Survival and apoptosis rates after vitrification in cryotop devices of in vitro-produced calf and cow blastocysts at different developmental stages

Morató, Roser; Izquierdo, Dolors; Paramio, M. T.; Mogas, T.

doi:10.1071/rd10013

Cited by 52 publications

(45 citation statements)

References 35 publications

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“…In a previous study, in which the embryos were not exposed to CLA, our group observed a distinct behavior after vitrification of the embryos at the same stage, but cryopreservation at D7 or D8 (MEZZALIRA et al, 2004). In this study, embryos at an early developmental stage (BLs) were less cryotolerant than those at a later developmental stage (EBs), similar to what was reported by Morató et al (2010). During BL formation and expansion, lipids are metabolized by the trophoblastic cells, which may explain the increased cryotolerance of EBs compared to the BL-stage IVP bovine embryos.…”

Section: Discussionsupporting

confidence: 75%

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Effect of the cryopreservation method used, the embryonic stage and the use of conjugated linoleic acid isomers on the cryotolerance of in vitro-produced bovine embryos

et al. 2015

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Effect of the cryopreservation method used, the embryonic stage and the use of conjugated linoleic acid isomers on the cryotolerance of in vitro-produced bovine embryos Efeito do método de criopreservação utilizado, estágio de desenvolvimento embrionário e uso de isômeros do ácido linoleico conjugado na criotolerância de embriões bovinos produzidos in vitro AbstractConjugated linoleic acid (CLA) might be able to improve the cryotolerance of in vitro-produced (IVP) embryos. The effect of two CLA isomers on the cryotolerance of bovine IVP embryos, as well as that of the stage of embryonic development and the method used for cryopreservation was evaluated by three experiments. In Experiment 1, oocytes (n = 3,917) were fertilized in vitro and cultured with 0, 50, 100, or 200 µM trans-10, cis-12 (t10, c12 CLA). In Experiment 2, fertilized oocytes (n = 2,131) were cultured with 100 µM t10, c12 or cis-9, trans-11 (c9, t11 CLA), or a combination of both isomers. The embryos were vitrified at the blastocyst (BL) or the expanded blastocyst (EB) stage. In Experiment 3, oocytes (n = 1,720) were fertilized and cultured with or without 100 µM t10, c12 CLA, and the blastocysts were vitrified or frozen. Blastocyst development rate as well as the rates of re-expansion and hatching after thawing was recorded. Moreover, the mean cell number and mRNA expression of acetyl-CoA carboxylase (ACC1) and stearoyl-CoA desaturase (SCD1) as well as fatty acid synthase (FASN) multienzyme complex were determined. In Experiment 1, the highest concentration of t10, c12 CLA that did not reduce blastocyst development rate was 100 µM. In Experiment 2, the rates of re-expansion and hatching among the EBs obtained through IVP after supplementation with t10, c12 CLA (73.1% and 57.7%), with c9, t11 CLA (80.0% and 68.6%), with the combination (78.3% and 52.2%), and with the control group (85.4% and 58.3%) were similar. At the BL stage, the rates of re-expansion and hatching were lower than those at the EB stage, and CLA combination allowed a hatching rate (8.0%) lower than that observed in the control group (40.0%). In Experiment 3, the hatching rates for vitrified EBs (vitrified control; 67.4%) and vitrified CLA EBs (65.8%) were higher than those obtained for frozen EBs, exposed (13.3%) or not exposed (28.6%) to CLA. In addition, in Experiment 3, the hatching rate was higher at the EB stage in vitrified groups, while the rates of BL and EB were similar in frozen groups, thus proving that vitrification was more efficient than freezing for IVP bovine embryos. In Experiment 3, CLA isomer t10, C12 did not influence the embryonic cell number or mRNA expression of ACC1 and SCD1 enzymes, but decreased the mRNA expression of FASN. In conclusion, 100 µM CLA did not affect subsequent embryonic development. However, neither CLA isomer improved the cryotolerance of IVP bovine embryos. c12 CLA). No Experimento 2, oócitos fecundados (n = 2.131) foram cultivados com 100 µM de t10, c12 ou cis-9, trans-11 (c9, t11 CLA), ou ainda com uma associação de ambos. Os embri...

