scholarly journals Antioxidant activity of oily extract obtained from Lippia origanoides improves the quality of bovine embryos produced in vitro

2019 ◽  
Vol 71 (3) ◽  
pp. 723-731
Author(s):  
N.V. Sollecito ◽  
E.C.M. Pereira ◽  
J.G.V. Grázia ◽  
B.P. Neves ◽  
B.V.R. Couto ◽  
...  
Keyword(s):  
Embryo Culture ◽  
Culture Medium ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Blastocyst Development ◽  
Embryo Culture Medium ◽  
Vitro Production

ABSTRACT The aim of this study was to evaluate the supplementation of embryo culture medium with antioxidant obtained from oily extract of Lippia origanoides on in vitro blastocyst development and quality. Oocytes collected from slaughterhouse ovaries were matured and fertilized in vitro following standard laboratory procedures. Zygotes were cultured in SOF medium supplemented according to the following treatments: T1 embryo culture medium without antioxidant supplementation; T2)50μM/mL Cysteamine; T3)2.5μg/mL; T4)5.0μg/mL and T5)10.0μg/mL of antioxidant obtained from oily extract of Lippia origanoides. On the seventh day of culture, the blastocysts were fixed and evaluated for apoptosis rates, number of total cell and inner cell mass cells by means of the TUNEL Test. The use of antioxidants during cultivation did not increase (P> 0.05) the final blastocyst production rate. The treatments T2, T3, T4 and T5 had the lowest (P< 0.05) apoptotic indexes (4.5±1.1%, 8.4±2.5%, 3.4±1.1% and 5.5±0.9%, respectively) when compared to T1 treatment (10.0±1.4%). The number of inner cell mass did not differ (P> 0.05) among embryos from different treatments. The addition of antioxidant obtained from oily extract of Lippia origanoides reduces the apoptosis rate and improves the quality without increasing the total in vitro production of bovine embryos.

scholarly journals Cell lineage allocation in equine blastocysts produced in vitro under varying glucose concentrations

Reproduction ◽  
2015 ◽  
Vol 150 (1) ◽  
pp. 31-41 ◽  
Author(s):  
Young-Ho Choi ◽  
Pablo Ross ◽  
Isabel C Velez ◽  
B Macías-García ◽  
Fernando L Riera ◽  
...  
Keyword(s):  
High Glucose ◽  
Culture Medium ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Lineage ◽  
Primitive Endoderm ◽  
Blastocyst Development ◽  
Embryo Culture Medium ◽  
Effect Of Glucose

Equine embryos developin vitroin the presence of high glucose concentrations, but little is known about their requirements for development. We evaluated the effect of glucose concentrations in medium on blastocyst development after ICSI. In experiment 1, there were no significant differences in rates of blastocyst formation among embryos cultured in our standard medium (DMEM/F-12), which contained >16 mM glucose, and those cultured in a minimal-glucose embryo culture medium (<1 mM; Global medium, GB), with either 0 added glucose for the first 5 days, then 20 mM (0-20) or 20 mM for the entire culture period (20-20). In experiment 2, there were no significant differences in the rates of blastocyst development (31–46%) for embryos cultured in four glucose treatments in GB (0-10, 0-20, 5-10, or 5-20). Blastocysts were evaluated by immunofluorescence for lineage-specific markers. All cells stained positively forPOU5F1. An inner cluster of cells was identified that included presumptive primitive endoderm cells (GATA6-positive) and presumptive epiblast (EPI) cells. The 5-20 treatment resulted in a significantly lower number of presumptive EPI-lineage cells than the 0-20 treatment did.GATA6-positive cells appeared to be allocated to the primitive endoderm independent of the formation of an inner cell mass, as was previously hypothesized for equine embryos. These data demonstrate that equine blastocyst development is not dependent on high glucose concentrations during early culture; rather, environmental glucose may affect cell allocation. They also present the first analysis of cell lineage allocation inin vitro-fertilized equine blastocysts. These findings expand our understanding of the factors that affect embryo development in the horse.

scholarly journals N-(2-mercaptopropionyl)-glycine enhances in vitro pig embryo production and reduces oxidative stress

