scholarly journals The Association of Kinetic Variables with Blastocyst Development and Ploidy Status

2021 ◽  
Author(s):  
Francesca Pennetta ◽  
Cristina Lagalla ◽  
Raffaella Sciajno ◽  
Nicoletta Tarozzi ◽  
Marco Nadalini ◽  
...  
Keyword(s):  
Blastocyst Stage ◽  
Human Embryos ◽  
Blastocyst Development ◽  
Developmental Potential ◽  
Logistic Regression Models ◽  
Kolmogorov Smirnov ◽  
Ploidy Changes ◽  
Study Hypothesis ◽  
Ploidy Status ◽  
Smirnov Test

Background: Despite a plethora of studies conducted so far, a debate is still unresolved as to whether TLM can identify predictive kinetic biomarkers or algorithms universally applicable. Therefore, this study aimed to elucidate if there is a relationship between kinetic variables and ploidy status of human embryos or blastocyst developmental potential. Methods: For conducting this retrospective cohort study, the normal distribution of data was verified using Kolmogorov-Smirnov test with the Lilliefors’ amendment and the Shapiro-Wilk test. Kinetic variables were expressed as median and quartiles (Q1, Q2, Q3, Q4). Mann-Whitney U-test was used to compare the median values of parameters. Univariate and multiple logistic regression models were used to assess relationship between blastocyst developmental potential or ploidy status and kinetics. Several confounding factors were also assessed. Results: Blastocyst developmental potential was positively correlated with the t4-t3 interval (s2) (OR=1.417, 95% CI of 1.288-1.560). s2 median value was significantly different between high- and low-quality blastocysts (0.50 and 1.33 hours post-insemination, hpi, respectively; p=0.003). In addition, timing of pronuclear appearance (tPNa) (OR=1.287; 95% CI of 1.131-1.463) had a significant relationship with ploidy changes. The median value of tPNa was statistically different (p=0.03) between euploid and aneuploid blastocysts (Euploid blastocysts=8.9 hpi; aneuploid blastocysts=10.3 hpi).  Conclusion: The present findings are in line with the study hypothesis that kinetic analysis may reveal associations between cleavage patterns and embryo development to the blastocyst stage and ploidy status.

scholarly journals 324SPHINGOSINE-1-PHOSPHATE PROTECTS CULTURED BOVINE OOCYTES FROM PHYSIOLOGICALLY RELEVANT THERMAL STRESS

2004 ◽  
Vol 16 (2) ◽  
pp. 282 ◽  
Author(s):  
Z. Roth ◽  
P.J. Hansen
Keyword(s):  
Thermal Stress ◽  
Heat Shock ◽  
Blastocyst Stage ◽  
Caspase Activity ◽  
Blastocyst Development ◽  
Cleavage Rate ◽  
Developmental Potential ◽  
Oocyte Competence ◽  
Cell Death Pathway

