- Email: [email protected]
Report of frozen equine embryos transfer in the Czech Republic
Abstracts / Journal of Equine Veterinary Science 41 (2016) 51e84
antitrypsin 1, alpha-1 antitrypsin 14), uteroglobin, uteroferrin, deleted in malignant brain tumor 1, mucins (4, 5b and 6). Mass spectrometry identified a total of 218 proteins within the CMP, with lactoferrin consistently the most abundant protein, followed closely by immunoglobulin A and G, respectively. The most abundant mucin in the CMP was mucin 5B, followed by mucin 5AC, mucin 4 and mucin 6. Ingenuity Pathway Analysis identified a total of 38 pathways, 20 of which were primarily related to immune functions. Gross histology identified two distinct regions within the CMP, a cellular region and an acellular region. The acellular region stained more strongly with PAS, suggesting this region contains a higher proportion of heavily glycosylated proteins such as mucins. Analysis by qPCR showed that the cervical mucosa contained transcripts for each of the examined mRNAs during pregnancy. Lactoferrin, uterine serpins, uteroglobin, uteroferrin, mucin 4 and mucin 5b were each significantly upregulated during pregnancy compared to diestrus mares (P < 0.05). Mucin 6 and deleted in malignant brain tumor 1 were not upregulated in the cervical mucosa during pregnancy, but did exhibit pregnancy-dependent fluctuations in the endometrium. Overall, we showed that the CMP is comprised of a complex milieu of proteins during late gestation, many of which are known to play an important role in immune function. Proteins traditionally considered to be endometrial proteins were found to be produced by the cervical mucosa as well as the endometrium and were upregulated during pregnancy in the cervical mucosa. In summary, composition of the equine CMP is specifically regulated during pregnancy, and the CMP provides both a structural barrier (due to its high mucin content) as well as antimicrobial properties to prevent infection of the conceptus during pregnancy. Key Words: Cervical Mucus Plug, Proteome, Mare, Cervical Mucosa, Endometrium
Acknowledgements The authors thank Dr. Igor Canisso for his help in obtaining materials used in this study. Supported by the Albert G. Clay Endowment at the University of Kentucky.
36 Report of frozen equine embryos transfer in the Czech Republic Josef Muller, Jindrich Muller Equine Reproduction Center, Pardubice-Mnetice, Czech Republic In the period of 2008 to 2015, we used as donors 17 Warmbloded sport-type mares, ranging in age from 2 to 22 years. These mares were inseminated with frozen semen fromone of 38 sport horse stallions, imported mainly from Germany, France and Holland. Insemination was performed by one of three insemination technicians, to the top of uterus horn, just before ovulation or within 2 to 4 hours after ovulation. An embryo collection attempt was performed 163 to 172 hours after ovulation. Embryos were frozen via vitrification method using a commercial equine vitrification kit (Bioniche) as recomended. Embryos were stored for 2 to 59 months in liqiud nitrogen before transfer. Embryo recovery varied between individual donor mares (12 to 140 %) and between stallions. Together a total of 118 flushings of donor mares were performed and 64 embryos (54%) were recovered. To date, a total of 33 frozen embryos have been nonsurgically transfered to the uterus of 25 warmbloded recepient mares, ranging in age from 2 to 14 years. A total of 17 mares were determined to be pregnant (52%); of these 14 foals have been born, 2 mares are still pregnant and 1 mare resorbed her pregnancy.