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
J. M. Cambra ◽  
C. A. Martinez ◽  
H. Rodriguez-Martinez ◽  
E. A. Martinez ◽  
C. Cuello ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Culture Medium ◽  
Control Culture ◽  
Related Gene ◽  
In Vitro Production ◽  
Embryo Culture Medium ◽  
Porcine Embryo ◽  
Vitro Production ◽  
Oxidative Stress Related Gene

Abstract This study evaluated the effects of different concentrations (1, 10, 25, 50, and 100 µM) of the antioxidant N-(2-mercaptopropionyl)-glycine (NMPG), during the culture of in vitro-fertilized porcine oocytes. While the highest concentrations of NMPG (50 and 100 µM) were toxic to the developing embryos during the first two days of culture, 25 µM NMPG achieved cleavage rates that were similar to those achieved by the control but did not sustain blastocyst production by Day 7 of culture. Compared to the control culture medium, the culture medium supplemented with 10 µM NMPG increased (P < 0.05) the rates of blastocyst formation, decreased (P < 0.05) the intracellular levels of reactive oxygen substances, and downregulated (P < 0.05) the expression of the oxidative stress related gene GPX1. In conclusion, these results demonstrated that supplementation of porcine embryo culture medium with 10 µM NMPG can attenuate oxidative stress and increase the yield of in vitro production of blastocysts.

In vitro production of bovine embryos in medium supplemented with a serum replacer: Effects on blastocyst development, cryotolerance and survival to term

2007 ◽  
Vol 68 (9) ◽  
pp. 1316-1325 ◽  
Author(s):  
K. Moore ◽  
C.J. Rodríguez-Sallaberry ◽  
J.M. Kramer ◽  
S. Johnson ◽  
E. Wroclawska ◽  
...  
Keyword(s):  
Bovine Embryos ◽  
In Vitro Production ◽  
Blastocyst Development ◽  
Vitro Production

scholarly journals Zinc supplementation during in vitro embryo culture increases inner cell mass and total cell numbers in bovine blastocysts1

2019 ◽  
Vol 97 (12) ◽  
pp. 4946-4950 ◽  
Author(s):  
Lydia K Wooldridge ◽  
Madison E Nardi ◽  
Alan D Ealy
Keyword(s):  
Embryo Culture ◽  
Zinc Supplementation ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Number ◽  
Retention Rates ◽  
Bovine Embryo ◽  
Blastocyst Development ◽  
Cell Numbers

Abstract Deficiencies in current embryo culture media likely contribute to the poor blastocyst development rates and pregnancy retention rates for in vitro produced (IVP) bovine embryos. Of special concern is the lack of micronutrients in these media formulations. One micronutrient of interest is zinc, an essential trace element involved with various enzyme and transcription factor activities. The objective of this work was to describe whether zinc sulfate supplementation during in vitro embryo culture affects bovine embryo development and blastomere numbers. Either 0, 2, 20, or 40 µM zinc sulfate was supplemented to presumptive zygotes cultured in synthetic oviductal fluid containing AAs and bovine serum albumin for 8 d. None of the treatments affected cleavage rates. Percentage of blastocysts on days 7 and 8 postfertilization was not affected by supplementing 2 or 20 µM zinc but were reduced (P &lt; 0.05) with 40 µM zinc. In blastocysts harvested on day 8, inner cell mass (ICM) and total cell number were increased (P &lt; 0.05) with 2 µM zinc supplementation but not with the other zinc concentrations. Numbers of trophectoderm cells were not affected by zinc treatment. In conclusion, supplementing zinc during bovine embryo culture did not impact blastocyst development but improved ICM cell numbers. This improvement in ICM cell number may have implications for improved pregnancy retention rates after IVP embryo transfer as smaller ICM sizes are associated with poor pregnancy success in cattle.

scholarly journals Bovine conceptus of Bos indicus produced by somatic cell nuclear transfer and parthenogenesis present morphological variations since the blastocyst stage

2015 ◽  
Vol 67 (6) ◽  
pp. 1483-1491
Author(s):  
F.D. Oliveira ◽  
J.R. Sangalli ◽  
F.V. Meirelles ◽  
F. Perecin ◽  
P.P.C. Silva Filho ◽  
...  
Keyword(s):  
Nuclear Transfer ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Bos Indicus ◽  
Bovine Embryo ◽  
In Vitro Production ◽  
Production Techniques ◽  
Morphological Differences ◽  
Vitro Production

