scholarly journals Evaluation of developmental competence of Sus scrofa domesticus (L.) oocyte-cumulus complexes after intra- and extraovarian vitrification

2021 ◽  
Vol 25 (6) ◽  
pp. 613-619
Author(s):  
T. I. Kuzmina ◽  
I. V. Chistyakova
Keyword(s):  
Developmental Competence ◽  
Chromatin State ◽  
Mitochondrial Activity ◽  
Highly Dispersed Silica ◽  
Maturation In Vitro ◽  
Sus Scrofa Domesticus ◽  
First Time ◽  
Cytogenetic Method ◽  
Mitotracker Orange

The aim of the present study was to identify the influence of extra- (EOV) and intraovarian vitrification (IOV) on mitochondrial activity (MA) and chromatin state in porcine oocytes during maturation in vitro. During EOV porcine oocytes were exposed in cryoprotective solutions (CPS): CPS-1 – 0.7 M dimethyl sulfoxide (DMSO)+0.9 M ethylene glycol (EG); CPS-2 – 1.4 M DMSO+1.8 M EG; CPS-3 – 2.8 M DMSO+3.6 M EG+0.65 M trehalose. At IOV the ovarian fragments were exposed in CPS-1 – 7.5 % EG+7.5 % DMSO, then in CPS-2 – 15 % EG, 15 % DMSO and 0.5 M sucrose. Straws with oocytes and ovarian fragments were plunged into LN2 and stored. For devitrification, the EOV oocytes were washed in solutions of 0.25, 0.19 and 0.125 M of trehalose, the IOV – in 0.5 and 0.25 М trehalose. Oocytes were cultured in NCSU-23 medium with 10 % fluid of follicles, follicular walls, hormones. 0.001 % of highly dispersed silica nanoparticles (ICP named after A.A. Chuyko of the NAS of Ukraine) were added to all media. The methods of fertilization and embryo culture are presented in the guidelines developed by us. MA and chromatin state were measured by MitoTracker Orange CMTMRos and the cytogenetic method. Significant differences in the level of oocytes with high-expanded cumulus between control and experimental vitrified groups (81 % versus 59 % and 52 %, respectively, p ≤ 0.001) were observed. The percentage of pyknotic cells in native oocytes was 19 %, EOV or IOV oocytes were 39 % and 49 %, respectively. After culture, the level of matured native oocytes was 86 %, 48 % EOV and 33 % IOV cells finished the maturation (p ≤ 0.001). Differences were also observed in the level of MA between groups treated by EOV and IOV (89.4±7.5 µA and 149.2±11.3 µA, respectively, p ≤ 0.05). For the first time, pre-implantation embryos were obtained from oocytes treated by IOV.

scholarly journals Glycine ameliorates mitochondrial dysfunction caused by ABT-199 in porcine oocytes

2021 ◽  
Author(s):  
Sicong Yu ◽  
Lepeng Gao ◽  
Yang Song ◽  
Xin Ma ◽  
Shuang Liang ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Mitochondrial Dysfunction ◽  
Oocyte Maturation ◽  
Mitochondrial Function ◽  
Protective Action ◽  
Damage Model ◽  
Developmental Competence ◽  
Free Calcium ◽  
Maturation In Vitro

Abstract Mitochondria play an important role in controlling oocyte developmental competence. Our previous studies showed that glycine can regulate mitochondrial function and improve oocyte maturation in vitro. However, the mechanisms by which glycine affects mitochondrial function during oocyte maturation in vitro have not been fully investigated. In this study, we induced a mitochondrial damage model in oocytes with the Bcl-2-specific antagonist ABT-199. We investigated whether glycine could reverse the mitochondrial dysfunction induced by ABT-199 exposure and whether it is related to calcium regulation. Our results showed that ABT-199 inhibited cumulus expansion, decreased the oocyte maturation rate and the intracellular glutathione (GSH) level, caused mitochondrial dysfunction, induced oxidative stress, which was confirmed by decreased mitochondrial membrane potential (Δ⍦m) and the expression of mitochondrial function-related genes (PGC-1α), and increased reactive oxygen species (ROS) levels and the expression of apoptosis-associated genes (Bax, caspase-3, CytC). More importantly, ABT-199-treated oocytes showed an increase in the intracellular free calcium concentration ([Ca 2+]i) and had impaired cortical type 1 inositol 1,4,5-trisphosphate receptors (IP3R1) distribution. Nevertheless, treatment with glycine significantly ameliorated mitochondrial dysfunction, oxidative stress and apoptosis, glycine also regulated [Ca 2+]i levels and IP3R1 cellular distribution, which further protects oocyte maturation in ABT-199-induced porcine oocytes. Taken together, our results indicate that glycine has a protective action against ABT-199-induced mitochondrial dysfunction in porcine oocytes.

