic techniques for oviductal obstruction in the mare are limited. Nonsurgical and retrograde flushing may be an attractive alternative to current treatment methods for oviductal blockage. Objective: To evaluate hysteroscopic selective hydrotubation as a treatment option for presumptive equine oviductal blockage. Methods: Retrospective case series. Methods: A quantity of 10 mL of saline was flushed through the oviducts in 28 standing sedated mares, which had reproductive histories of unexplained subfertility, by inserting a catheter into the uterotubal junction under endoscopic guidance...
Murcia-Robayo RY, Jouanisson E, Beauchamp G, Diaw M.Evaluation of sperm morphology is part of the assessment of fertility in human and animal reproduction. Analyses can be performed using different techniques, including the use of staining methods In our prospective study, the morphology of equine sperm was evaluated using 3 staining methods Diff-Quik, eosin-nigrosin, and SpermBlue, the latter being a relatively new stain commonly used for human sperm. Our hypotheses were that (1) SpermBlue allows easier assessment of horse sperm morphology and facilitates better identification of sperm abnormalities, and (2) sperm morphology classification dif...
Cabrera T, Ramires-Neto C, Belaz KRA, Freitas-Dell'aqua CP, Zampieri D, Tata A, Eberlin MN, Alvarenga MA, Souza FF.The membrane of spermatozoa, which contributes to cellular cryoresistance, contains numerous lipids with a composition that directly affects membrane fluidity and the fertilization process. In light of variations in the degree of sensitivity in equine seminal freezing, this study aimed to correlate equine semen lipids with post-thawing characteristics of spermatozoa. We used ejaculates from 34 stallions, which were evaluated (total motility ≥ 60%), frozen and thawed and reevaluated for motility of spermatozoa, membrane integrity and lipid peroxidation. Lipid extraction of the fresh semen s...
Shojaeian K, Nouri H, Kohram H.Overproduction of reactive oxygen species during sperm freeze-thawing process leads to membrane lipid peroxidation, DNA damage, motility loss, and subsequent death. This oxidative stress can be alleviated by the addition of some antioxidants to semen extenders prior to freezing. This study was performed to evaluate the in vitro effectiveness of MnTBAP (a cell permeable antioxidant) on stallion sperm freezability and in vivo fertility rate. Twenty-one ejaculates were, collected with missouri model artificial vagina (n = 3 stallions, seven ejaculate each), and diluted (1:2 v/v) with phosphoc...
Oldenhof H, Zhang M, Narten K, Bigalk J, Sydykov B, Wolkers WF, Sieme H.Nonviable freeze-dried sperm have intact chromatin and can be used for fertilization via intracytoplasmic sperm injection. Freeze-dried sperm preferably should be stored at 4°C or lower, because DNA damage accumulates during storage at room temperature. Disaccharides are known to protect biomolecules both during freezing and drying, by forming a glassy state. Their use is challenging because cellular membranes are normally impermeable for disaccharides. In the current study, we demonstrate that membrane impermeable compounds, including lucifer yellow and trehalose, are taken up by stallion sp...
Allen WR, Wilsher S.Over the past 50 years, per season pregnancy rate in Thoroughbred mares have risen from 70 to >90% and foaling rates from 55 to >80%. Some of the significant pharmacological treatments and diagnostic methods which have driven this marked improvement in breeding efficiency are reviewed. They include the application of artificial lighting to hasten the onset of ovulatory oestrous cyclicity in early Spring, rapid steroid hormone assays to aid in determining the stage and normality of the mare's cycle, prostaglandin analogues, synthetic progestagens and Gonadotrophin-releasing Hormone (GnRH) anal...
Rui BR, Angrimani D, Bicudo LC, Losano J, Nichi M, Pereira R.Sperm DNA fragmentation is a condition that interferes directly in the reproductive efficiency. Currently, there are several methods for assessing the sperm DNA integrity, such as Alkaline Comet, TUNEL and Sperm Chromatin Structure Assay. However, many of these techniques are laborious and require high-precision equipment. Thus, the development of new techniques can optimize the evaluation of sperm DNA damage. Therefore, the aim of this study was to standardize the toluidine blue (TB) stain technique for the analysis of DNA fragmentation of dog, cat, bull, stallion and ram spermatozoa. For thi...
