scholarly journals Effect of bovine oviductal extracellular vesicles on embryo development and quality in vitro

Reproduction ◽  
2017 ◽  
Vol 153 (4) ◽  
pp. 461-470 ◽  
Author(s):  
Ricaurte Lopera-Vasquez ◽  
Meriem Hamdi ◽  
Veronica Maillo ◽  
Alfonso Gutierrez-Adan ◽  
Pablo Bermejo-Alvarez ◽  
...  
Keyword(s):  
Gene Expression ◽  
Embryo Development ◽  
Extracellular Vesicles ◽  
Water Channel ◽  
Calf Serum ◽  
Bovine Embryos ◽  
Analysis System ◽  
Oviductal Fluid

The aim of this study was to evaluate the effect of extracellular vesicles (EV) from oviductal fluid (OF), either from the ampulla or isthmus, on the development and quality of in vitro-cultured bovine embryos. Zygotes were cultured in synthetic oviduct fluid (SOF + 3 mg/mL BSA) without calf serum (C− group), in the presence of 3 × 105 EV/mL from ampullary or isthmic OF at either 1 × 104 g (10 K) or 1 × 105 g (100 K), and compared with SOF + 5% FCS (C+ group). OF-EV size and concentration were assessed by electron microscopy and nanotracking analysis system. Embryo development was recorded on Days 7–9, and blastocyst quality was assessed through cryotolerance and gene expression analysis. Lower blastocyst yield was observed on Day 7 in the C− and OF-EV groups (12.0–14.3%) compared with C+ (20.6%); however, these differences were compensated at Days 8 and 9 (Day 9: 28.5–30.8%). Importantly, the survival rate of blastocysts produced with isthmic 100 K OF-EV was higher than that of C+ and C− group at 72 h after vitrification and warming (80.1 vs 34.5 and 50.5% respectively, P < 0.05). In terms of gene expression, blastocysts produced in the presence of 100 K isthmic OF-EV upregulated the water channel AQP3 and DNMT3A and SNRPN transcripts compared with the C+, with the expression in C− being intermediate. The lipid receptor LDLR was downregulated in C+ compared with all other groups. In conclusion, the addition of oviductal fluid extracellular vesicles from isthmus, to in vitro culture of bovine embryos in the absence of serum improves the development and quality of the embryos produced.

Effect of bovine oviductal fluid on development and quality of bovine embryos produced in vitro

2017 ◽  
Vol 29 (3) ◽  
pp. 621 ◽  
Author(s):  
Ricaurte Lopera-Vasquez ◽  
Meriem Hamdi ◽  
Veronica Maillo ◽  
Valeriano Lloreda ◽  
Pilar Coy ◽  
...  
Keyword(s):  
In Vitro Culture ◽  
Embryo Development ◽  
Cell Mass ◽  
Water Channel ◽  
Calf Serum ◽  
Cell Counting ◽  
Bovine Embryos ◽  
Oviductal Fluid

To evaluate the effect of bovine oviductal fluid (OF) supplementation during in vitro culture of bovine embryos on their development and quality, in vitro-produced zygotes were cultured in synthetic oviductal fluid (SOF; negative control; C–) supplemented with OF or 5% fetal calf serum (positive control; C+). Embryo development was recorded on Days 7–9 after insemination and blastocyst quality was assessed through cryotolerance, differential cell counting of the inner cell mass and trophectoderm, and gene expression. OF was added to the culture medium at concentrations ranging from 0.625% to 25%. The higher OF concentrations (5%, 10% and 25%) had a detrimental effect on embryo development. Lower OF concentrations (1.25% and 0.625%) supported embryo development until Day 9 (27.5%) and produced higher-quality blastocysts, as reflected by their cryotolerance (53.6% and 57.7% survival at 72 h, respectively, vs 25.9% in C+) and total cell number (mean (± s.e.m.) 165.1 ± 4.7 and 156.2 ± 4.2, respectively, vs 127.7 ± 4.9 in C– and 143.1 ± 4.9 in C+). Consistent with these data, upregulation of the water channel aquaporin 3 (AQP3) mRNA was observed in blastocysts supplemented with 1.25% OF compared with C– and C+. Serum supplementation resulted in a reduction in the expression of glucose and lipid metabolism-related genes and downregulation of the epigenetic-related genes DNA methyltransferase 3A (DNMT3A) and insulin-like growth factor 2 receptor (IGF2R). In conclusion, in vitro culture with low concentrations of OF has a positive effect on the development and quality of bovine embryos.