scholarly journals Maturation of human oocytes in vitro and their developmental competence

Reproduction ◽  
2001 ◽  
pp. 51-75 ◽  
Author(s):  
A Trounson ◽  
C Anderiesz ◽  
G Jones
Keyword(s):  
Embryo Development ◽  
Oocyte Maturation ◽  
Growth Phase ◽  
Developmental Competence ◽  
Minimum Size ◽  
Success Rates ◽  
Human Oocytes ◽  
Maturation In Vitro

Complete maturation of oocytes is essential for the developmental competence of embryos. Any interventions in the growth phase of the oocyte and the follicle in the ovary will affect oocyte maturation, fertilization and subsequent embryo development. Oocyte size is associated with maturation and embryo development in most species examined and this may indicate that a certain size is necessary to initiate the molecular cascade of normal nuclear and cytoplasmic maturation. The minimum size of follicle required for developmental competence in humans is 5-7 mm in diameter. Maturation in vitro can be accomplished in humans, but is associated with a loss of developmental competence unless the oocyte is near completion of its preovulatory growth phase. This loss of developmental competence is associated with the absence of specific proteins in oocytes cultured to metaphase II in vitro. The composition of culture medium used successfully for maturation of human oocytes is surprisingly similar to that originally developed for maturation of oocytes in follicle culture in vitro. The presence of follicle support cells in culture is necessary for the gonadotrophin-mediated response required to mature oocytes in vitro. Gonadotrophin concentration and the sequence of FSH and FSH-LH exposure may be important for human oocytes, particularly those not exposed to the gonadotrophin surge in vivo. More research is needed to describe the molecular and cellular events, the presence of checkpoints and the role of gene expression, translation and protein uptake on completing oocyte maturation in vitro and in vivo. In the meantime, there are very clear applications for maturing oocytes in human reproductive medicine and the success rates achieved in some of these special applications are clinically valuable.

253 OXIDATIVE STRESS MAY IMPAIR OOCYTE QUALITY IN DAIRY COWS OF REPRODUCTIVE AGE WITH A REDUCED ANTRAL FOLLICLE COUNT

2013 ◽  
Vol 25 (1) ◽  
pp. 274 ◽  
Author(s):  
I. Tessaro ◽  
F. Franciosi ◽  
V. Lodde ◽  
D. Corbani ◽  
A. M. Luciano ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Nitric Oxide ◽  
Antral Follicle ◽  
Oocyte Quality ◽  
Antral Follicle Count ◽  
Oxide System ◽  
Reproductive Age ◽  
Developmental Competence ◽  
Mitochondrial Activity