Bonilla-Correal S, Noto F, Garcia-Bonavila E, Rodríguez-Gil JE, Yeste M, Miro J.Aquaporins (AQPs) are transmembrane proteins found in all cells and are responsible for the transport of water and small solutes. While these proteins have been found in the spermatozoa of humans, rodents, pigs and cattle, where not only do they play a role for the regulation of sperm volume but are also related with the sperm resilience to withstand freeze-thawing procedures, their presence in stallion sperm is yet to be reported. Therefore, the objectives of this work were as follows: (i) to determine whether AQP3, AQP7 and AQP11 are present in stallion sperm and (ii) to investigate whether ...
Müller-Unterberg M, Wallmann S, Distl O.The Black Forest Draught horse (BFDH) is an endangered German coldblood breed with its origin in the area of the Black Forest in South Germany. In this retrospective study, the influence of the inbreeding coefficient on foaling rates was investigated using records from ten breeding seasons. Due to the small population size of BFDH, the level of inbreeding is increasing and may have an effect on foaling rates.The data of the present study included all coverings reported for 1024 BFDH mares in the years 2001-2009. These mares were covered by 32 BFDH stallions from the State Stud Marbach. Data fr...
Wynn MAA, Esteller-Vico A, Legacki EL, Conley AJ, Loux SC, Stanley SD, Curry TE, Squires EL, Troedsson MH, Ball BA.During the latter half of gestation in mares, there is a complex milieu of pregnanes in peripheral blood. Progesterone concentrations are often assessed by immunoassay during late gestation as a measure of pregnancy well-being; however, interpretation of results is complicated by the numerous cross-reacting pregnanes present in high concentrations during late gestation. Further, many mares are supplemented with an exogenous progestin, altrenogest, which may also cross-react with existing assays and further confound interpretation. The objectives of this study were: 1) to compare differences in...
Segabinazzi LGTM, Oba E, Alvarenga MA.Equine practitioners often prescribe the combined use of hCG and GnRH to hasten ovulation due to presumed synergistic effects. Therefore, this study aimed to test whether the combination of hCG and deslorelin acetate to hasten ovulation in mares would show any effect in inducing ovulation more efficiently than when either drug is used by itself, and to verify whether this association would affect progesterone concentrations; corpus luteum (CL) diameter and blood flow; and pregnancy outcome in recipient mares after embryo transfer (ET). Seventeen mares had the ovulation hastened (≥35 mm folli...
Consuegra C, Crespo F, Bottrel M, Ortiz I, Dorado J, Diaz-Jimenez M, Pereira B, Hidalgo M.The aim of this study was to assess different concentrations of sucrose-based extenders combined with bovine serum albumin (BSA) as an alternative to stallion sperm cryopreservation with permeable cryoprotectants. Semen samples (n = 16) were collected from six stallions. Sperm was cooled, filled in 0.5 mL straws and frozen in nitrogen vapor. Post-thaw sperm kinetic parameters, plasma and acrosome membrane integrity were statistically compared among treatments. In Experiment 1, extenders containing 1% of BSA and different concentrations of sucrose (mmol/L, M): 0, 50, 100, 250, 350 and 450...
Ball BA.Fertilization rates were similar for normal and subfertile mares, and much of the difference in fertility between normal and subfertile mares was due to embryonic loss. Fertilization rate estimates for mares ranged from 71 to 96 per cent. The incidence of embryonic loss detected by ultrasonography between Days 11 and 50 was approximately 9 per cent for normal mares, and the estimated incidence of embryonic loss before Day 14 was also 9 per cent. Therefore, the estimated incidence of embryonic loss in normal mares between fertilization and Day 50 is approximately 18 per cent (Fig. 1). In subfer...
Troedsson MH, Ababneh MM, Ohlgren AF, Madill S, Vetscher N, Gregas M.The objective of this study was to determine whether periovulatory treatments with PGF2alpha affects the development of the CL, and whether the treatment was detrimental to the establishment of pregnancy. Reproductively sound mares were assigned randomly to one of the following treatment groups during consecutive estrus cycles: 1. 3,000 IU hCG within 24 hours before artificial insemination and 500 microg cloprostenol (PGF2alpha analogue) on Days 0, 1, and 2 after ovulation (n=8), 2. 2 mL sterile water injection within 24 hours before artificial insemination and 500 microg cloprostenol on Days ...