Sphingosine-1-phosphate (S1P) is a sphingolipid metabolite that can block the sphingomyelin cell-death pathway by suppressing ceramide-induced apoptosis. The present study was performed to test whether S1P protects oocytes from heat shock during in vitro maturation. Cumulus-oocyte complexes obtained by slicing follicles were placed in maturation medium with or without 50nM S1P and cultured at 38.5°C (CON) or 41°C (41C) for the first 12h of maturation. Incubation during the last 10h of maturation (22-h total maturation time), fertilization, and embryonic development were performed at 38.5°C and 5% (v/v) CO2. Blastocyst development was recorded at 8 days post-insemination (dpi) and activity of group II caspases in 8-day blastocysts was determined using a fluoroprobe, PhiPhiLux-G1D2 (OncoImmunin, Gaithersburg, MD, USA). Data were analysed by least-squares ANOVA with the GLM procedure of SAS. Percentage data were subjected to arcsin transformation before analysis. Exposure of oocytes to thermal stress during the first 12h of maturation reduced cleavage rate (P<0.01) and the number of oocytes developing to the blastocyst stage (P<0.04). There was a temperature x S1P interaction for cleavage rate (P<0.03) because S1P blocked effects of thermal stress on cleavage rate. Without S1P, the percentage of oocytes that cleaved by 3 dpi were 83.6±2.7% and 65.8±2.7% for CON and 41C, respectively. In the presence of S1P, percent cleavage was 86.7±2.7% and 83.9±2.7% for CON and 41C, respectively. There was a trend (P=0.06) for a temperature x S1P interaction for percent oocytes developing to blastocyst stage because S1P blocked effects of heat shock on development. Without S1P, the percentages of oocytes that developed to the blastocyst stage were 28.7±3.0% and 15.2±3.0% for CON and 41C, respectively. In the presence of S1P, percent blastocysts were 24.3±3.4% and 23.9±3.0% for CON and 41C, respectively. When development was expressed as percentage of cleaved embryos, however, there were no effects of temperature, S1P, or temperature x S1P on percent development to the blastocyst stage. Blastocyst caspase activity was not affected by temperature or S1P. In summary, exposure to physiologically relevant thermal stress during the first 12h of maturation has a deleterious effect on oocyte competence and this effect can be reduced by S1P. The fact that heat shock reduced the percentage of oocytes but not the percentage of cleaved embryos that became blastocysts suggests that oocytes that survive effects of heat shock and cleave have normal potential to develop to the blastocyst stage. Moreover, since heat shock did not affect caspase activity, it is likely that blastocysts from heat-shocked oocytes have normal developmental potential, at least as determined by caspase activity. Support: BARD FI-330-2002 and USDA Grants 2002-35203-12664 and 2001-52101-11318.

The effects of permeating cryoprotectants on intracellular free-calcium concentrations and developmental potential of in vitro-matured feline oocytes

2016 ◽  
Vol 28 (5) ◽  
pp. 599 ◽  
Author(s):  
Jason R. Herrick ◽  
Chunmin Wang ◽  
Zoltan Machaty
Keyword(s):  
Enzymatic Digestion ◽  
Blastocyst Stage ◽  
Intracellular Free Calcium ◽  
In Vitro Fertilisation ◽  
Free Calcium ◽  
Blastocyst Development ◽  
Oocyte Vitrification ◽  
Developmental Potential ◽  
Parthenogenetic Activation

Embryos produced from vitrified feline oocytes have resulted in pregnancies, but the efficiency of oocyte vitrification in cats is still low. Our objectives were to evaluate the effects of exposing feline oocytes to ethylene glycol (EG), propanediol (PrOH) and dimethyl sulfoxide (DMSO) on changes in intracellular free-calcium concentrations ([Ca2+]i), the time needed for enzymatic digestion of the zona pellucida (ZP), the incidence of parthenogenetic activation and degeneration and embryonic development following in vitro fertilisation (IVF). All of the chemicals tested altered [Ca2+]i, but changes in [Ca2+]i, resistance of the ZP to enzymatic digestion and the incidence of parthenogenetic activation (<5% for all treatments) were not affected (P > 0.05) by extracellular Ca2+. Exposure to EG (>44.1%) and DMSO (19.7%) increased (P < 0.05) oocyte degeneration compared with control oocytes and oocytes exposed to PrOH (≤2.5%). Following exposure to a combination of PrOH and DMSO (10% v/v each), blastocyst development (per cleaved embryo; 52.1%) was similar (P > 0.05) to control oocytes (64.4%). When oocytes were vitrified with PrOH and DMSO, 28.3% of surviving (intact plasma membrane) oocytes cleaved following IVF, but no blastocyst developed. When a non-permeating cryoprotectant (galactose, 0.25 M) was added to the vitrification medium, 47.7% of surviving oocytes cleaved and 14.3% developed to the blastocyst stage.

scholarly journals Use of Insulin to Increase Epiblast Cell Number: Towards a New Approach for Improving ESC Isolation from Human Embryos