In dairy cattle, oocytes isolated from ovaries with a reduced antral follicle count (AFC) have a low embryonic developmental competence. This may be related to oxidative stress, as indicated by our recent finding that ovaries with reduced AFC show a defective endothelial nitric oxide synthase/nitric oxide system. To further test this hypothesis, we evaluated whether the poor developmental competence of these oocytes was possibly due 1) to an imbalance of the reduced glutathione (GSH) system, because GSH is the major antioxidant compound stored within the oocyte and protects the zygote and early embryos from oxidative damage, and 2) to reduced mitochondrial activity. Ovaries were obtained from the abattoir, and oocytes were collected from ovaries with reduced AFC, with fewer than 10 follicles of 2 to 6 mm in diameter, and aged-matched controls, with more than 10 follicles of 2 to 6 mm in diameter. Oocyte GSH content was evaluated using the 5,5′-dithio-bis(2-nitrobenzoic acid)-GSH reductase recycling micro-GSH assay before and after in vitro maturation (IVM) in the presence or absence of 100 µM cysteamine, a GSH precursor. At the same time the developmental competence after IVF was assessed. Moreover, the mitochondrial activity during IVM was evaluated in additional oocytes from the two ovarian categories by specific MitoTracker dyes (MitoTracker FM Green and MitoTracker Orange CMTMRos, Invitrogen, Carlsbad, CA, USA) and subsequent image analysis (ImageJ software). All data were analysed by ANOVA followed by Fisher’s least significant differences test, and P-values <0.05 were considered significant. Experiments were repeated at least three times. Oocytes isolated from ovaries with a low AFC had a similar GSH content compared with oocytes isolated from control ovaries (n = 65 and 85, respectively; 4.31 ± 0.41 v. 4.51 ± 0.42 pmol oocyte–1). After IVM, oocytes from ovaries with reduced AFC showed a significantly lower GSH content compared with control oocytes (n = 55 and 65, respectively; 4.36 ± 0.31 v. 6.59 ± 0.39 pmol oocyte–1); however, cysteamine supplementation during IVM induced GSH accumulation similar to the control (n = 80 and 85, respectively; 9.88 ± 0.77 v. 10.45 ± 0.88 pmol oocyte–1). It is interesting that the increase in intracellular GSH content significantly improved the developmental competence of oocytes from ovaries with a reduced AFC (n = 196 and 201, respectively; 20.1 ± 2.9% v. 6.2 ± 1.6%), although the blastocyst rate remained lower than the control either with or without cysteamine (n = 218 and 212, respectively; 33.3 ± 3.8% and 34.2 ± 2.4%). Further, immature oocytes from ovaries with a low AFC showed a reduced mitochondrial membrane potential compared with control oocytes (n = 13 and 18, respectively; 1.74 ± 1.19 v. 2.22 ± 1.72, calculated as the ratio between the fluorescence of active and total mitochondria), whereas at the end of IVM, it declined in both categories at a comparable level (n = 17 and 24, respectively; 1.19 ± 0.10 and 1.30 ± 0.06). Our data confirmed the hypothesis that both the GSH imbalance and defective mitochondrial activity contribute to the limited developmental competence of oocytes from ovaries with a reduced AFC. This work was supported by Dote ricerca applicata-FSE, Regione Lombardia, Italy (VL, IT).

Meiotic progression, mitochondrial features and fertilisation characteristics of porcine oocytes with different G6PDH activities

2010 ◽  
Vol 22 (5) ◽  
pp. 830 ◽  
Author(s):  
István Egerszegi ◽  
Hannelore Alm ◽  
József Rátky ◽  
Bassiouni Heleil ◽  
Klaus-Peter Brüssow ◽  
...  
Keyword(s):  
Developmental Competence ◽  
Mitochondrial Activity ◽  
G6pdh Activity ◽  
Cresyl Blue ◽  
Meiotic Progression ◽  
Germinal Vesicles ◽  
Nuclear Development ◽  
Chromatin Configuration ◽  
Glucose 6 Phosphate Dehydrogenase