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Section: Discussionsupporting

confidence: 75%

“…This is possible by embryo culture in serum-free systems (ORDOÑEZ-LEON et al, 2014) or by mechanical removal of intracellular lipids (KAWAKAMI et al, 2008). However, both approaches reduce the developmental viability of embryos.…”

Section: Introductionmentioning

confidence: 99%

Effect of the cryopreservation method used, the embryonic stage and the use of conjugated linoleic acid isomers on the cryotolerance of in vitro-produced bovine embryos

et al. 2015

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“…This damage may occur due to the use of high concentrations of cryoprotectant solutions employed in the current vitrification technique to generate a glass-like physical state (Campos-Chillòn et al, 2006), which increases the risk of osmotic and toxic damage (Barceló-Fimbres & Seidel, 2007b). Conversely, Morató et al (2010) did not observe any difference in apoptosis rates between fresh and vitrified blastocysts. However, the authors justify that the period between warming and DNA fragmentation examination (3 h) was not enough to identify the extention of damage caused by the vitrification-warming procedure.…”

Section: Figurecontrasting

confidence: 56%

Crucial surviving aspects for vitrified in vitro-produced bovine embryos

Sudano

Paschoal

Rascado

et al. 2012

Zygote

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The objective of the present study was to correlate some parameters (cleavage, blastocyst production, quality degree score, total cell number, fresh apoptosis and lipid content) with embryo survival after cryopreservation. A total of 1727 in vitro-produced bovine blastocysts were used to establish the parameters (mean ± standard error of the mean (SEM)) for cleavage (85.6 ± 0.8), blastocyst production (39.9 ± 1.4), quality degree score (1.6 ± 0.1), total cell number (140.1 ± 2.9), fresh apoptosis (20.8 ± 1.1) and lipid content (21.3 ± 0.8 droplets). On the same way 1316 blastocysts were vitrified for the determination of post-cryopreservation embryo survival (49.4 ± 1.9). Fresh apoptosis rate and total lipid droplets value were correlated (P < 0.05) with embryo survival after cryopreservation (r = 0.91 and r = 0.59; respectively). However, cleavage, blastocyst production, quality degree score and total cell number were not correlated (P > 0.05) with embryo cryotolerance (r = 0.23, r = 0.38, r = 0.22 and r = 0.28; respectively). Therefore, the increased lipid content was moderately correlated with apoptosis in vitrified blastocysts. On the other hand, increased apoptosis in fresh blastocysts was strongly correlated with apoptosis in vitrified blastocysts, which indicated that the apoptosis rate in fresh embryos was a better parameter than the lipid content to predict post-vitrification embryo survival.

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“…The successful cryopreservation of IVP bovine embryos is essential to the establishment of its use on a commercial scale. With recent advances in freezing protocols, cryopreservation of IVP embryos has become a fast and reliable tool that allows the indefinite storage of embryos with higher potentials for in vitro development (Morató et al 2010). However, there has been little attention on the effects of antioxidants on the cryosurvival of vitrified-warmed embryos (Hosseini et al 2009).…”

Section: Introductionmentioning

confidence: 99%

Reduced levels of intracellular reactive oxygen species and apoptotic status are not correlated with increases in cryotolerance of bovine embryos produced in vitro in the presence of antioxidants

Rocha-Frigoni

Leão

Nogueira

et al. 2014

Reprod. Fertil. Dev.

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The effects of intracellular (cysteine and β-mercaptoethanol) and extracellular (catalase) antioxidant supplementation at different times during in vitro production (IVM and/or in vitro culture (IVC)) on bovine embryo development, intracellular reactive oxygen species (ROS) levels, apoptosis and re-expansion rates after a vitrification-thawing process were examined. Blastocyst frequencies were not affected by either antioxidant supplementation (40.5%-56.4%) or the timing of supplementation (41.7%-55.4%) compared with control (48.7%; P>0.05). Similarly, antioxidants and the moment of supplementation did not affect (P>0.05) the total number of blastomeres (86.2-90.5 and 84.4-90.5, respectively) compared with control (85.7). However, the percentage of apoptotic cells was reduced (P<0.05) in groups supplemented during IVM (1.7%), IVC (2.0%) or both (1.8%) compared with control (4.3%). Intracellular ROS levels measured in Day 7 blastocysts were reduced (P<0.05) in all groups (0.60-0.78), with the exception of the group supplemented with β-mercaptoethanol during IVC (0.88), which did not differ (P>0.05) from that in the control group (1.00). Re-expansion rates were not affected (P>0.05) by the treatments (50.0%-93.0%). In conclusion, antioxidant supplementation during IVM and/or IVC reduces intracellular ROS and the rate of apoptosis; however, supplementation does not increase embryonic development and survival after vitrification.

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Survival and apoptosis rates after vitrification in cryotop devices of in vitro-produced calf and cow blastocysts at different developmental stages

Cited by 52 publications

References 35 publications

Effect of the cryopreservation method used, the embryonic stage and the use of conjugated linoleic acid isomers on the cryotolerance of in vitro-produced bovine embryos

Effect of the cryopreservation method used, the embryonic stage and the use of conjugated linoleic acid isomers on the cryotolerance of in vitro-produced bovine embryos

Crucial surviving aspects for vitrified in vitro-produced bovine embryos

Reduced levels of intracellular reactive oxygen species and apoptotic status are not correlated with increases in cryotolerance of bovine embryos produced in vitro in the presence of antioxidants

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