99 EXTRACELLULAR VESICLES OF BOVINE OVIDUCTAL FLUID MODIFY THE GENE EXPRESSION ON BOVINE IN VITRO-DERIVED EMBRYOS

2016 ◽  
Vol 28 (2) ◽  
pp. 179 ◽  
Author(s):  
R. Lopera-Vasquez ◽  
M. Hamdi ◽  
V. Maillo ◽  
C. Nunez ◽  
M. Yanez-Mo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Embryo Development ◽  
Extracellular Vesicles ◽  
Membrane Trafficking ◽  
Dna Methyltransferase ◽  
De Novo ◽  
Expression Patterns ◽  
Water Channel ◽  
Developmental Competence

Extracellular vesicles (EVs) act as intercellular communicators through their protein, lipid, and mRNA content. The interaction of EVs from oviducal environment and the first stages of embryo development is currently an enigma. The aim of the present study was to evaluate the developmental competence and the expression profile of bovine blastocysts cultured with previously purified EVs recovered from ampullary and isthmic oviducal fluid (OF) under different centrifugal forces. OF-EVs recovered from oviducts of slaughtered heifers in early luteal phase were quantified with a nanoparticle tracking analysis system, and their integrity and size were assessed by electron microscopy. In vitro-produced zygotes were cultured in SOF+3 mg mL–1 BSA (C–), C– with 3 × 105 OF-EVs/mL from the ampulla (A) and isthmus (I) isolated at 1 × 103 (A10k and I10k, respectively) and 1 × 105 (A100k and I100k, respectively) × g. A control culture group of SOF+5% FCS (C+) was included. Blastocyst development was recorded on Day 7, 8, and 9 (D0: day of fertilization). Blastocysts on Days 7/8 cultured in C–, C+, I10k, and I100k were used to measure the relative mRNA expression of genes related with membrane trafficking (AQP3, AQP11, and ATP1A1), metabolism (LDLR and LDHA), and epigenetics (DNMT3A, IGF2R, GRB10, and SNRPN) by RT-qPCR. One-way ANOVA was used for statistical analysis. The size of ampullary and isthmic OF-EVs was similar with a mean of 220 nm. The concentration of I10k was significantly lower compared with A100k (3.6 × 108 v. 10.5 × 108 EVs/mL, respectively; P < 0.05); however, no differences were found in the rest of the groups with a mean concentration of 7.6 × 108 EVs/mL. EVs and C– groups showed a delayed embryo development at Day 7 compared with C+ (range: 12.0–13.8 v. 20.6%, respectively, P < 0.05); however, it was compensated at Days 8 and 9 (Day 9 range: 28.5–30.8%). The water channel related protein AQP3, associated with blastocoel formation, water, and cryoprotectant movement during cryopreservation, was up-regulated in I10k and I100k blastocysts compared with C+. The lipid receptor LDLR, proposed as a regulator of lipid uptake in blastocysts, was significantly down-regulated in C+ compared with the other groups, a possible consequence of a higher concentration of lipids in the C+ group. The de novo DNA methyltransferase DNMT3A and the imprinting gene SNRPN were down-regulated in the C+ compared with I100k, suggesting alterations in imprinting. In conclusion, bovine isthmic OF-EVs supplementation in in vitro embryo culture has a positive effect on gene expression patterns of developmental related genes compared with serum supplementation, suggesting an association between the oviducal environment and the developing embryo. Funded by the Spanish Ministry of Science and Innovation (AGL2012–37510 and AGL2012–39652-C02–01).