The aim of the present study was to investigate the developmental competence, mitochondrial characteristics and chromatin status of immature follicular porcine oocytes selected for their glucose-6-phosphate dehydrogenase (G6PDH) activity by brilliant cresyl blue (BCB) staining. In Experiment 1, the oocyte parameters were determined in parallel right after BCB staining (T0), after 22 h of in vitro maturation (IVM) (T22) and after 44 h of IVM (T44) (n = 496). BCB-stained oocytes (BCB+) at T0 were characterised by fibrillated chromatin filaments in their germinal vesicles (GV) and diakinesis stages whereas unstained (BCB–) oocytes at T0 contained in their GV mainly condensed stages of chromatin (P < 0.05). After 22 h of IVM BCB+ oocytes showed a prominent chromatin configuration of metaphase I and after 44 h the majority developed a M II nuclear configuration in contrast to the BCB– group (P < 0.0001). Differences were also observed between the two oocyte populations in their mitochondrial activity (P < 0.05). At the beginning of IVM BCB+ oocytes were characterised by high mitochondrial activity in their cytoplasm. The BCB+ oocytes showed clear visible homogenous distributions of mitochondria (P < 0.005) and contained more aggregated clusters of mitochondria in contrast to BCB– oocytes (P < 0.005). In Experiment 2, 318 oocytes were tested for their G6PDH activity and introduced to IVM and IVF. Only oocytes from the BCB+ group, which were matured after 44 h up to the stage of M II (81.6%) were fertilised (17.4%), penetrated (46%) or activated (15.6%) after IVF. These results indicate a relationship between the G6PDH activity of porcine oocytes before IVM and their subsequent nuclear development, mitochondrial activity and aggregation.

scholarly journals Transient meiotic arrest maintained by DON (6-diazo-5-oxo-l-norleucine) enhances nuclear/cytoplasmic maturation of porcine oocytes

Reproduction ◽  
2019 ◽  
Vol 158 (6) ◽  
pp. 543-554
Author(s):  
Hae-Jun Yang ◽  
Sanghoon Lee ◽  
Bo-Woong Sim ◽  
Pil-Soo Jeong ◽  
Seon-A Choi ◽  
...  
Keyword(s):  
Formation Rate ◽  
Developmental Competence ◽  
Cortical Granules ◽  
Meiotic Arrest ◽  
Cellular Survival ◽  
Cytoplasmic Maturation ◽  
First Time ◽  
A Current

The developmental competence of in vitro-matured oocytes is still lower than that of the in vivo-matured oocytes due to precocious meiotic resumption and inappropriate cytoplasmic maturation. Although numerous efforts have been attempted to accomplish better in vitro maturation (IVM) condition, only limited progress has been achieved. Thus, a current study was conducted to examine the effects of 6-diazo-5-oxo-l-norleucine (DON, an inhibitor of hyaluronan synthesis) during the first half period of IVM on nuclear/cytoplasmic maturation of porcine oocytes and subsequent embryonic development. Based on the observation of the nucleus pattern, metaphase II (MII) oocyte production rate in 1 µM DON group was significantly higher than other groups at 44 h of IVM. The 1 µM of DON was suggested to be optimal for porcine IVM and was therefore used for further investigation. Meiotic arrest effect of DON was maximal at 6 h of IVM, which was supported by the maintenance of significantly higher intra-oocyte cAMP level. In addition, increased pERK1/2 levels and clear rearrangement of cortical granules in membrane of MII oocytes matured with DON provided the evidence for balanced meiosis progression between nuclear and cytoplasmic maturation. Subsequently, DON significantly improved blastocyst formation rate, total cell numbers, and cellular survival in blastocysts after parthenogenetic activation, in vitro fertilization, and somatic cell nuclear transfer. Altogether, our results showed for the first time that 1 µM DON can be used to increase the yield of developmentally competent MII oocytes by synchronizing nuclear/cytoplasmic maturation, and it subsequently improves embryo developmental competence.

scholarly journals Antioxidant Nobiletin Enhances Oocyte Maturation and Subsequent Embryo Development and Quality

2020 ◽  
Vol 21 (15) ◽  
pp. 5340
Author(s):  
Yulia N. Cajas ◽  
Karina Cañón-Beltrán ◽  
Magdalena Ladrón de Guevara ◽  
María G. Millán de la Blanca ◽  
Priscila Ramos-Ibeas ◽  
...  
Keyword(s):  
Embryo Development ◽  
Biological Effects ◽  
Calf Serum ◽  
Cumulus Cells ◽  
Developmental Competence ◽  
Mitochondrial Activity ◽  
Cortical Granules ◽  
Cleavage Rate ◽  
Cytoplasmic Maturation