2013 ◽  
Vol 2013 ◽  
pp. 1-7
Author(s):  
Jared M. Campbell ◽  
Michelle Lane ◽  
Ivan Vassiliev ◽  
Mark B. Nottle
Keyword(s):  
Embryonic Stem ◽  
Cell Number ◽  
Poor Quality ◽  
Blastocyst Stage ◽  
Human Embryos ◽  
Cleavage Stage ◽  
Embryo Viability ◽  
Blastocyst Development ◽  
Cell Stage ◽  
Hesc Derivation

Human embryos donated for embryonic stem cell (ESC) derivation have often been cryopreserved for 5–10 years. As a consequence, many of these embryos have been cultured in media now known to affect embryo viability and the number of ESC progenitor epiblast cells. Historically, these conditions supported only low levels of blastocyst development necessitating their transfer or cryopreservation at the 4–8-cell stage. As such, these embryos are donated at the cleavage stage and require further culture to the blastocyst stage before hESC derivation can be attempted. These are generally of poor quality, and, consequently, the efficiency of hESC derivation is low. Recent work using a mouse model has shown that the culture of embryos from the cleavage stage with insulin to day 6 increases the blastocyst epiblast cell number, which in turn increases the number of pluripotent cells in outgrowths following plating, and results in an increased capacity to give rise to ESCs. These findings suggest that culture with insulin may provide a strategy to improve the efficiency with which hESCs are derived from embryos donated at the cleavage stage.

104 EMBRYO MANIPULATION TECHNIQUES ALTER PRE-IMPLANTATION DEVELOPMENT AND GENE EXPRESSION IN MOUSE EMBRYOS

2016 ◽  
Vol 28 (2) ◽  
pp. 182 ◽  
Author(s):  
D. Jones ◽  
M. Paczkowski ◽  
T. Kuehl
Keyword(s):  
Gene Expression ◽  
Transcript Abundance ◽  
Blastocyst Stage ◽  
Mouse Embryos ◽  
Human Embryos ◽  
Assisted Hatching ◽  
Long Term Effects ◽  
Slow Cooling ◽  
Developmental Potential ◽  
Cell Counts

Micromanipulation techniques are commonly employed with human embryos; however, the long-term effects on growth and development of embryos are unknown. Subjective techniques, such as embryo grading, may not be sensitive enough to show differences in developmental potential and more objective assessments are needed. The objective of this experiment was to evaluate whether manipulation techniques and cryopreservation alter morphologic and gene expression endpoints of embryos. Two-cell mouse embryos were cultured to blastocyst stage and divided into 6 treatment groups: nonmanipulated control (UNMAN; n = 252), laser-assisted hatching (LAH; n = 124), LAH with slow cooling cryopreservation (LAHcryo; n = 78), simulated biopsy (BIOP; n = 90) performed by laser disruption of 1/8 of the embryo’s cellular volume, BIOP with slow cooling cryopreservation (BIOPcryo; n = 91), and BIOP with vitrification (BIOPvit; n = 66). After manipulation or thaw, embryos were cultured 28 h to blastocyst stage. Fully hatched and hatching blastocysts were collected for cell counts and quantitative PCR to assay absolute transcript abundance for Plac8, Glut1, Oct4, and Cox2, 18s rRNA, and Ppia using standard curves generated from serial dilutions of digested plasmids. Data were analysed using ANOVA with Duncan’s post hoc test and significance was defined as P < 0.05. Manipulation groups differed in developmental stage at 28 h postmanipulation (P = 0.03), with a larger percentage of embryos completely hatched in LAH (46%) and BIOP (45%) groups compared to UNMAN (11%). Cryopreservation reduced percentages of completely hatched embryos (LAHcryo = 26%; BIOPcryo = 31%; BIOPvit = 19%). The number of cells per embryo also varied (P < 0.001); UNMAN and LAH groups had similar numbers of blastomeres (UNMAN = 71; LAH = 74), whereas BIOP groups had reduced cell counts (BIOP = 53; BIOPcryo = 56; BIOPvit = 48). Patterns of gene expression varied between groups; the BIOP group had more cDNA per cell (UNMAN = 125 ng cell–1; BIOP = 202 ng cell–1; P = 0.004), whereas BIOPcryo group did not show this effect (BIOPcryo = 111 ng cell–1). Transcript abundance of Cox2, Oct4, and Glut1 per embryo was significantly decreased (P < 0.05) in biopsied embryos that were cryopreserved, either by slow-cooling or vitrification (Cox2: UNMAN = 643, BIOP = 490, BIOPcryo = 259, BIOPvit = 268; Oct4: UNMAN = 266, BIOP = 181, BIOPcryo = 95, BIOPvit = 97; Glut1: UNMAN = 643, BIOP = 490, BIOPcryo = 259, BIOPvit = 268). The data suggests that micromanipulation alters developmental timelines and gene expression of embryos. While manipulations differ in degree and nature of effect, our investigation implies all manipulations have some effect on the transcriptome of the developing embryo. Differences in cDNA quantity in the BIOP group may indicate a stress response or recovery attempt that becomes blunted in biopsied embryos which are subsequently cryopreserved. Thus, our study suggests that cryopreservation of biopsied embryos may come at a price and offers scientific support for only judiciously selected and medically indicated embryo biopsies.