Effects of bovine oviduct epithelial cells, fetal calf serum and bovine serum albumin on gene expression in single bovine embryos produced in the synthetic oviduct fluid culture system

2005 ◽  
Vol 17 (8) ◽  
pp. 751 ◽  
Author(s):  
Mona E. Pedersen ◽  
Øzen Banu Øzdas ◽  
Wenche Farstad ◽  
Aage Tverdal ◽  
Ingrid Olsaker
Keyword(s):  
Gene Expression ◽  
Fetal Calf Serum ◽  
Calf Serum ◽  
Developmental Competence ◽  
Bovine Embryos ◽  
Glut 1 ◽  
Significant Difference ◽  
Oviduct Fluid

In this study the synthetic oviduct fluid (SOF) system with bovine oviduct epithelial cell (BOEC) co-culture is compared with an SOF system with common protein supplements. One thousand six hundred bovine embryos were cultured in SOF media supplemented with BOEC, fetal calf serum (FCS) and bovine serum albumin (BSA). Eight different culture groups were assigned according to the different supplementation factors. Developmental competence and the expression levels of five genes, namely glucose transporter-1 (Glut-1), heat shock protein 70 (HSP), connexin43 (Cx43), β-actin (ACTB) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), analysed as mRNA by using reverse transcription–polymerase chain reaction, were measured on bovine embryos cultured for 9 days. Gene expression of these in vitro-produced embryos was compared with the gene expression of in vivo-produced embryos. There was no significant difference found in embryo developmental competence between the Day 9 embryos in BOEC co-culture, FCS and BSA supplements in SOF media. However, differences in gene expression were observed. With respect to gene expression in in vivo and in vitro embryos, BOEC co-culture affected the same genes as did supplementation with FCS and BSA. HSP was the only gene that differed significantly between in vitro and in vivo embryos. When the different in vitro groups were compared, a significant difference between the BOEC co-culture and the FCS supplementation groups due to Glut-1 expression was observed.

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

Zygote ◽  
2012 ◽  
Vol 22 (2) ◽  
pp. 146-157 ◽  
Author(s):  
Daniela Martins Paschoal ◽  
Mateus José Sudano ◽  
Midyan Daroz Guastali ◽  
Rosiára Rosária Dias Maziero ◽  
Letícia Ferrari Crocomo ◽  
...  
Keyword(s):  
Culture Medium ◽  
Fetal Calf Serum ◽  
Calf Serum ◽  
Similar Rate ◽  
Bovine Embryos ◽  
Embryo Survival ◽  
Vitrification Solution ◽  
Oviductal Fluid

SummaryThe objective of this study was to assess the viability and cryotolerance of zebu embryos produced in vitro with or without the addition of fetal calf serum (FCS) and forskolin (F). Embryos produced in vivo were used as a control. Presumptive zygotes were cultured in modified synthetic oviductal fluid supplemented with amino acids (SOFaa), bovine serum albumin (BSA) and with (2.5%) or without (0%) FCS. On day 6 of growth, the embryos from each group were divided into treatments with or without 10 μM F to induce embryonic lipolysis, comprising a total of four experimental groups: 2.5% FCS, 0% FCS, 2.5% + F and 0% + F. For vitrification, embryos were exposed to vitrification solution 1 (5 M EG (ethylene glycol)) for 3 min and then transferred to vitrification solution 2 (7 M EG, 0.5 M galactose solution and 18% (w/v) Ficoll 70) before being introduced to liquid nitrogen. The presence of FCS in the culture medium resulted in the production of embryos with a similar rate of damaged cells compared with in vivo-produced embryos. After vitrification, the 2.5% FCS group had a significantly higher rate of damaged cells when compared with the other groups (P < 0.05). The results of this experiment indicated that the omission of FCS and the addition of forskolin do not have deleterious effect on embryo production rates. In addition, embryos produced in the presence of FCS had greater sensitivity to cryopreservation, but this effect was reversed when forskolin was added to the medium, which improved embryo survival without affecting embryo development and quality after vitrification.