Nobiletin is a polymethoxylated flavonoid isolated from citrus fruits with wide biological effects, including inhibition of reactive oxygen species (ROS) production and cell cycle regulation, important factors for oocyte in vitro maturation (IVM). Therefore, the objective of the present study was to evaluate the antioxidant activity of nobiletin during IVM on matured bovine oocyte quality (nuclear and cytoplasmic maturation; oocyte mitochondrial activity; intracellular ROS and glutathione (GSH) levels) and their developmental competence, steroidogenesis of granulosa cells after maturation, as well as quantitative changes of gene expression in matured oocytes, their cumulus cells, and resulting blastocysts. Bovine cumulus-oocyte complexes were in vitro matured in TCM-199 +10% fetal calf serum (FCS) and 10 ng/mL epidermal growth factor (EGF) (Control) supplemented with 10, 25, 50, or 100 μM of nobiletin (Nob10, Nob25, Nob50, and Nob100, respectively) or 0.1% dimethyl sulfoxide (CDMSO: vehicle for nobiletin dilution). A significantly higher percentage of matured oocytes in metaphase II was observed in Nob25 and Nob50 compared to other groups. Similarly, cleavage rate and cumulative blastocyst yield on Days 7 and 8 were significantly higher for Nob25 and Nob50 groups. Oocytes matured with 25 and 50 μM nobiletin showed a higher rate of migration of cortical granules and mitochondrial activity and a reduction in the ROS and GSH content in comparison with all other groups. This was linked to a modulation in the expression of genes related to metabolism (CYP51A1), communication (GJA1), apoptosis (BCL2), maturation (BMP15 and MAPK1), and oxidative stress (SOD2 and CLIC1). In conclusion, nobiletin offers a novel alternative for counteracting the effects of the increase in the production of ROS during IVM, improves oocyte nuclear and cytoplasmic maturation, and subsequent embryo development and quality in cattle.

75 Improvement of Developmental Competence of Bovine In Vitro-Produced Embryos by Using Charcoal:Dextran-Stripped Fetal Bovine Serum on In Vitro Culture Media

2018 ◽  
Vol 30 (1) ◽  
pp. 176
Author(s):  
A. Mesalam ◽  
R. Kong ◽  
B.-H. Choi ◽  
K.-L. Lee ◽  
B.-Y. Park ◽  
...  
Keyword(s):  
In Vitro Culture ◽  
Bovine Serum ◽  
Culture Media ◽  
Fetal Bovine Serum ◽  
Developmental Competence ◽  
Mitochondrial Activity ◽  
Mrna Levels ◽  
Fetal Bovine