scholarly journals Individual commitment to a group effect: strengths and weaknesses of bovine embryo group culture

Reproduction ◽  
2014 ◽  
Vol 148 (5) ◽  
pp. 519-529 ◽  
Author(s):  
Eline Wydooghe ◽  
Leen Vandaele ◽  
Sofie Piepers ◽  
Jeroen Dewulf ◽  
Etienne Van den Abbeel ◽  
...  
Keyword(s):  
Classical Group ◽  
Blastocyst Stage ◽  
Human Embryos ◽  
Bovine Embryo ◽  
Bovine Embryos ◽  
Blastocyst Development ◽  
Group Culture ◽  
Individual Culture ◽  
Better Than

Recently, new culture devices such as Corral and Primo Vision dishes have been designed for the culture of human embryos to allow the combination of group culture plus follow-up of individual embryos. Bovine inseminated oocytes were allocated to Primo Vision dishes, Corral dishes, individual culture or classical group culture. Blastocyst development in Primo Vision dishes was similar to classical group culture (34.3 and 39.0% respectively), and better than Corral dishes or individual culture (28.9 and 28.5% respectively). In Primo Vision dishes, a higher number of ‘slow’ embryos developed to the blastocyst stage compared with their individually cultured counterparts, while no differences were observed for ‘fast’ embryos. ‘Slow’ embryos in a ‘standard drop’ had a higher chance of becoming a blastocyst compared with individual culture (OR: 2.3), whereas blastulation of ‘fast’ embryos was less efficient in a ‘delayed drop’ than in individual culture (OR: 0.3). The number of non-cleaved embryos in Primo Vision dishes did not negatively influence blastocyst development. Likewise, removing non-cleaved embryos (NC removed) and regrouping the cleaved embryos afterwards (ReGR) did not affect blastocyst development and quality compared with group culture in Primo Vision dishes (CTRL, 31.6%, NC removed, 29.3% and ReGR, 29.6%). The experiments revealed that group culture of bovine embryos in Primo Vision dishes is superior to individual culture, primarily because of the higher blastocyst rate achieved by slow embryos. Non-cleaved or arrested embryos do not hamper the ability of co-cultured bovine embryos to reach the blastocyst stage in group culture.

scholarly journals Effect of the Re-Vitrification of Embryos at Different Stages on Embryonic Developmental Potential

2021 ◽  
Vol 12 ◽  
Author(s):  
Jingyu Li ◽  
Shun Xiong ◽  
Yanhua Zhao ◽  
Chong Li ◽  
Wei Han ◽  
...  
Keyword(s):  
Embryo Transfer ◽  
Blastocyst Stage ◽  
Expression Level ◽  
Delivery Rate ◽  
Human Embryos ◽  
Control Group ◽  
Cell Stage ◽  
Developmental Potential ◽  
Group 2 ◽  
Group 1