scholarly journals Bovine Follicular Fluid and Extracellular Vesicles Derived from Follicular Fluid Alter the Bovine Oviductal Epithelial Cells Transcriptome

2020 ◽  
Vol 21 (15) ◽  
pp. 5365 ◽  
Author(s):  
Mohammad Mehedi Hasan ◽  
Janeli Viil ◽  
Freddy Lättekivi ◽  
James Ord ◽  
Qurat Ul Ain Reshi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Embryo Development ◽  
Extracellular Vesicles ◽  
Follicular Fluid ◽  
Early Embryo ◽  
Transcriptional Responses ◽  
Early Embryo Development ◽  
Sperm Survival

While follicular fluid (FF) is well known to provide an optimal environment for oogenesis, its functional roles following its release into the oviduct during ovulation are currently elusive. We hypothesized that FF and FF-derived extracellular vesicles (EVs) may be conveyors of signals capable of inducing functionally-relevant transcriptional responses in oviductal cells. The aim of this study was, therefore, to evaluate the effect of FF and FF-derived EVs on the transcriptome of primary bovine oviductal epithelial cells (BOECs). We examined the gene expression of BOECs in three conditions: BOECs cultured with FF, FF-derived EVs, and without supplementations. For each condition, cells were cultured for 6 and 24 h. RNA sequencing results revealed that FF had a stronger effect on BOECs gene expression compared to EVs. We detected 488 and 1998 differentially expressed genes (DEGs) with FF treatment in 6 and 24 h, respectively, whereas only 41 DEGs were detected at 6 h following EV treatment. Pathway analysis of the FF-induced DEGs showed that several pathways were highly enriched, notably oxidative phosphorylation, thermogenesis, arachidonic acid metabolism, and steroid hormone biosynthesis. Some of these pathways have a role in sperm survival, fertilization, and early embryo development. In conclusion, the findings of our study demonstrate for the first time that bovine FF and FF-derived EVs can induce changes in the gene expression of the bovine oviductal cells which, although observed in vitro, may be reflective of in vivo responses which may contribute to a favorable periconceptional microenvironment for sperm survival, fertilization, and early embryo development.

Production of in vitro bovine embryos supplemented with l-carnitine in different oxygen tensions and the relation to nitric oxide

Zygote ◽  
2020 ◽  
Vol 28 (5) ◽  
pp. 403-408
Author(s):  
Daniela Moraes Pereira ◽  
Christopher Junior Tavares Cardoso ◽  
Wilian Aparecido Leite da Silva ◽  
Mirela Brochado Souza-Cáceres ◽  
Mariana Santos ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Embryo Development ◽  
Calf Serum ◽  
Hatching Rate ◽  
Bovine Embryos ◽  
Low Oxygen ◽  
Cleavage Rate ◽  
Treatment Groups ◽  
Oxygen Tensions

SummaryThe aim of this study was to evaluate the production of bovine embryos in vitro when supplemented with l-carnitine for 24 h beginning on day 5 (d 5) under two different oxygen tensions (20% or 5%) and the relationship of nitric oxide (NO) in in vitro culture (IVC) medium to embryo development. Cumulus–oocyte complexes (COC; n = 837) were matured in vitro for 24 h and fertilization was performed for 18 h. Zygotes were cultured in vitro for 9 days after in vitro fertilization in synthetic oviductal fluid (SOF) medium with 5% fetal calf serum. At d 5 the plates were assigned to one of four treatment groups: high (20%) or low (5%) O2 tension either with or without the addition of 3.03 mM l-carnitine (High-Cont, High-Lcar, Low-Cont, Low-Lcar). The concentration of NO in the culture medium was evaluated on d 5, d 6 and d 9. On d 7, parts of the embryos were submitted for evaluation of intracellular lipid droplets. The cleavage rate was similar (P > 0.05) between high and low O2 tension and the blastocyst rate was similar in all conditions evaluated. The hatching rate was higher (P < 0.05) for Low-Cont. The NO concentration was higher at d 9 under low O2 tension (P < 0.1). The addition of 3.03 mM l-carnitine between d 5 and d 6 of IVC was not efficient in reducing cytoplasmic lipid content of bovine embryos. Additionally, IVC at a low oxygen tension without l-carnitine promoted better conditions for embryo development. A higher concentration of NO in medium was observed under low O2 tension.