Serum has widely been used as a main supplement to embryo in vitro culture media as it contains embryotrophic factors. Charcoal:dextran treatment of fetal bovine serum (FBS) removes lipophilic chemicals and certain steroid hormones and growth factors. The objective of this study was to investigate the effects of charcoal:dextran-stripped fetal bovine serum (CDS FBS) and heat-inactivated FBS (HI FBS) in embryo culture medium (SOF-BE1 medium supplemented with 10% of serum) on their ability to support in vitro development of bovine embryos. The developmental ability and quality of bovine embryos were determined by assessing their cell number, lipid content, mitochondrial activity, gene expression, and cryo-tolerance. The experiment was conducted in 6 replicates (350 oocytes per group). The differences in embryo development, integrated optical intensity, and expression levels of the various genes between experimental groups were analysed by one-way ANOVA. Duncan’s multiple range tests were used to test the differences between the treatments. The level of statistical significance was set at P < 0.05. The percentages of embryos that underwent cleavage and formed a blastocyst were significantly (P < 0.05) higher in medium containing CDS FBS than in medium containing HI FBS (42.84 ± 0.78% v. 36.85 ± 0.89%, respectively). The total number of cells per Day 8 blastocyst was not significantly different (P > 0.05) between the CDS FBS group (208.40 ± 14.77) and the HI FBS group (195.11 ± 19.15). Furthermore, the beneficial effects of CDS FBS on embryos were associated with a significantly increased mitochondrial activity, as identified by MitoTracker Green, and reduced intracellular lipid content, as identified by Nile red staining, which increased their cryo-tolerance. The post-thaw survival rate of blastocysts was significantly (P < 0.05) higher after 24 h in the CDS FBS than in the HI FBS group (85.33 ± 4.84% v. 68.67 ± 1.20%). Quantitative reverse transcription PCR showed that the mRNA levels of lipid metabolism-related genes, acyl-CoA synthetase long-chain family member 3, acyl-coenzyme A dehydrogenase long-chain, and the cholesterol metabolism related gene hydroxymethylglutaryl-CoA reductase were significantly increased upon culture with CDS FBS. Moreover, the mRNA levels of survival gene sirtuin 1, antioxidant gene superoxide dismutase 2, and anti-apoptotic associated gene B-cell lymphoma 2 in frozen–thawed blastocysts were significantly (P < 0.05) higher in the CDS FBS group than in the HI FBS group; however, the mRNA level of the pro-apoptotic gene BCL2-associated X protein was significantly reduced. In conclusion, these data suggest that supplementation of in vitro culture medium with CDS FBS improves in vitro bovine embryo developmental competence and the quality of blastocysts in terms of their crytolerance and gene expression. This research was supported by grant from the Next-Generation BiogGeen21 (No. PJ01107703), IPET (No. 315017-5 and 117029-3), Allergy free cat (Co.. Felix Pets), BK21plus, and KGSP.

Acquisition of developmental competence during maturation in vitro

1986 ◽  
Vol 25 (1) ◽  
pp. 150 ◽  
Author(s):  
E.S. Critser ◽  
M.L. Leibfried-Rutledge ◽  
W.H. Eyestone ◽  
D.L. Northey ◽  
N.L. First
Keyword(s):  
Developmental Competence ◽  
Maturation In Vitro

scholarly journals Seasonal Effect on Developmental Competence, Oxidative Status and Tubulin Assessment of Prepubertal Ovine Oocyte

Animals ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1886
Author(s):  
Elisa Serra ◽  
Sergio Domenico Gadau ◽  
Giovanni Giuseppe Leoni ◽  
Salvatore Naitana ◽  
Sara Succu
Keyword(s):  
In Vitro Maturation ◽  
Reproductive Activity ◽  
Oocyte Quality ◽  
Developmental Competence ◽  
Seasonal Effect ◽  
Mitochondrial Activity ◽  
Reproductive Seasonality ◽  
Embryo Production

The reproductive seasonality of domestic animals is often manipulated in order to have more reproductive periods for commercial purposes related to the production of milk and meat. It is scientifically proven that such an alteration of the reproductive activity in sheep entails a deterioration in oocyte quality, leading to an inability to generate embryos. Since oocytes obtained from prepubertal ewes can be incorporated into an in vitro embryo production system and considering that their quality is crucial to the success of in vitro procedures, the aim of this work was to investigate the effect of seasons on the quality of prepubertal ovine oocytes collected in autumn and spring. Ovaries were collected from a local slaughterhouse from 30–40-day-old suckling lambs during both seasons. Following 24 h of in vitro maturation, oocytes developmental competence, reactive oxygen species (ROS) intracellular levels, and mitochondrial activity were evaluated, and a tubulin assessment was performed. The results on embryo production, as a percentage of first divisions and number of blastocysts obtained, were significantly higher in oocytes collected in the spring. Mitochondrial activity in oocytes was higher, and ROS production significantly lower, in spring than in autumn. Tubulin PTMs (tyrosinated and acetylated α-tubulin) showed a higher immunoreactivity in oocytes collected in spring compared with autumn sampling. Our data showed that seasons may affect the developmental competence, energetic status, and tubulin assessment of oocytes recovered from prepubertal ewes. Therefore, special care should be taken when choosing the period of the year for prepuberal ovine oocytes collection aimed at in vitro embryo reproduction programs.

Sign in / Sign up

Export Citation Format

Share Document