In cattle, embryo development is characterized by the appearance of two distinct cell layers, the trophectoderm and the inner cell mass. The latter will undergo differentiation to form the embryonic disc consisting of the epiblast and hypoblast. The aim of this study was to ultrastructurally characterize the bovine embryo from different in vitro production techniques, with emphasis on trophectoderm and inner cell mass cells. Bovine embryos on day 7 (conception = D1) of pregnancy, derived via in vitro production techniques, were fixed for light and transmission electron microscopy processing. Results suggested that embryos produced by nuclear transfer of somatic cells and parthenogenesis showed significant changes in macroscopic and microscopic structure. Size was reduced, and the inner cell mass had no defined shape. Furthermore, organelles responsible for the absorption processes, communication, growth, and cellular metabolism were fewer and had changes in shape, when compared to results in embryos produced by in vitrofertilization. We concluded that embryos produced by parthenogenesis and SCNT exhibit morphological differences when compared with IVF embryos, such as undeveloped blastocoel, poorly defined distribution of ICM, and morphological differences in organelles.

91 INTERACTION OF OXYGEN TENSION AND CYSTEAMINE SUPPLEMENTATION DURING IN VITRO CULTURE OF BOVINE EMBRYOS

2012 ◽  
Vol 24 (1) ◽  
pp. 158
Author(s):  
L. M. Stauber ◽  
G. E. Seidel
Keyword(s):  
In Vitro Culture ◽  
Embryo Culture ◽  
Culture Medium ◽  
Cumulus Cells ◽  
Bovine Embryos ◽  
Cell Stage ◽  
Embryo Culture Medium ◽  
Mammalian Embryos ◽  
Defined Culture

Reactive oxygen species damage early mammalian embryos, so culture of bovine in vitro-produced (IVP) embryos at 5% O2 is clearly superior to 20% O2. The thiol compound cysteamine is an antioxidant and improves in vitro blastocyst production when added to in vitro maturation (IVM) medium. The purpose of this study was to investigate supplementation of IVP embryo culture medium with cysteamine at different oxygen tensions. Bovine ovaries from feedlot heifers were collected from a local abattoir and cumulus–oocyte complexes (COC) were aspirated from 3- to 8-mm follicles. COC were matured in maturation medium with 100μM cysteamine (Reprod. Fertil. Dev. 18, 585) for 23 h in a humidified incubator at 38.5°C in 5% CO2 in air. COC at 23 h of maturation were co-incubated with sperm for 18 h. Cumulus cells were then removed and presumed zygotes were cultured in our chemically defined culture medium system (Reprod. Fertil. Dev. 18, 585) in a 2 × 2 factorial arrangement: with or without 50 μM cysteamine × atmospheres of either 5% O2, 5% CO2, 90% N2, or 5% CO2 in air. There were no treatment differences (P > 0.01) in percentages of oocytes cleaving or reaching the 8-cell stage (Table 1). However, blastocyst production rates were lower (P < 0.01) in the group cultured without cysteamine at 20% O2 compared with all the other groups. Adding cysteamine for embryo culture at 20% O2 resulted in blastocyst rates similar to those cultured at 5% O2 with or without cysteamine. Cysteamine was not beneficial at 5% O2. Table 1.Cysteamine supplementation during in vitro culture of bovine embryos

scholarly journals Biological differences between in vitro produced bovine embryos and parthenotes

Reproduction ◽  
2009 ◽  
Vol 137 (2) ◽  
pp. 285-295 ◽  
Author(s):  
Enrique Gómez ◽  
Alfonso Gutiérrez-Adán ◽  
Carmen Díez ◽  
Pablo Bermejo-Alvarez ◽  
Marta Muñoz ◽  
...  
Keyword(s):  
De Novo ◽  
Inner Cell Mass ◽  
Relative Proportion ◽  
Cell Mass ◽  
Blastocyst Stage ◽  
Bovine Embryos ◽  
Blastocyst Development ◽  
Cell Counts ◽  
Biological Differences

