38 Vitrification of Bos Taurus Indicus and Bos Taurus Indicus×bos Taurus Taurus Embryos Produced in the Presence or Absence of Fetal Calf Serum

Abstract

In vitro-produced Bos taurus indicus (zebu) and Bos taurus indicus × Bos taurus taurus (cross-bred) embryos behave differently when vitrified. The present experiment aimed to examine the effect of vitrification on embryos produced in the presence or absence of FCS. Cumulus-oocyte complexes (COC) were matured in TCM-199 and fertilized in human tubal fluid medium with frozen Nelore bull semen. On Day 1 (Day 0 = IVF), presumptive zygotes were cultured with SOFaa + BSA in the presence of FCS (Group 2.5%) or in the absence of FCS (Group 0%) until Day 7. The cleavage was analysed on Day 3 and the blastocyst rate on Day 7. Blastocysts were vitrified and, after warming (Campos-Chillòn et al. 2006) the viability was evaluated. Data were analysed with ANOVA, using the general linear model (GLM) of SAS (SAS Inst Inc., Cary, NC, USA). Sources of variation in the model included FCS concentration and first-order interactions; all factors were considered fixed effects. The arcsine transformation (√y/100) was applied to percentage data. If the ANOVA was significant, means were separated using the Tukey test. There was no difference in cleavage (for zebu embryos: Group 0%: 87.2 ± 6.8; Group 2.5%: 87.4 ± 9.5; for cross-bred embryos: Group 0%: 79.6 ± 11.9; Group 2.5%: 73.1 ± 13.7; P > 0.05). On the other hand, zebu embryos cultured in the presence of FCS reached blastocysts at a higher rate than cross-bred embryos in the absence of FCS (for zebu embryos: Group 0%: 33.3 ± 12.4ab; Group 2.5%: 46.8 ± 13.2a; for cross-bred embryos: Group 0%: 21.8 ± 8.3b; Group 2.5%: 33.6 ± 10.1ab; P < 0.05). After vitrification and warming, no significant differences in re-expansion rate (zebu embryos: Group 0%: 82.7 ± 13.1; Group 2.5%: 75.0 ± 9.8; cross-bred embryos: Group 0%: 93.7 ± 8.8; Group 2.5%: 84.1 ± 11.3; P > 0.05) and cell number per embryo (zebu embryos: Group 0%: 65.1 ± 34.7; Group 2.5%: 42.6 ± 17.2; cross-bred embryos: Group 0%: 64.3 ± 44.2; Group 2.5%: 52.0 ± 31.5; P > 0.05) between species groups and within species were seen. However for zebu embryos, Group 0% showed a lower damaged cell rate than Group 2.5%. The same effect was not observed in the cross-bred embryos (zebu embryos: Group 0%: 20.3 ± 22.7c; Group 2.5%: 63.3 ± 27.0d; cross-bred embryos: Group 0%: 25.4 ± 24.3cd; Group 2.5%: 45.8 ± 34.6cd; P < 0.05). The addition of 2.5% FCS had a higher deleterious effect on zebu embryos than cross-bred (zebu × taurine) embryos after vitrification. These results also reinforce the species differences observed between zebu and cross-bred, as they behaved differently in relation to the addition of FCS in the culture medium and in relation to their cryopreservation sensitivity. Supported by FAPESP 10/50410-2.

Forskolin effect on the cryosurvival ofin vitro-produced bovine embryos in the presence or absence of fetal calf serum

“…This factor is one of the reasons that led to our experiment to highlight the cleavage and blastocyst rates compared with the results in the literature. In addition, these results reinforce the differences in breeds observed between zebu and crossbreed, as they behaved differently in relation to the addition of FCS to the culture medium (Paschoal et al ., 2011).…”

Tauroursodeoxycholic Acid Supplementation in In Vitro Culture of Indicine Bovine Embryos: Molecular and Cellular Effects on the In Vitro Cryotolerance

“…These adverse factors negatively impact ER functions and protein synthesis and folding, resulting in the activation of ER stress and the UPR signaling pathways in in vitro-produced embryos [ 14 , 15 , 16 , 17 ]. Furthermore, there is evidence that IVPEs are more sensitive to cryopreservation than in vivo derived from embryos [ 34 , 35 ] and that this reduced cryotolerance may be associated with the high lipid content present in the cytoplasm as well as the lipid profile of the cell membrane of these embryos [ 28 , 29 , 34 ]. Although TUDCA has been demonstrated to exert efficient cytoprotective activity in relieving ER stress [ 17 , 18 , 20 , 26 ], recent studies reported on its new potential and molecular modes of action as a weight-reducing agent, modulating lipid metabolism through or independently of ER modulation [ 36 , 37 ].…”

“…The reduced post-warming cell viability is also associated with the abnormal amount and/or the type of lipids in the blastomeres that contributes to the occurrence of cryogenic fractures during the freezing process [ 28 , 34 ]. The cell membrane fluidity is related to the lipid profile and the capacity to support cryo-injuries during the cryopreservation process [ 34 ].…”

“…The reduced post-warming cell viability is also associated with the abnormal amount and/or the type of lipids in the blastomeres that contributes to the occurrence of cryogenic fractures during the freezing process [ 28 , 34 ]. The cell membrane fluidity is related to the lipid profile and the capacity to support cryo-injuries during the cryopreservation process [ 34 ]. The IVPEs have a different lipid profile from their in vivo-derived counterparts [ 34 , 35 ] and could possess a cryotolerance enhanced by changes in the lipid metabolism (e.g., TUDCA treatment).…”

“…The cell membrane fluidity is related to the lipid profile and the capacity to support cryo-injuries during the cryopreservation process [ 34 ]. The IVPEs have a different lipid profile from their in vivo-derived counterparts [ 34 , 35 ] and could possess a cryotolerance enhanced by changes in the lipid metabolism (e.g., TUDCA treatment). Although the evaluation of the lipid content was not assessed in this work, we cannot rule out that TUDCA treatment may have modulated the lipid content of the hatched embryo (down-regulated mRNA abundance of PLIN3 and SREBF2 in T200 group).…”