Irvine CH, Alexander SL.Conflicting views exist on the mode of gonadotrophin-releasing hormone (GnRH) secretion during the ovulatory LH surge and the relative importance of changes in pituitary responsiveness to GnRH in generating the LH surge. This disagreement may stem from species differences and/or methodological problems. To provide data on the exact relationship between GnRH and gonadotrophin secretion during the spontaneous LH surge, we collected pituitary venous (PV) blood every 30 s for 3-4 h from eight mares and then assayed GnRH (in six of the mares), FSH and LH. Jugular blood was also collected from twelv...
Flores-Flores G, Velázquez-Cantón E, Boeta M, Zarco L.The effects of repeated cloprostenol administration were compared in mares impregnated by horses and mares impregnated by donkeys in order to assess the role of eCG on the development of pregnancy-associated resistance to the luteolytic and abortifacient effects of PGF2α. Eleven mares impregnated by donkey (mule pregnancy) and 9 mares impregnated by horse (horse pregnancy) were used. Six mares with mule pregnancy and four with horse pregnancy were injected with cloprostenol (0.25 mg) when they were between day 65 and day 75 of pregnancy, and the treatment was repeated 48, 72 and 96 h latter...
Dini P, Bogado Pascottini O, Ducheyne K, Hostens M, Daels P.In the present study, we examined the effect of holding equine oocytes in Syngro embryo holding medium (EHM) overnight at either 4 °C, 17 °C, or 22 °C to 25 °C, on the time to maturation and developmental competence. We also examined the effect of placing denuded oocyte without extruded polar body back in maturation condition on subsequent maturation rate. In experiment 1, cumulus-oocyte complexes (COCs) were recovered postmortem and placed in EHM at 22 °C to 25 °C for 18 to 20 hours (OH) or placed directly in maturation (DM). The maturation rate was assessed after 22, 24, or 28 hou...
Allen WR, Wilsher S, Morris L, Crowhurst JS, Hillyer MH, Neal HN.Mares are occasionally encountered that consistently fail to conceive when inseminated, naturally or artificially, with fertile stallion semen in the absence of any identifiable pathology of either the structure or function of their reproductive tract. Objective: Temporary blockage of the oviducts by accumulations of naturally occurring oviducal masses may be preventing oviducal transport of the embryo to the uterus. Methods: Mares, with known reproductive histories, that had exhibited inexplicable failure of conception were treated by laparoscopically guided administration of PGE2-laced triac...
Bugno M, Jablonska Z, Tischner M, Klukowska-Rötzler J, Pienkowska-Schelling A, Schelling C, Slota E.The aim of our study was to diagnose aneuploidy in equine spermatozoa by multicolour fluorescence in situ hybridization (FISH) technique using specific molecular probes for equine sex chromosomes and autosome pair four (EGFR probe) labeled by different fluorochromes. These were applied on decondensed spermatozoa of four stallions. In total, more than 8800 sperm cells were examined. The total frequency of aberrant cells was 0.496%: aneuploidy of XX (0.135%), YY (0.023%), XY (0.102%), diploidy (0.057%), lack of sex chromosome (0.18%). In one stallion the ratio of normal X- and Y-bearing cells wa...
Donadeu FX, Ginther OJ.The temporal relationships in the changes in concentrations of follicular fluid factors during follicle selection were characterized in mares. All follicles > or =5 mm were ablated 10 days after ovulation, followed by follicular fluid collection from the three largest follicles (F1, F2, and F3) when F1 of the new wave reached a diameter of 8.0-11.9, 12.0-15.9, 16.0-19.9, 20.0-23.9, 24.0-27.9, or 28.0-31.9 mm (n = 4-8 mares/range). Diameter deviation between F1 and F2 began during the 20.0- to 23.9-mm range, as indicated by a greater difference in diameter between the two follicles at the 24...
