[Show abstract][Hide abstract] ABSTRACT: To identify embryos individually during in vitro development, we previously developed the well-of-the-well (WOW) dish, which contains 25 microwells. Here we investigated the effect of embryo density (the number of embryos per volume of medium) on in vitro development and gene expression of bovine in vitro-fertilized embryos cultured in WOW dishes. Using both conventional droplet and WOW culture formats, 5, 15, and 25 bovine embryos were cultured in 125 μl medium for 168 h. The blastocysts at Day 7 were analyzed for number of cells and expression of ten genes (CDX2, IFN-tau, PLAC8, NANOG, OCT4, SOX2, AKR1B1, ATP5A1, GLUT1 and IGF2R). In droplet culture, the rates of formation of >4-cell cleavage embryos and blastocysts were significantly lower in embryos cultured at 5 embryos per droplet than in those cultured at 15 or 25 embryos per droplet, but not in WOW culture. In both droplet and WOW culture, developmental kinetics and blastocyst cell numbers did not differ among any groups. IFN-tau expression in embryos cultured at 25 embryos per droplet was significantly higher than in those cultured at 15 embryos per droplet and in artificial insemination (AI)-derived blastocysts. Moreover, IGF2R expression was significantly lower in the 25-embryo group than in the 5-embryo group and in AI-derived blastocysts. In WOW culture, these expressions were not affected by embryo density and were similar to those in AI-derived blastocysts. These results suggest that, as compared with conventional droplet culture, in vitro development and expression of IFN-tau and IGF2R in the microwell system may be insensitive to embryo density.
Journal of Reproduction and Development 11/2012; 59(2). DOI:10.1262/jrd.2012-113 · 1.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mitochondrial bioenergetics in mammalian oocytes has not been sufficiently characterized. In this study, the function of oxidative phosphorylation (OXPHOS), a major pathway in mitochondria, was investigated in individual bovine oocytes by monitoring oxygen consumption using modified scanning electrochemical microscopy (SECM). At the germinal vesicle (GV) stage, 65% of basal respiration was used for mitochondrial respiration, which was inhibited by complex IV inhibitor. Around 63% of mitochondrial respiration was coupled to ATP synthesis, as determined by sensitivity to an ATP synthase inhibitor, and the remaining 37% was attributed to proton leak. In contrast, 50% and 43% of mitochondrial respiration were used for ATP synthesis in in vivo- and in vitro-derived metaphase II (MII)-stage oocytes, respectively. ATP-linked respiration, in both in vivo- and in vitro-derived MII-stage oocytes, was significantly lower than in GV-stage oocytes, suggesting that OXPHOS in bovine oocytes is more active at the GV stage compared with the MII stage. Interestingly, basal respiration in in vitro-derived MII oocytes was significantly higher than for in vivo-derived oocytes, reflecting an increase in proton leak. Next, we assessed respiration in MII oocytes cultured for 8 h. The aged oocytes had a significantly reduced maximum respiratory capacity, which was stimulated by a mitochondrial uncoupler, and reduced ATP-linked respiration compared with non-aged oocytes. However, the aging-related phenomenon could be prevented by caffeine treatment. We conclude that OXPHOS in bovine oocytes varies in the transition from GV to MII stage, in vitro maturation and the aging process. This approach will be particularly useful for analyzing mitochondrial bioenergetics in individual mammalian oocytes.
Journal of Reproduction and Development 07/2012; 58(6). DOI:10.1262/jrd.2012-082 · 1.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Conventionally, in vitro-fertilized (IVF) bovine embryos are morphologically evaluated at the time of embryo transfer to select those that are likely to establish a pregnancy. This method is, however, subjective and results in unreliable selection. Here we describe a novel selection system for IVF bovine blastocysts for transfer that traces the development of individual embryos with time-lapse cinematography in our developed microwell culture dish and analyzes embryonic metabolism. The system can noninvasively identify prognostic factors that reflect not only blastocyst qualities detected with histological, cytogenetic, and molecular analysis but also viability after transfer. By assessing a combination of identified prognostic factors--(i) timing of the first cleavage; (ii) number of blastomeres at the end of the first cleavage; (iii) presence or absence of multiple fragments at the end of the first cleavage; (iv) number of blastomeres at the onset of lag-phase, which results in temporary developmental arrest during the fourth or fifth cell cycle; and (v) oxygen consumption at the blastocyst stage--pregnancy success could be accurately predicted (78.9%). The conventional method or individual prognostic factors could not accurately predict pregnancy. No newborn calves showed neonatal overgrowth or death. Our results demonstrate that these five predictors and our system could provide objective and reliable selection of healthy IVF bovine embryos.