BackgroundUsing re-vitrified human embryos for frozen-warmed embryo transfer (FET) is a valuable option when there are no other cryopreserved embryos to use, however, except for the PGT cases, no published data are available for FET with human embryos that were re-vitrified at different developmental stages.ObjectiveTo evaluate the effect of re-vitrification of embryos at different stages on embryonic developmental potential.MethodThis study included clinical retrospective and mouse experimental studies. For the retrospective study, a total of 25 FET cycles with re-vitrified day 3 embryos (re-vitrification group 1) and 54 FET cycles with re-vitrified day 5 blastocysts (re-vitrification group 2) between January 2015 and December 2019 were included in this study. The corresponding FET cycles with once-vitrified embryos were identified using propensity score (PS) matching according to the time of embryo transfer. For the mouse experimental study, we divided embryos into 5 groups: fresh (group 1), vitrified at the 8-cell stage (group 2), vitrified at the early blastocyst stage (group 3), vitrified at the 8-cell stage, and re-vitrified at the 8-cell (group 4) or early blastocyst stage (group 5). The fresh embryos was selected as control group. The primary outcome in this study was delivery outcomes.ResultsNo significant difference in delivery rate was detected between re-vitrification group 1 (24.00%) and the corresponding control group (28.00%). However, re-vitrification group 2 (46.3%) showed a significant decrease in delivery rate compared with the two corresponding control groups (63.89% and 64.12%) (P &lt; 0.05). Our experiment using mouse embryos also confirmed the clinical data, and showed that re-vitrification at the blastocyst stage following the first round of vitrification at the 8-cell stage reduced the delivery rate. In addition, both re-vitrified groups showed a significantly higher expression level of BAX. However, only re-vitrification at the blastocyst stage increased the expression level of CASPASE3.ConclusionsRe-vitrification at the 8-cell and blastocyst stages has different effects on embryonic developmental potential, as re-vitrification at blastocyst stage following a previous vitrification at 8-cell stage reduced the delivery rate, while vitrification at the 8-cell stage twice achieved comparable pregnancy outcomes to the once-vitrified group.

scholarly journals 275FOLLICULAR ATRESIA IN SMALL, NON-FSH-DEPENDENT BOVINE FOLLICLES IS ASSOCIATED WITH INCREASED DEVELOPMENTAL POTENTIAL OF OOCYTES FOLLOWING IVP

2004 ◽  
Vol 16 (2) ◽  
pp. 258
Author(s):  
H. Irving-Rodgers ◽  
S. Morris ◽  
R. Collett ◽  
K. Catanzariti ◽  
T. Peura ◽  
...  
Keyword(s):  
Basal Lamina ◽  
Calf Serum ◽  
Blastocyst Stage ◽  
Developmental Competence ◽  
Early Cleavage ◽  
Blastocyst Development ◽  
Chi Square ◽  
Developmental Potential ◽  
Chi Square Analysis