Jasiński T, Zdrojkowski Ł, Ferreira-Dias G, Kautz E, Juszczuk-Kubiak E, Domino M.Endometrosis is a frequently occurring disease decreasing mares' fertility. Thus, it is an important disease of the endometrium associated with epithelial and stromal cell alterations, endometrium gland degeneration and periglandular fibrosis. Multiple degenerative changes are found in uterine mucosa, the endometrium. However, their pathogenesis is not well known. It is thought that nuclear factor-κB (NF-κB), a cell metabolism regulator, and its activation pathways take part in it. The transcription of the profibrotic pathway genes of the NF-κB in fibrotic endometria differed between the fo...
Dawod A, Miro J, Elbaz HT, Fahmy H, Abdoon AS.This study was designed to examine the effect of the intrauterine infusion of platelet-rich plasma (PRP) or equine lyophilized growth factor (L-GFequina) on the follicular growth, endometrial thickness, estrus cycle length, and pregnancy rate in purebred Arabian mares. A total of 73 purebred Arabian mares who experienced repeat breeding for three successive cycles were randomly divided into the following three groups: control group, without treatment; second Group (PRP group), in which mares were intrauterine infused with 20 mL of fresh PRP on the second day after the end of physic estrus phas...
Paradowska-Dogan A, Fernandez A, Bergmann M, Kretzer K, Mallidis C, Vieweg M, Waliszewski P, Zitzmann M, Weidner W, Steger K, Kliesch S.Highly compacted sperm DNA in protamine toroids and a minor fraction of nucleohistones are prerequisites for the efficient transmission of the paternal genome into the oocyte at fertilization. The objective of this study was to evaluate whether protamines might serve as a prognostic factor for stallion fertility. In situ hybridization detected specific expression of P1 mRNA in the cytoplasm of stage I to VII spermatids, whereas comparable immunohistochemical stainings showed that protein expression was delayed till elongating spermatids in differentiation stages III to VIII. No staining was de...
Rantala M, Attia S, Koukila-Kähkölä P, de Hoog S, Anttila M, Katila T.We present an unusual equine endometritis case associated with Cladophialophora bantiana in a 15-year-old mare. The mare displayed infertility and uterine fluid accumulation with numerous black, hairy granules. Microscopically, the fluid revealed numerous septate, dark fungal hyphae and conidia in chains. Culture yielded C. bantiana (CBS 138271); the species was confirmed by internal transcribed spacer (ITS) sequencing. Treatment was unsuccessful. C. bantiana causes cerebral phaeohyphomycosis in humans, while animal cases are rare. Animal cases are reviewed.
Grüninger B, Schoon HA, Schoon D, Menger S, Klug E.The morphology of endometrial blood vessels in uterine biopsy specimens from mares of varying age and reproductive status was examined by light (n = 117) and electron microscopy (n = 13), and additionally after elastase digestion (n = 86). Inflammatory vascular alterations were observed in 20.5% of the specimens. Smaller and larger arterial and venous vessels demonstrated mild to severe degenerative lesions. Unaltered vessels were detected only in maiden mares. Vessels in older maiden mares were frequently affected by angiosclerotic changes, characterized by mild to moderate perivascular and i...
Lagares MA, Petzoldt R, Sieme H, Klug E.The swelling of cells in a hypo-osmotic medium has been described as an important criterion for assessing the functional integrity of the sperm plasma membrane. The resistance of equine spermatozoa to osmolarity changes was studied by extending 98 semen samples collected from nine stallions in media at five osmolarities (300, 200, 150, 100, and 50 mOsmol l(-1)). The response of the cells was measured by the spermatocrit technique and eosin staining. Spermatocrit determines the increase on spermatozoal volume under hypo-osmotic conditions, a sign of functional integrity of sperm plasma membrane...
Meyers S, Burruel V, Kato M, de la Fuente A, Orellana D, Renaudin C, Dujovne G.In this study we examined the timeline of mitotic events of invitro-produced equine embryos that progressed to blastocyst stage using non-invasive time-lapse microscopy (TLM). Intracytoplasmic sperm injection (ICSI) embryos were cultured using a self-contained imaging incubator system (Miri®TL; Esco Technologies) that captured brightfield images at 5-min intervals that were then generated into video for retrospective analysis. For all embryos that progressed to the blastocyst stage, the initial event of extrusion of acellular debris preceded all first cleavages and occurred at mean (±s.e.m.)...