56 Nobiletin affects gene expression profiles of the ERK1/2 pathway in bovine embryos produced invitro

2021 ◽  
Vol 33 (2) ◽  
pp. 135
Author(s):  
Y. N. Cajas ◽  
K. E. Cañón-Beltrán ◽  
C. L. V. Leal ◽  
A. Gutierrez-Adán ◽  
E. González ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Cell Cycle ◽  
Embryo Development ◽  
Expression Profiles ◽  
Calf Serum ◽  
Early Embryo ◽  
Bovine Embryos ◽  
Cell Stage ◽  
And Oxidative Stress

During embryo development the embryonic genome activation (EGA) is one of the most important events and in bovine embryos it occurs at the 8- to 16-cell stage. Invitro embryo production increases the levels of reactive oxygen species (ROS), which leads to the low quality of the produced blastocysts, possibly by affecting EGA. Nobiletin is an antioxidant that affects cell cycle regulation (Huang et al. 2016 Evid. Based. Complement. Alternat. Med. 2016, 2918796, https://doi.org/10.1155/2016/2918796). Therefore, we aimed to evaluate the effect of nobiletin supplementation, in two key periods of early embryo development, on blastocyst yield and expression of selected genes of the ERK1/2 pathway and oxidative stress on produced embryos. Invitro zygotes were cultured in synthetic oviductal fluid (SOF) with 5% fetal calf serum (control, C); C with 5 or 10µM nobiletin (MedChemExpress) (N5, N10); or C with 0.03% dimethyl sulfoxide (CDMSO; vehicle for nobiletin dilution) during the minor (21–54h post-insemination (hpi): 2- to 8-cell; MNEGA; 12 replicates) or major (54–96 hpi: 8- to 16-cell; MJEGA; 10 replicates) phase of EGA. The speed of development was considered and embryos that reached ≥8 cells at 54 hpi from MNEGA phase and ≥16 cells at 96 hpi from MJEGA phase, were selected and further cultured in control medium until Day 7. Embryos at ≥8 cell (MNEGA), ≥16 cell (MJEGA) stage, and Day 7 blastocysts from both periods were snap-frozen in liquid N2 for gene expression analysis (3 pools of 10 embryos/treatment). The expression of genes related to ERK1/2 pathway (H3–3B, H3–3A, NFE2L2) and oxidative stress (GPX1) were measured by quantitative PCR; H2AFZ and ACTB were used as housekeeping genes. Statistical analysis was assessed by one-way ANOVA. At 54 hpi, irrespective of nobiletin supplementation, no differences were found in the proportion of embryos that reached the 8-cell stage between groups in both phases (≈60%). At 96 hpi, nobiletin during MJEGA showed a higher proportion of embryos reaching the 16-cell stage than control groups (≈70% vs. ≈60%, respectively; P&lt;0.001). Blastocyst yield for MNEGA and MJEGA was higher (P&lt;0.001) for N5 (40.0±0.8% and 46.7±0.8%) and N10 (41.0±0.9% and 54.5±1.1%) compared with C (32.0±0.6% and 38.4±1.1%) and CDMSO (31.2±0.4% and 35.8±1.0%) groups, while N10 was higher (P&lt;0.05) compared to N5 group in MJEGA. The expression of H3–3B and H3–3A were higher (P&lt;0.05) in 8-cell embryos from N5 and N10 groups during MNEGA; while in 16-cell embryos, H3–3B and NFE2L2 were higher (P&lt;0.05) only in the N10 group compared with both controls during MJEGA. GPX1 was upregulated in nobiletin-supplemented groups from both phases (8- and 16-cell embryos and blastocysts) compared with controls (P&lt;0.05). In conclusion, nobiletin supplementation during minor or major EGA has a positive effect in pre-implantation embryo development and modifies the transcription of cell cycle and oxidative stress genes in early embryos. These benefits can be attributed to its bioactivity and indicate that it might be a tool to overcome EGA and ROS disorders in bovine invitro-produced embryos.This research was funded by MINECO-Spain AGL2015-70140-R, PID2019-111641RB-I00, RTI2018-093548-B-I00; SENESCYT-Ecuador; FAPESP-Brazil 2017/20339-3, CNPq-Brazil 304276/2018-9.