PLoS ONE 05/2012; 7(5):e36627. DOI:10.1371/journal.pone.0036627 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We examined the influence of recipient oocytes on in vitro development, oxygen consumption, and gene expression in the resulting cloned bovine embryos. Oocytes derived from slaughterhouse ovaries and ovum pickup (OPU)-derived oocytes were used as recipient cytoplasts for the production of cloned embryos. A series of OPU sessions was conducted on Holstein cows without follicular growth treatment (FGT). In the same cows, we then performed dominant follicle ablation and subsequently administered follicle-stimulating hormone and prostaglandin F(2α) with controlled internal drug release device before a second series of OPU. Cumulus cells collected from single Holstein cows were used as donor cells. After measurement of oxygen consumption at the blastocyst stage with modified scanning electrochemical microscopy, analysis of 10 genes (CDX2, IFN-tau, PLAC8, OCT4, SOX2, NANOG, ATP5A1, GLUT1, AKR1B1, and IGF2R) was performed with real-time RT-PCR. Rates of fusion, cleavage, and blastocyst formation were not different among the treatment groups. Levels of oxygen consumption in cloned blastocysts derived from slaughterhouse ovaries or OPU without FGT were significantly lower than in blastocysts derived from artificial insemination (AI). However, oxygen consumption was increased in cloned blastocysts derived from OPU with FGT, depending on the individual oocyte donor. Furthermore, gene expression of IFN-tau and OCT4 in cloned blastocysts derived from OPU with FGT was similar to that in AI-derived blastocysts, whereas expression of those genes in cloned blastocysts derived from slaughterhouse ovaries or OPU without FGT was significantly different from that in AI-derived blastocysts. Thus, recipient oocytes collected by OPU in combination with manipulation of follicular growth in donor cows are suitable for producing cloned embryos.
[Show abstract][Hide abstract] ABSTRACT: More than 300000 embryos have been transferred all over the world (Stroud 2010 IETS Newsl. 27(4), 11-21). We have reported that embryos that showed the abnormal cleavage pattern at the first cell division can develop to the blastocyst stage (Somfai et al. 2010 J. Reprod. Dev. 56, 200-207). However, we have limited knowledge about the consequences of the pattern of first embryonic cleavage on their post-transfer developmental competence. The present study was conducted to determine the developmental competence of bovine blastocysts showing different cleavage patterns at their first cell division. Cumulus-oocyte complexes were collected by ovum pickup from Japanese Black cows and were subjected to in vitro maturation and IVF as reported previously (Imai et al. 2006 J. Reprod. Dev. 52, S19-S29 suppl). Inseminated oocytes were cultured in CR1aa medium supplemented with 5% calf serum covered by mineral oil at 38.5°C in 5% CO(2) in air with micro-droplets or 5% CO(2), 5% O(2) and 90% N(2). The kinetics of embryo development were analysed by time-lapse cinematography for 168h after IVF by using a Cultured Cell Monitoring System (CCM-M1.4ZS, Astec, Fukuoka, Japan). A total of 673 photographs of each embryo were taken (1 photograph in every 15min) during in vitro culture. Image stacks were analysed by the CCM-M1.4 software. Embryos were classified in 5 groups according to the pattern of first cleavage as normal cleavage (NC), direct cleavage from 1 cell to 3 to 4 blastomeres (3-4BL), unequal blastomeres (UB), multiple fragments (MF) and protrusion formation (PT). Blastocysts developing from each group were transferred into the ipsilateral uterine horn of each synchronized recipient on Day 7 or 8 after oestrus. Data on conception at Day 60, abortion and delivery were then recorded. Data were analysed by chi-square test and Student's t-test. In total, 43 embryos were transferred, 17 conceptions (39.5%) were established and 16 recipients (94.1%) were delivered. Only 1 abortion was detected at Day 223 in the NC group. The highest conception rate was observed in the NC group (55%, n=20) and the 3-4BL (n=12), UB (n=6) and PT (n=3) groups showed similar conception rates of 33.3% (1 implanted embryo belonged to 2 classes in UB and PT) and none of the embryos derived from the MF group (n=3) could cause conception. There was a significant difference (P<0.05) in conception rates between the NC group and totals of each of the other cleavage groups. No significant difference was found in gestation lengths and birth weights between the NC group (282.2±4.4 days, 30.6±3.8kg, respectively) and totals of each of the other cleavage groups (282.8±5.3 days, 30.3±1.9kg, respectively). These results indicate that embryos showing abnormal cleavage patterns at first cell division can develop to normal calves with normal gestation lengths and birth weights; however, their post-transfer viability is lower than for NC embryos.