Parthenotes may represent an alternate ethical source of stem cells, once biological differences between parthenotes and embryos can be understood. In this study, we analyzed development, trophectoderm (TE) differentiation, apoptosis/necrosis, and ploidy in parthenotes andin vitroproduced bovine embryos. Subsequently, using real-time PCR, we analyzed the expression of genes expected to underlie the observed differences at the blastocyst stage.In vitromatured oocytes were either fertilized or activated with ionomycin +6-DMAP and cultured in simple medium. Parthenotes showed enhanced blastocyst development and diploidy and reduced TE cell counts. Apoptotic and necrotic indexes did not vary, but parthenotes evidenced a higher relative proportion of apoptotic cells between inner cell mass and TE. The pluripotence-relatedPOU5F1and the methylationDNMT3Agenes were downregulated in parthenotes. Among pregnancy recognition genes,TP-1was upregulated in parthenotes, whilePGRMC1andPLAC8did not change. Expression ofp66shcandBAX/BCL2ratio were higher, andp53lower, in parthenotes. Among metabolism genes,SLC2A1was downregulated, whileAKR1B1,PTGS2,H6PD, andTXNwere upregulated in parthenotes, andSLC2A5did not differ. Among genes involved in compaction/blastulation,GJA1was downregulated in parthenotes, but no differences were detected withinATP1A1andCDH1. Within parthenotes, the expression levels ofSLC2A1,TP-1, andH6PD, and possiblyAKR1B1, resemble patterns described in female embryos. The pro-apoptotic profile is more pronounced in parthenotes than in embryos, which may differ in their way to channel apoptotic stimuli, throughp66shcandp53respectively, and in their mechanisms to control pluripotency andde novomethylation.

scholarly journals In vitro production of bovine embryos using flow-cytometrically sexed sperm

2006 ◽  
Vol 49 (2) ◽  
pp. 133-140 ◽  
Author(s):  
L. Kątska-Książkiewicz ◽  
B. Ryńska ◽  
M. Bochenek ◽  
J. Opiela ◽  
J. Jurkiewicz
Keyword(s):  
Bovine Embryos ◽  
In Vitro Production ◽  
Blastocyst Development ◽  
Vitro Fertilization ◽  
Bovine Spermatozoa ◽  
Embryo Cleavage ◽  
And Control ◽  
Vitro Production ◽  
Sperm Freezing

Abstract. The investigation aimed to compare the effect of fresh and frozen-thawed X and Y fractions of flow-cytometrically sorted bovine spermatozoa on in vitro fertilization of bovine in vitro matured oocytes and subsequent blastocyst development. Sperm cells sorted in MoFloSX cytometer were used either for IVF or frozen and stored in liquid nitrogen. Immature oocytes recovered from ovaries of slaughtered animals and matured in vitro in TCM-199 containing 20% estrus cow serum and additional granulosa cells were fertilized in vitro with fresh or frozen-thawed fractions of sorted sperm. Simultaneously, control, fresh or frozen/thawed sperm was used for IVF. A total number of 2712 IVM oocytes were fertilized with sorted and control sperm of 6 bulls. Embryo cleavage rates were significantly affected by bull (P<0.0001), sperm sexing (P<0.0001) and sperm freezing (P<0.01). Blastocysts development was affected by sperm freezing (P<0.04) and sperm sexing (P<0.01). The significant differences were shown between unsorted and sorted sperm, however no differences in embryo cleavage rates and blastocysts rates were observed between X- and Y-sperm fractions, both fresh and frozen/ thawed. There were significant differences in cleavage rates among fresh, control sperm (52.7%), X fraction (26.8%) and Y fraction (24.7%). Similar differences in cleavage rates were shown for frozen/thawed control sperm (52.8%), X fraction (33.9%) and Y fraction (26.2%). The female blastocysts were frozen for further transfer, while the hatched male blastocysts were analysed by PCR revealing 76.2% accuracy. The results suggest that there were significant differences in cleavage rates and blastocyst rates due to sperm sorting in comparison to unsorted sperm and no differences between effectiveness of X and Y fractions of spermatozoa.

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