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37 Influence of mare parity on weight, height, thoracic circumference and vitality of neonatal foals F.J. Affonso, M.G. Meirelles, M.A. Alonso, C.F. Guimaraes, K.M. Lemes, M. Nichi, C.B. Fernandes* Department of Animal Reproduction, College of Veterinary Medicine ~o Paulo, SP, Brazil and Animal Science, University of Sa *Corresponding author: [email protected] In equine embryo transfer programs, there is a great deal of emphasis placed on recipient mare characteristics such as size and parity. Studies have demonstrated that multiparous females have a more developed intrauterine environment and are better able to provide nutritional support for fetal and placental growth [Wilsher S Equine Vet J 2003;35;476-483]. Elliott C. Theriogenology 2009;71;683e9]. A positive correlation has been established between placental weight and foal birth weight, as well as between parity and foal birth weight, with an average of an 0.8 kg increase for every extra foaling [2]. A previous study performed by our group noted a positive correlation between parity and placental weight, as well as placental weight and foal size [Fernandes CB et.al .J Equine Vet Sci 2014; 34:225-7]. However, there are few studies that relate maternal characteristics with foal size and vitality at birth. The assessment of neonatal characteristics and attitude/behavior of the newborn are important to identify and correct neonatal disorders within the first hours of life. The aim of this study was to correlate parity with weight, height, thoracic circumference and neonatal vitality of Mangalarga Paulista and Brazilian Sport Horse foals. The foaling of 38 mares was observed, with the mares placed into three groups based on parity. The groups were defined as G1: nulliparous (n:14); G2: 1-3 births (n:15) and G3: 4 births (n:9). Data collected included measures of neonatal vitality (APGAR Score at 3 and 60 min, and the time taken to sit sternally, to observe the suckling reflex, to stand, to nurse and to eliminate meconium), as well as weight, height and thoracic circumference at birth, 7 and 30 days of life. The data were analyzed by ANOVA (GLM), with post hoc analysis performed using Tukey's procedure. G1 newborns had a lower birth weight, height and thoracic circumference at birth, 7 and 30 days after birth (p<0.05); however, G2 and G3 did not differ significantly. Most neonatal parameters were similar between the three groups, with the exception of time to stand which was greater for G3 compared with G1 (p<0.05), and likely related to the larger foal size seen in G3. It has been demonstrated that foals from larger breeds take longer to stand than those from smaller breeds [Stoneham SJ. In: Paradis MR. Equine neonatal medicine: a casebased approach, Philadelphia; 2006, p 1-11]. Thus, parity influences the size of the foal and time to stand, but not the additional vitality parameters studied. As observed in previous studies nulliparous mares produced smaller foals with a lower birth weight. This is likely because these mares show a reduction in microcotyledon surface density with a subsequent decrease in fetomaternal contact, demonstrating the importance of the first pregnancy in preparing the intrauterine environment for subsequent pregnancies In contrast, pluriparous mares produced taller and heavier foals. We noted that the influence of parity on foal size is not about the number of pregnancies, since no differences were observed between the groups G2 and G3, but is seen with any previous pregnancy. The differences in foal size were maintained at 7 and 30 days after birth. Key Words: recipient mare, placenta, neonatology, equine
Acknowledgments Financial Support FAPESP 2012.08929-6.
antitrypsin 1, alpha-1 antitrypsin 14), uteroglobin, uteroferrin, deleted in malignant brain tumor 1, mucins (4, 5b and 6). Mass spectrometry identified a total of 218 proteins within the CMP, with lactoferrin consistently the most abundant protein, followed closely by immunoglobulin A and G, respectively. The most abundant mucin in the CMP was mucin 5B, followed by mucin 5AC, mucin 4 and mucin 6. Ingenuity Pathway Analysis identified a total of 38 pathways, 20 of which were primarily related to immune functions. Gross histology identified two distinct regions within the CMP, a cellular region and an acellular region. The acellular region stained more strongly with PAS, suggesting this region contains a higher proportion of heavily glycosylated proteins such as mucins. Analysis by qPCR showed that the cervical mucosa contained transcripts for each of the examined mRNAs during pregnancy. Lactoferrin, uterine serpins, uteroglobin, uteroferrin, mucin 4 and mucin 5b were each significantly upregulated during pregnancy compared to diestrus mares (P < 0.05). Mucin 6 and deleted in malignant brain tumor 1 were not upregulated in the cervical mucosa during pregnancy, but did exhibit pregnancy-dependent fluctuations in the endometrium. Overall, we showed that the CMP is comprised of a complex milieu of proteins during late gestation, many of which are known to play an important role in immune function. Proteins traditionally considered to be endometrial proteins were found to be produced by the cervical mucosa as well as the endometrium and were upregulated during pregnancy in the cervical mucosa. In summary, composition of the equine CMP is specifically regulated during pregnancy, and the CMP provides both a structural barrier (due to its high mucin content) as well as antimicrobial properties to prevent infection of the conceptus during pregnancy. Key Words: Cervical Mucus Plug, Proteome, Mare, Cervical Mucosa, Endometrium
Acknowledgements The authors thank Dr. Igor Canisso for his help in obtaining materials used in this study. Supported by the Albert G. Clay Endowment at the University of Kentucky.