Oocytes from small, non-FSH-dependent follicles are associated with reduced developmental competence following in vitro embryo production (IVP) compared to oocytes from larger follicles. It has been suggested that, for small follicles, oocytes derived from atretic follicles are more developmentally competent than those from healthy follicles (Blondin P and Sirard MA, 1995 Mol. Reprod. Dev. 41, 54–62). Little is known of the characteristics of small follicles that support developmentally competent oocytes. Here we examine the development to blastocyst stage of oocytes collected from histologically-assessed bovine 2–5mm follicles. Ovaries were obtained at a local abattoir;; 4 follicles were dissected from each ovary and oocytes were recovered. A section of each follicle wall was taken and fixed in 2.5% glutaraldehyde for histological assessment of the follicle and characterization of the morphology of the follicular basal lamina by electron microscopy (Irving-Rodgers HF and Rodgers RJ, 2000 J. Reprod. Fert. 118, 221–228). Oocytes recovered from follicles underwent IVP utilizing a novel single IVP system. Oocytes were matured for 24h (10μL per COC) in TCM199, supplemented with FSH, hCG, FCS, cysteamine and pyruvate. Mature oocytes were inseminated with 1×106 sperm mL−1 for an additional 24h using Bovine Fertilization Medium (10μL per COC;; Cook, Australia). Following insemination, putative zygotes were stripped of remaining cells and placed within individual micro-wells prepared in 1% agar in Bovine Early Cleavage Medium, Cook, Australia. The agar (350μL) was prepared within wells of a 4-well plate and small plugs of agar were removed to form micro-wells. The agar was over-laid with 450μL of Early Cleavage Medium and 250μL mineral oil, and equilibrated overnight before putative zygotes were placed individually within micro-wells. Culture was performed under 7% O2, 6% CO2, and 87% N2 at 39°C. On Day 5 following insemination, fetal calf serum (final concentration 10% v/v) was added to facilitate blastocyst development. Blastocyst formation was assessed on Day 8. A total of 211 oocytes were cultured and 69% were from healthy follicles;; 67 oocytes (32%) had developed to the blastocyst stage by Day 8. Forty-three percent of oocytes recovered from atretic follicles (28/65) had developed to the blastocyst stage by Day 8, as compared to only 27% (39/146) oocytes recovered from healthy follicles, this difference was significant (P&lt;0.05, chi-square analysis). Seventy-eight percent (14/18) of oocytes from healthy follicles with additional follicular basal lamina material (Irging-Rodgers HF and Rodgers RJ, 2000 J. Reprod. Fert. 118, 221–228) failed to develop, whereas only 44% (4/9) of oocytes from healthy follicles with a normal basal lamina failed to develop (P&gt;0.08). The present study finds a direct association between the follicle morphology and oocyte maturational potential within non-FSH dependent follicles, revealing that high levels of development (&gt;40%) can be obtained from atretic follicles. Furthermore, differences between healthy follicles may also contribute to developmental variation.

158 Predicting embryo development success with physical parameters of oocytes

2019 ◽  
Vol 31 (1) ◽  
pp. 204
Author(s):  
C. L. Timlin ◽  
A. Lynn ◽  
R. R. White ◽  
K. Lee ◽  
V. R. G. Mercadante
Keyword(s):  
Polar Body ◽  
Digital Camera ◽  
Blastocyst Stage ◽  
Physical Parameters ◽  
Oocyte Diameter ◽  
Blastocyst Development ◽  
Noninvasive Methods ◽  
Developmental Potential ◽  
Maturation Medium ◽  
Blastocyst Rate