scholarly journals 160 IN VIVO-CULTURE OF BOVINE EMBRYOS: TRANSFER OF SEMEN PRE-INCUBATED OOCYTES, ZYGOTES AND 4 TO 8 CELL STAGE EMBRYOS INTO THE BOVINE OVIDUCT

2005 ◽  
Vol 17 (2) ◽  
pp. 231
Author(s):  
V. Havlicek ◽  
F. Wetscher ◽  
T. Huber ◽  
M. Gilles ◽  
D. Tesfaye ◽  
...  
Keyword(s):  
Embryo Development ◽  
Calf Serum ◽  
Bovine Embryos ◽  
Cell Stage ◽  
Group Iii ◽  
Group I ◽  
In Vivo Culture ◽  
Group Ii

Oviduct as well as oocyte and embryo development are subject to developmental changes which have crucial effects on the application of in vivo culture. The present study aimed at optimizing in vivo culture of IVP bovine embryos at different developmental stages in the bovine oviduct. Cumulus oocyte complexes (COC) were collected from slaughterhouse ovaries, matured in vitro for 22 h and assigned to four groups. In groups I and II, oocytes were pre-incubated for 3 to 4 h with 5 × 106 sperm/mL, and then immediately transferred to recipients, which had just completed ovulation (group I), or kept in vitro for a further 12 to 18 h and transferred to Day 1 synchronized recipients (group II). In groups III and IV, COC were subjected to standard IVF/IVC; then embryos were either transferred at the 4- to 8-cell stage on Day 3 into the oviducts of Day 3-synchronized recipients (group III) or kept in vitro for a further 4 to 5 days (group IV). Thirty-four 18- to 30-month-old temporary recipients were synchronized using a standard Ovsynch protocol. COC and embryos were transferred and re-collected by transvaginal endoscopy. COC or embryos were loaded into a 180° curved glass capillary, which was inserted via the infundibulum 5 to 8 cm deep into the ampulla ipsilateral to the CL. On recipient Day 7, a 90° curved metal canula served for tubal flushing prior to conventional uterine embryo flushing. Sixty mL of PBS containing 1% fetal calf serum were rinsed through the oviduct into the uterus and a further 400 mL of medium were finally used for flushing of the uterine horn and collected via an embryo filter. Embryo development was evaluated directly after flushing (Day 7) and on Day 8. For statistical analysis (ANOVA), the blastocyst rates (Days 7 and 8) in group III were related to COC corrected by the collection rate. In group I, 575 COC were transferred to 11 recipients and 420 (73%) were re-collected as oocytes or embryos. The blastocyst yields on Day 7 and Day 8 were 23% (97) and 25% (104), respectively. In group II, the transfer of 489 presumptive zygotes into 13 heifers resulted in only 175 re-collected (36%), of which 15% developed into blastocysts (Day 7: 26; Day 8: 27). Ten heifers (group III) served for in vivo culture of 643 embryos at the 4- to 8-cell stage. On Day 7, 568 (88%) embryos were flushed and 171 (30%) reached the blastocyst stage. A further 24 h culture in vitro finally resulted in 244 (42%) blastocysts. The complete in vitro production system delivered 13% (63/477) blastocysts on Day 7 and 34% (161/477) blastocysts on Day 8. The collection rates (P < 0.001) and the blastocyst rates on Day 7 (P < 0.05) and Day 8 (P < 0.001) differed significantly in all groups. The present data demonstrate that the developmental stage of transferred complexes has an influence on embryo recovery as well as an embryo development. This work was supported by Austrian BMBWK and BMLFUW (#1227).