36 Report of frozen equine embryos transfer in the Czech Republic Josef Muller, Jindrich Muller Equine Reproduction Center, Pardubice-Mnetice, Czech Republic In the period of 2008 to 2015, we used as donors 17 Warmbloded sport-type mares, ranging in age from 2 to 22 years. These mares were inseminated with frozen semen fromone of 38 sport horse stallions, imported mainly from Germany, France and Holland. Insemination was performed by one of three insemination technicians, to the top of uterus horn, just before ovulation or within 2 to 4 hours after ovulation. An embryo collection attempt was performed 163 to 172 hours after ovulation. Embryos were frozen via vitrification method using a commercial equine vitrification kit (Bioniche) as recomended. Embryos were stored for 2 to 59 months in liqiud nitrogen before transfer. Embryo recovery varied between individual donor mares (12 to 140 %) and between stallions. Together a total of 118 flushings of donor mares were performed and 64 embryos (54%) were recovered. To date, a total of 33 frozen embryos have been nonsurgically transfered to the uterus of 25 warmbloded recepient mares, ranging in age from 2 to 14 years. A total of 17 mares were determined to be pregnant (52%); of these 14 foals have been born, 2 mares are still pregnant and 1 mare resorbed her pregnancy.
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37 Influence of mare parity on weight, height, thoracic circumference and vitality of neonatal foals F.J. Affonso, M.G. Meirelles, M.A. Alonso, C.F. Guimaraes, K.M. Lemes, M. Nichi, C.B. Fernandes* Department of Animal Reproduction, College of Veterinary Medicine ~o Paulo, SP, Brazil and Animal Science, University of Sa *Corresponding author: [email protected] In equine embryo transfer programs, there is a great deal of emphasis placed on recipient mare characteristics such as size and parity. Studies have demonstrated that multiparous females have a more developed intrauterine environment and are better able to provide nutritional support for fetal and placental growth [Wilsher S Equine Vet J 2003;35;476-483]. Elliott C. Theriogenology 2009;71;683e9]. A positive correlation has been established between placental weight and foal birth weight, as well as between parity and foal birth weight, with an average of an 0.8 kg increase for every extra foaling [2]. A previous study performed by our group noted a positive correlation between parity and placental weight, as well as placental weight and foal size [Fernandes CB et.al .J Equine Vet Sci 2014; 34:225-7]. However, there are few studies that relate maternal characteristics with foal size and vitality at birth. The assessment of neonatal characteristics and attitude/behavior of the newborn are important to identify and correct neonatal disorders within the first hours of life. The aim of this study was to correlate parity with weight, height, thoracic circumference and neonatal vitality of Mangalarga Paulista and Brazilian Sport Horse foals. The foaling of 38 mares was observed, with the mares placed into three groups based on parity. The groups were defined as G1: nulliparous (n:14); G2: 1-3 births (n:15) and G3: 4 births (n:9). Data collected included measures of neonatal vitality (APGAR Score at 3 and 60 min, and the time taken to sit sternally, to observe the suckling reflex, to stand, to nurse and to eliminate meconium), as well as weight, height and thoracic circumference at birth, 7 and 30 days of life. The data were analyzed by ANOVA (GLM), with post hoc analysis performed using Tukey's procedure. G1 newborns had a lower birth weight, height and thoracic circumference at birth, 7 and 30 days after birth (p<0.05); however, G2 and G3 did not differ significantly. Most neonatal parameters were similar between the three groups, with the exception of time to stand which was greater for G3 compared with G1 (p<0.05), and likely related to the larger foal size seen in G3. It has been demonstrated that foals from larger breeds take longer to stand than those from smaller breeds [Stoneham SJ. In: Paradis MR. Equine neonatal medicine: a casebased approach, Philadelphia; 2006, p 1-11]. Thus, parity influences the size of the foal and time to stand, but not the additional vitality parameters studied. As observed in previous studies nulliparous mares produced smaller foals with a lower birth weight. This is likely because these mares show a reduction in microcotyledon surface density with a subsequent decrease in fetomaternal contact, demonstrating the importance of the first pregnancy in preparing the intrauterine environment for subsequent pregnancies In contrast, pluriparous mares produced taller and heavier foals. We noted that the influence of parity on foal size is not about the number of pregnancies, since no differences were observed between the groups G2 and G3, but is seen with any previous pregnancy. The differences in foal size were maintained at 7 and 30 days after birth. Key Words: recipient mare, placenta, neonatology, equine
Acknowledgments Financial Support FAPESP 2012.08929-6.