There is a continual search for reliable, noninvasive methods of selecting viable oocytes and embryos. Previous studies have indicated that the physical size of oocytes may reflect their developmental potential. The objective of this study was to observe the correlation between an oocyte’s diameter [including zona pellucida (ZP), cell area, and ZP thickness] and its ability to develop. Bovine cumulus-oocyte complexes were collected from abattoir-derived ovaries and incubated for 24h in TCM-199-based maturation medium. The cumulus-oocyte complexes were denuded by vortexing with hyaluronidase, and mature oocytes were selected based on presence of a visible polar body. Selected oocytes were artificially activated by incubating in 5 μM ionomycin for 5min followed by incubation in 2mM 6-DMAP for 3h (n=723). After activation, oocytes were placed in individual 5-μL culture droplets under oil and photographed using an inverted scope with digital camera. Oocytes were then group cultured in 50-μL droplets in a polyester micromesh for identification of individual oocytes. Development to the blastocyst stage was noted on Days 7 and 8 of culture. ImageJ was used to measure diameter, area, and ZP thickness from images. A logistic regression using the lme4 package in R was run with Day 7 or 8 blastocyst development as the response; diameter, area, and ZP thickness as predictors; and replicate and culture droplet as random effects. The residual estimates of area and ZP thickness from diameter were used to account for correlation between predictors. Only significant interactions were kept in the model. The ZP thickness ranged from 6.1 to 17.7μm with a mean of 12μm. There was a significant correlation between diameter and ZP thickness (P&lt;0.01, R2=0.12) with a correlation coefficient of 0.35. Oocyte diameter had a significant effect on subsequent blastocyst development on Day 7 (P&lt;0.01) and Day 8 (P&lt;0.01), with larger oocytes more likely to develop on both days. Oocytes were also grouped into quantiles by diameter. Larger groups were more likely to develop to the blastocyst stage on Day 7 (P&lt;0.001) and Day 8 (P&lt;0.001). Blastocyst rate on Day 8 for oocytes with diameters &lt;149.5μm was 24.2%, whereas blastocyst rate on Day 8 for those with diameters=159μm was 41.2%. In addition, ZP thickness also had an effect: oocytes with thinner ZP were more likely to develop to the blastocyst stage on Day 7 (P&lt;0.001) and Day 8 (P&lt;0.001). Blastocyst rate for oocytes with a ZP thickness &lt;11μm was 37%, whereas the rate for those with a ZP thickness of 12.9μm was 27.6%. Area did not have an effect on blastocyst formation on Day 7 or 8 (P=0.21). Here we have demonstrated that the oocyte diameter, including the ZP and ZP thickness, affects its probability of development. Larger oocytes and those with thinner ZP are more likely to develop to the blastocyst stage. Differences in the size of the perivitelline space could explain why diameter had a significant effect on development and area did not. Further studies will focus on determining the relationship between these physical parameters of oocytes and embryo quality.

Applying metabolomic analyses to the practice of embryology: physiology, development and assisted reproductive technology

2015 ◽  
Vol 27 (4) ◽  
pp. 602 ◽  
Author(s):  
Rebecca L. Krisher ◽  
Adam L. Heuberger ◽  
Melissa Paczkowski ◽  
John Stevens ◽  
Courtney Pospisil ◽  
...  
Keyword(s):  
Amino Acids ◽  
Culture Medium ◽  
Blastocyst Stage ◽  
Small Samples ◽  
Human Embryos ◽  
Blastocyst Development ◽  
Oct4 Expression ◽  
Complex Culture

The advent of metabolomics technology and its application to small samples has allowed us to non-invasively monitor the metabolic activity of embryos in a complex culture environment. The aim of this study was to apply metabolomics technology to the analysis of individual embryos from several species during in vitro development to gain an insight into the metabolomics pathways used by embryos and their relationship with embryo quality. Alanine is produced by both in vivo- and in vitro-derived human, murine, bovine and porcine embryos. Glutamine is also produced by the embryos of these four species, but only those produced in vitro. Across species, blastocysts significantly consumed amino acids from the culture medium, whereas glucose was not significantly taken up. There are significant differences in the metabolic profile of in vivo- compared with in vitro-produced embryos at the blastocyst stage. For example, in vitro-produced murine embryos consume arginine, asparagine, glutamate and proline, whereas in vivo-produced embryos do not. Human embryos produce more alanine, glutamate and glutamine, and consume less pyruvate, at the blastocyst compared with cleavage stages. Glucose was consumed by human blastocysts, but not at a high enough level to reach significance. Consumption of tyrosine by cleavage stage human embryos is indicative of blastocyst development, although tyrosine consumption is not predictive of blastocyst quality. Similarly, although in vivo-produced murine blastocysts consumed less aspartate, lactate, taurine and tyrosine than those produced in vitro, consumption of these four amino acids by in vitro-derived embryos with high octamer-binding transcription factor 4 (Oct4) expression, indicative of high quality, did not differ from those with low Oct4 expression. Further application of metabolomic technologies to studies of the consumption and/or production of metabolites from individual embryos in a complete culture medium could transform our understanding of embryo physiology and improve our ability to produce developmentally competent embryos in vitro.

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