Reproduction Fertility and Development 12/2011; 24(1):191-2. DOI:10.1071/RDv24n1Ab159 · 2.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In cattle, the prediction of embryonic viability after embryo transfer is an important research target. A previous study has indicated that the duration of the fourth cell cycle at the time of maternal-zygotic transition, which is involved in in vitro embryonic development, may be an indicator of blastocyst formation; this study showed that embryos with a short fourth cell cycle have a better potential of developing into blastocysts than those with a long fourth cell cycle (Lequarre et al. 2003 Biol. Reprod. 69, 1707-1713). However, the relationship between the fourth cell cycle duration and post-transfer viability of embryos is unclear. The aim of the present study was to examine the effect of the fourth cell cycle duration on embryo development after embryo transfer. Twenty-five IVF bovine embryos were cultured in well-of-the-well culture dishes contained 125 μ of CR1aa supplemented with 5% calf serum at 38.5°C in 5% O(2) and 5% CO(2) for 168h after insemination. In vitro development of the embryos was monitored using time-lapse cinematography (Sugimura et al. 2010 Biol. Reprod. 83, 970-978). We found that 61% of the blastocysts had a long fourth cell cycle (41.5±5.9h), which is commonly referred to as the lag phase, whereas the remaining embryos had a short fourth cell cycle (7.4±4.5h). All the embryos with a short fourth cell cycle exhibited a lag phase in the next cell cycle (32.9±6.6h). Moreover, embryos with a short fourth cell cycle were found to have a higher blastocyst rate (75.8%) than those with a long fourth cell cycle (48.1%; Student's t-test, P<0.01). However, embryonic cell number, apoptosis incidence, chromosomal abnormality and O(2) consumption were found to be identical between the 2 groups (Student's t-test, P>0.05). Real-time reverse-transcription PCR results of the individual blastocysts showed that the relative expression of 5 genes related to pregnancy reorganization, placentation and fetal growth-namely, CDX2, IFN-τ, PLAC8, AKR1B1 and IGF2R-did not differ between the 2 groups (Student's t-test, P>0.05). Furthermore, blastocysts derived from embryos with long (n=30) and short (n=19) fourth cell cycles were transferred into 49 recipient cows; we did not observe any difference between the long and short fourth cell cycles on the rates of pregnancy (long vs short fourth cell cycle, 30.0 vs 52.6%) and delivery (long vs short fourth cell cycle, 30.0 vs 47.4%; Yates' corrected chi-square test, P>0.10). These results show that blastocysts derived from embryos with either long or short fourth cell cycles have identical developmental competence after embryo transfer. Therefore, the fourth cell cycle duration during maternal-zygotic transition appears to be unavailable as the indicator of post-transfer viability of IVF bovine embryos.
Reproduction Fertility and Development 12/2011; 24(1):191. DOI:10.1071/RDv24n1Ab158 · 2.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Changes in sensory traits of longissimus muscle (LM) from 20-30-month-old cattle were investigated using somatic cell clones of Japanese black steers slaughtered at 20-, 25- and 30-months-old (n=3, 4 and 4 respectively). The fat content of LM samples at 20, 25 and 30 months were 23.7, 38.7 and 41.1%, respectively. Soluble collagen content and collagen solubility at 20 months was greater than at 25 and 30 months. In terms of sensory traits, initial tenderness and juiciness at 25 months was greater than at 20 months, and fattiness at 25 and 30 months was greater than at 20 months. These results demonstrate that the changes in physicochemical traits of beef accompanying the differences in slaughter age affect the sensory traits although the desirable effects of the sensory traits do not continue throughout the entire fattening period.