Vafaei F, Kohram H, Zareh-Shahne A, Ahmad E, Seifi-Jamadi A.This study was aimed to evaluate the effect of permeable cryoprotectants in combination with trehalose or sucrose on the freezing capacity of stallion sperm. For this purpose, the ejaculates (n = 24) were collected from four healthy mature Turkmen stallions. The ejaculates were pooled and diluted with one of the extenders containing a combination of 5% of permeating (dimethylacetamide [DMA]; dimethylformamide [DMF] or glycerol) and 50 mM of nonpermeating cryoprotectant agents (CPAs) (sucrose or trehalose) to a final concentration of 200 × 10 spermatozoa/mL. The extended samples were cryopr...
Merkl M, Ertl R, Handschuh S, Aurich C, Schäfer-Somi S.In the present study, we assessed the presence of the ATP-binding-cassette (ABC) transporter molecules ABCA1 in spermatozoa of adult stallions and in testicular and epididymal tissue of prepubertal and adult stallions. For this purpose, semen samples from six fertile Shetland pony stallions aged 4 to 19 years were collected. Semen was collected from each stallion on three consecutive days. Ejaculates were analyzed immediately after collection, and only ejaculates meeting minimal requirements for fertile stallions were further evaluated. ABCA1 immunosignal was localized after staining of semen ...
de la Torre J, Sánchez-Martín P, Gosálvez J, Crespo F.Sperm quality was assessed in normozoospermic human (n = 10) and Spanish breed stallion (n = 10) after sperm fractionation during ejaculation. The first ejaculated fraction was separated from the second. A third sample was reconstituted using equivalent proportion of both fractions (RAW). Fraction 1, Fraction 2 and RAW semen were incubated for 30 min at 37°C to homogenise the impact of iatrogenic damage between both species. Sperm concentration, motility and sperm DNA damage were assessed in each fraction and RAW semen. The results showed two important facts: (i) spermatozoa confined at ...
Brinsko SP, Blanchard TL, Rigby SL, Love CC, Varner DD.The aim of this study was to determine if dead spermatozoa reduced motility or membrane integrity of live spermatozoa in fresh and cooled-stored equine semen. Three ejaculates from each of three stallions were centrifuged and virtually all seminal plasma was removed. Spermatozoa were resuspended to 25 x 10(6) spermatozoa/ml with EZ-Mixin CST extender and 10% autologous seminal plasma, then divided into aliquots to which 0 (control), 10, 25, 50, or 75% (v/v) dead spermatozoa were added. Dead spermatozoa preparations contained 25 x 10(6) spermatozoa/ml and 10% seminal plasma from pooled ejaculat...
Betteridge KJ, Eaglesome MD, Flood PF.Two experiments were conducted using 14 mares. In Exp. 1, mares were inseminated with semen treated with TEPA, which, in other species, has been shown to lead to an arrest in ovum cleavage at 2--4 cells. The oviducts and/or uterus were then flushed 7--10 days after ovulation in 6 mares (Group A) or 2--6 days after ovulation in 5 mares (Group B). Fresh eggs were found in the oviduct flushes of 5 Group A and 5 Group B mares: 9 of the 10 eggs appeared to have cleaved, but none had developed beyond 16-cells. Seven eggs contained spermatozoa and 3 of 4 eggs from each group showed evidence of fertil...
Grøndahl C, Hyttel P, Grøndahl ML, Eriksen T, Gotfredsen P, Greve T.The objectives were to describe the ultrastructure of equine oocytes aspirated from small and preovulatory follicles, and to relate the ultrastructural features to follicle size and follicular fluid steroid concentrations. Mares were examined every second day by transrectal ultrasonography, and follicles measuring > 30 mm were aspirated (in vivo) using a 20-cm-long 12-gauge needle through the flank. Following slaughter, both large and small follicles were aspirated (in vitro) from six mares. The oocytes were isolated under a stereomicroscope and processed for transmission electron microscop...