137 PHENAZINE ETHOSULFATE AND FETAL CALF SERUM EFFECTS ON THE DEVELOPMENT AND APOPTOSIS OF IN VITRO PRODUCED BOVINE EMBRYOS

2011 ◽  
Vol 23 (1) ◽  
pp. 173
Author(s):  
M. J. Sudano ◽  
D. M. Paschoal ◽  
T. S. Rascado ◽  
L. C. O. Magalhães ◽  
L. F. Crocomo ◽  
...  
Keyword(s):  
Embryo Development ◽  
Culture Medium ◽  
Fetal Calf Serum ◽  
Bos Indicus ◽  
Calf Serum ◽  
Bovine Embryos ◽  
Blastocyst Quality ◽  
Phenazine Ethosulfate

Phenazine ethosulfate (PES) is a metabolic regulator that inhibits fatty acid synthesis and favours the pentose-phosphate pathway. Supplementation of fetal calf serum (FCS) during culture has been correlated with the reduction of quality of in vitro produced bovine embryos (IVPE). The aim of the present study was to evaluate embryo development and apoptosis in blastocysts after the supplementation of PES and FCS in culture medium of IVPE. Oocytes (N = 4320) were matured and fertilized in vitro (Day 0). The zygotes (Bos indicus) were cultured in SOFaa medium with 4 concentrations of FCS (0, 2.5, 5, and 10%) and with the use or not of 0.3 μM PES from Day 4 (after 96 h of embryo culture). Embryo development was evaluated after 7 days of culture. Apoptosis in blastocysts (N = 60–80) was accessed through TUNEL reaction. Embryos (Bos indicus) recovered from superstimulated cows were used as in vivo control (n = 15). Data were analysed by ANOVA followed by LSD using PROC GLIMMIX (SAS; SAS Institute Inc., Cary, NC, USA) means ± SEM. Increasing FCS concentration in the culture media did not change cleavage (86.7 ± 1.7, 82.3 ± 1.6, 86.3 ± 1.4, 87.0 ± 1.5, P > 0.05) and augmented blastocyst production (30.5 ± 2.5a, 41.8 ± 2.4b, 40.5 ± 2.6b, 47.2 ± 2.8b, P < 0.05), respectively, for 0, 2.5, 5, and 10%. Additionally, increasing FCS concentration increased apoptosis in blastocysts (13.8 ± 1.2b, 19.1 ± 1.8b, 20.7 ± 1.9bc, 28.4 ± 2.3c, P < 0.05, respectively, for 0, 2.5, 5, and 10%). The addition of PES from Day 4 in the culture medium did not affect (P > 0.05) cleavage (87.0 ± 1.3 and 84.4 ± 1.3), blastocyst production (42.0 ± 2.8 and 43.0 ± 2.0), and apoptosis in blastocysts (20.7 ± 2.0b and 18.9 ± 2.1b), respectively, for control and PES Day 4 groups. Independent of FCS withdrawal or PES addition to culture medium, the in vivo control group presented the lowest apoptosis rate (6.3 ± 1.1a). Therefore, increasing FCS concentration augmented embryo development and reduced blastocyst quality. However, the addition of 2.5% of FCS in the culture medium increased the embryo development without the reduction of blastocyst quality. Moreover, the PES supplementation from Day 4 did not affect embryo development and blastocyst quality. São Paulo Research Foundation – FAPESP.

Sign in / Sign up

Export Citation Format

Share Document