[Show abstract][Hide abstract] ABSTRACT: The reproductive ability, milk-producing capacity, survival time and relationships of these parameters with telomere length were investigated in 4 groups of cows produced by somatic cell nuclear transfer (SCNT). Each group was produced using the same donor cells (6 Holstein (1H), 3 Holstein (2H), 4 Jersey (1J) and 5 Japanese Black (1B) cows). As controls, 47 Holstein cows produced by artificial insemination were used. The SCNT cows were artificially inseminated, and multiple deliveries were performed after successive rounds of breeding and conception. No correlation was observed between the telomere length and survival time in the SCNT cows. Causes of death of SCNT cows included accidents, accident-associated infections, inappropriate management, acute mastitis and hypocalcemia. The lifetime productivity of SCNT cows was superior to those of the controls and cell donor cows. All SCNT beef cows with a relatively light burden of lactation remained alive and showed significantly prolonged survival time compared with the cows in the SCNT dairy breeds. These results suggest that the lifetime productivity of SCNT cows was favorable, and their survival time was more strongly influenced by environmental burdens, such as pregnancy, delivery, lactation and feeding management, than by the telomere length.
Journal of Reproduction and Development 06/2011; 57(5):572-8. DOI:10.1262/jrd.10-174A · 1.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to develop an in-straw dilution method suitable for 1-step bovine embryo transfer of vitrified embryos using the Cryotop vitrification-straw dilution (CVSD) method. The development of embryos vitrified using the CVSD method was compared with those of embryos cryopreserved using in-straw vitrification-dilution (ISVD) and conventional slow freezing, outside dilution of straw (SFODS) methods. In Experiment 1, in vitro-produced (IVP) embryos cryopreserved using the CVSD method were diluted, warmed and exposed to the dilution solution at various times. When vitrified IVP embryos were exposed to the dilution solution for 30 min after warming, the rates of embryos developing to the hatched blastocyst stage after 72 h of culture (62.0-72.5%) were significantly lower (P<0.05) than those of embryos exposed to the solution for 5 and 10 min (82.4-94.3%), irrespective of supplementation with 0.3 M sucrose in the dilution solution. In Experiment 2, the rate of embryos developing to the hatching blastocyst stage after 48 h of culture in IVP embryos cryopreserved using the SFODS method (75.0%) was significantly (P<0.05) lower than those of embryos cryopreserved using the CVSD and ISVD methods (93.2 and 97.3%, respectively). In Experiment 3, when in vivo-produced embryos that had been cryopreserved using the CVSD, ISVD and SFODS methods and fresh embryos were transferred to recipient animals, no significant differences were observed in the conception and delivery rates among groups. In Experiment 4, when IVP embryos derived from oocytes collected by ovum pick-up that had been cryopreserved using the CVSD and ISVD methods and fresh embryos were transferred to recipient animals, no significant differences were observed in the conception rates among groups. Our results indicate that this simplified regimen of warming and diluting Cryotop-vitrified embryos may enable 1-step bovine embryo transfer without the requirement of a microscope or other laboratory equipment.
Journal of Reproduction and Development 04/2011; 57(4):437-43. DOI:10.1262/jrd.10-154M · 1.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We have developed a polystyrene-based well of-the-well system (WOW) using injection moulding to track individual embryos throughout culture using time-lapse cinematography (TLC). The WOW-cultured bovine embryos following in vitro fertilization (IVF) were compared with conventional droplet (control)-cultured embryos on in vitro and in vivo development. Twenty-five of zygotes were cultured in each culture system containing 125μL of CR1aa medium supplemented with 5% calf serum for 168h after IVF. No differences between control- and WOW-cultured embryos were observed during development to the blastocyst stage. Morphological quality, inner cell mass (ICM), and trophectoderm (TE) cell numbers and post-vitrification survival rates were not different between control- and WOW-derived blastocysts; however, incidence of apoptosis in the ICM and TE cells was reduced in WOW culture (P<0.01). Oxygen consumption in WOW-derived blastocysts was closer to physiological level than that of control-derived blastocysts. Moreover, WOW culture improved embryo viability, as indicated by increased pregnancy rates at days 30 and 60 after embryo transfer (P<0.05). The TLC monitoring was performed to evaluate the cleavage pattern and the duration of the first cell cycle of embryos from oocytes collected by ovum pick-up; correlations with success of pregnancy were determined. Logistic regression analysis indicated that the cleavage pattern correlated with success of pregnancy (P<0.05), but cell cycle length did not. Higher pregnancy rates (66.7%) were observed for animals in which transferred blastocysts had undergone normal cleavage, identified by the presence of 2 blastomeres of the same size without fragmentation, than among those with abnormal cleavage (33.3%). These results suggest that our microwell culture system is a powerful tool for producing and selecting healthy embryos and for identifying viability biomarkers.