The objective of this study was to enhance the in vitro sperm quality and in vivo fertility of frozen-thawed equine semen by the addition of l-carnitine (LC) to post-thawed semen. Different concentrations of LC were added to thawed samples to obtain four treatments control and 0.5, 1 and 2 mM LC. In the in vitro experiments, sperm motility and kinematics, membrane integrity and intracellular calcium ion concentration ([Ca ] ) were investigated, and the antioxidant bioactivity of LC was assessed by measuring hydrogen peroxide and nitrite concentrations (NO ). The fertility rate was assessed v...
Lagares MA, Castanheira PN, Amaral DC, Vasconcelos AB, Veado JC, Arantes RM, Stahlberg R.The aim of the present study was to evaluate the in vitro viability of equine embryos vitrified in three different solutions. Day 6 and 6.5 embryos were measured and morphologically evaluated. Only grade 1 or 2 morulae and early blastocysts were vitrified. Eighteen embryos were distributed in Group 1: 40 percent ethylene glycol in PBS, Group: 2 and 3: 40 percent ethylene glycol, 18 percent Ficoll, 0.3M sucrose or 0.3M trehalose in PBS, respectively. The vitrified embryos were loaded individually into 0.25 ml straws, which were cooled and immersed in liquid nitrogen. After warming at 20 degree ...
Deleuze S, Goudet G, Caillaud M, Lahuec C, Duchamp G.In vivo techniques, such as intraoviductal oocyte transfer (OT) and intrafollicular oocyte transfer (IFOT), can be considered as alternatives to bypass the lack of efficient superovulation treatments and the inadequacy of conventional in vitro fertilization techniques in the horse. We compared embryo production after transfer of in vivo recovered oocytes (1) into a recipient's oviduct or (2) into her preovulatory follicle either immediately after ovum pick-up or (3) after in vitro maturation (IVM). Recipients were inseminated with fresh semen of a stallion with a known normal fertility. Ten da...
Nogueira BG, Sampaio BF, Souza MI, Costa E Silva EV, Zúccari CE.Biotechnology applied for equine semen increases the levels of reactive oxygen species and reduces the natural antioxidant defence, by both dilution and removal of seminal plasma. Therefore, the aims of this study were to evaluate the effect of adding coenzyme Q10 (CoQ10) and α-tocopherol (α-TOH) to the cooling extender, singly or in combination, on sperm parameters, and their effectiveness in preventing lipid peroxidation (LPO) of equine semen during cooling at 5°C for 72 h. Ten adult stallions of proven fertility were used, using two ejaculates each, subjecting them to the treatments with...
Reilas T, Katila T, Mäkelä O, Huhtinen M, Koskinen E.Intrauterine fluid (IUF) was collected using a tampon from mid-oestrous mares (n = 57) with and without ultrasonically detectable accumulations of free intraluminal fluid. Bacteria were cultured and neutrophils counted from all samples (n = 57). Total protein concentration, trypsin-inhibitor capacity (TIC), and plasmin, beta-glucuronidase (B-Gase) and N-acetyl-beta-D-glucosaminidase (NAGase) activities were determined in 27 IUF samples. The motility of spermatozoa in the presence of IUF, IUF extended with Kenney's medium (1:1) and Kenney's medium alone was analysed in 9 samples using a Hamilto...
Moros-Nicolás C, Douet C, Reigner F, Goudet G.In spite of many attempts to establish an in vitro fertilization (IVF) technique in the equine, no efficient conventional IVF technique is available. The presence of oviductal fluid or oviductal cells during IVF helps to improve embryo production in vitro but is not sufficient to reach high fertilization rates. Thus, our aim was to perform equine IVF either after sperm pre-incubation with oviductal fluid or in the presence of oviductal cells, and to evaluate the effect of cumulus removal from the oocyte or sperm pre-incubation with progesterone. In experiments 1 and 2, IVF was performed in the...
Cary JA, Madill S, Farnsworth K, Hayna JT, Duoos L, Fahning ML.The purpose of this study was to evaluate 2 methods of semen collection that could be used as terminal procedures in stallions with irreparable conditions, such as fractures or colic. Electroejaculation was attempted under general anesthesia. Forty-eight hours later, the ponies were castrated and 2 different epididymal sperm collection techniques were attempted by using a flushing or floating method. Additionally, the effect of supplemental seminal plasma was evaluated. Experimentally, electroejaculation was found to be a safe but ineffective method of terminal semen collection. Viable sperm c...