Reproduction Fertility and Development 01/2011; 23(1):173. DOI:10.1071/RDv23n1Ab138 · 2.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We previously reported that superstimulatory (SS) treatment-induced follicular wave synchronization after ovum pickup (OPU) was effective in enhancing the quality of obtained oocytes and blastocysts derived from in vitro maturation (IVM) and fertilization (IVF; Imai et al. 2010 Reprod. Fertil. Dev. 22, 296). The present study was designed to examine the efficiency of embryo production by 4 sessions of OPU-IVF using a series of the SS treatment-induced follicular wave synchronizations. For the SS protocols, 3 consecutive SS (3CSS) and 2 separated SS (2SSS) were used. In the 3CSS group, the first OPU was performed on random days of the oestrous cycle (Day 0) and all follicles larger than 2mm in diameter were aspirated. On Day 5, follicles larger than 8mm in diameter were aspirated and a CIDR (InterAg, Hamilton, New Zealand) was inserted. The cows then received 20 armour units of FSH (Kawasaki-Seiyaku, Kawasaki, Japan) in twice-daily decreasing doses by IM injection from Day 7 to 10. Cloprostenol (PGF; 0.75mg, Fujita-Pharm, Tokyo, Japan) was administered on the morning of Day 9. The second OPU was performed 48h after PGF administration on Day 11; the CIDR was removed from the cows just before OPU. After the second OPU, donors were treated consecutively with the SS protocol mentioned above for the third and fourth OPU sessions. In the 2SSS group, donors received 2 sets of the SS treatment mentioned above, with an interval of 11 days between the second and the third OPU session. Four OPU sessions were performed every 11 days on all cows. In this study, 8 Japanese Black cows were divided into the 3CSS and 2SSS groups, and the treatment for each group was reversed after a 65-day interval as crossover trials. After OPU, Grade 1 and 2 oocytes were used for IVM and IVF, and putative zygotes were cultured as described by (Imai et al. 2006 J. Reprod. Dev. 52, S19-S29 suppl.). A part of the zygotes were cultured in a micro-well system. Data were analysed by Student's t-test and chi-square test. There were differences (P<0.05) in the mean (±SD) number of follicles, collected oocytes, and cultured oocytes in the 3CSS (35.0±8.6 and 24.4±11.2, respectively) and 2SSS (30.8±10.5 and 20.2±9.0, respectively) groups. There were no differences in mean percentage of blastocyst formation and Grade 1 blastocyst rates between the 3CSS (38.5 and 55.8%, respectively) and 2SSS (34.8 and 54.8%, respectively) groups. However, the mean number of blastocysts produced per OPU session was significantly (P<0.05) higher in the 3CSS group (8.1±6.3) compared with the 2SSS group (5.8±4.4). These results indicate that a series of 3 consecutive SS treatments had greater efficiency in producing OPU-IVF embryos.
Reproduction Fertility and Development 01/2011; 23(1):205. DOI:10.1071/RDv23n1Ab212 · 2.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We have developed a polystyrene-based well-of-the-well (WOW) system using injection molding to track individual embryos throughout culture using time-lapse cinematography (TLC). WOW culture of bovine embryos following in vitro fertilization was compared with conventional droplet culture (control). No differences between control- and WOW-cultured embryos were observed during development to the blastocyst stage. Morphological quality and inner cell mass (ICM) and trophectoderm (TE) cell numbers were not different between control- and WOW-derived blastocysts; however, apoptosis in both the ICM and TE cells was reduced in WOW culture (P < 0.01). Oxygen consumption in WOW-derived blastocysts was closer to physiological level than that of control-derived blastocysts. Moreover, WOW culture improved embryo viability, as indicated by increased pregnancy rates at Days 30 and 60 after embryo transfer (P < 0.05). TLC monitoring was performed to evaluate the cleavage pattern and the duration of the first cell cycle of embryos from oocytes collected by ovum pickup; correlations with success of pregnancy were determined. Logistic regression analysis indicated that the cleavage pattern correlated with success of pregnancy (P < 0.05), but cell cycle length did not. Higher pregnancy rates (66.7%) were observed for animals in which transferred blastocysts had undergone normal cleavage, identified by the presence of two blastomeres of the same size without fragmentation, than among those with abnormal cleavage (33.3%). These results suggest that our microwell culture system is a powerful tool for producing and selecting healthy embryos and for identifying viability biomarkers.
Biology of Reproduction 12/2010; 83(6):970-8. DOI:10.1095/biolreprod.110.085522 · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of the present study was to optimise the culture conditions for the in vitro production of bovine embryos. The development of in vitro fertilised bovine oocytes in CR1aa supplemented with 5% calf serum and IVD101 culture media were compared using traditional microdrops and Well of the Well (WOW) culture systems either under 5% or 20% oxygen tension. After 7 days of culture, a significantly higher blastocyst formation rate was obtained for embryos cultured in CR1aa medium compared to those cultured in IVD101, irrespective of O2 tensions and culture systems. The blastocyst formation in IVD101 was suppressed under 20% O2 compared to 5% O2 . Despite their similar total cell numbers, higher rates of inner cell mass (ICM) cells were observed in blastocysts developed in IVD101 medium than in those developed in CR1aa, irrespective of O2 tensions. There was no significant difference in blastocyst formation, total, ICM and trophectoderm (TE) cell numbers between embryos obtained by microdrop and WOW culture systems irrespective of the culture media and O2 tensions used. In conclusion, CR1aa resulted in higher blastocyst formation rates irrespective of O2 tension, whereas IVD101 supported blastocyst formation only under low O2 levels but enhanced the proliferation of ICM cells.
[Show abstract][Hide abstract] ABSTRACT: This study was conducted to study the kinetics of initial cell divisions in relation with the cleavage patterns in viable (with the ability to develop to the blastocyst stage) and non-viable bovine embryos and parthenotes. The kinetics of in vitro development and cleavage patterns were observed by time lapse cinematography. The length of the first and second but not third cell cycle differed significantly between the viable and non-viable embryos after IVF or parthenogenesis. Viable embryos had significantly shorter first and second cell cycles than non-viable ones. The presence of fragments, protrusions and unequally-sized blastomeres was associated with an extended one-cell stage and reduced ability to develop to the blastocyst stage; however, the lengths of the second and third cell cycles were not altered. Oocytes showing direct division from one cell to 3 or 4 blastomeres showed similar developmental ability and embryonic cell numbers to those showing normal division, although, with a high frequency of chromosomal abnormalities. Our results suggest that the differences in the first cell cycles between viable and non-viable embryos were not sperm-related, whereas direct cleavage of 1-cell embryos to 3 or more blastomeres and protrusion formation are related to sperm-driven factors. The length of the first and second cell cycles and the cleavage pattern should be examined simultaneously to predict developmental competence of embryos at early cleavage stages.
Journal of Reproduction and Development 04/2010; 56(2):200-7. DOI:10.1262/jrd.09-097A · 1.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The purpose of this study was to assess the feasibility of polyester mesh culture for the in vitro production of bovine embryos, as polyester mesh is an alternative way for tracking individual embryos throughout culture using time-lapse cinematography (TLC). Bovine embryos were isolated during in vitro culture using sections of three different polyethylene terephthalate (PET) mesh products. In vitro matured and fertilized bovine oocytes were cultured in the 217 × 217, 230 × 230 or 238 × 238-μm openings of PET mesh sections or in simple micro-drops (control) for 7 days under either 20% or 5% O(2) tensions. No difference in embryo developmental rates was found between the culture groups in terms of cleavage, blastocyst formation and blastocyst expansion irrespective of O(2) tension. In contrast, under 20% O(2) tension, blastocysts that developed in PET mesh with 217 × 217-μm opening had significantly higher numbers of total and trophectoderm (TE) cells than control embryos; however, the numbers and proportions of inner cell mass (ICM) cells did not differ. Under 5% O(2) tension, no difference was found among the culture groups in the numbers of total, ICM and TE cells in embryos. All three PET mesh products investigated in this study were proven to be effective to prevent embryo movement. The results demonstrate that bovine embryos can be cultured in PET mesh sections without negative side-effects and suggest that embryo distance determined by the mesh affects embryo quality at atmospheric oxygen tension. Polyethylene terephthalate mesh with 217 × 217-μm openings was found to be the most suitable for further application in TLC.