scholarly journals Tannin Supplementation Improves Oocyte Cytoplasmic Maturation and Subsequent Embryo Development in Pigs

Antioxidants ◽  
2021 ◽  
Vol 10 (10) ◽  
pp. 1594
Author(s):  
Zhi Yin ◽  
Jing-Tao Sun ◽  
Hong-Di Cui ◽  
Chao-Qian Jiang ◽  
Yu-Ting Zhang ◽  
...  
Keyword(s):  
Embryonic Development ◽  
Cumulus Cells ◽  
Cyclin B1 ◽  
High Concentration ◽  
Cumulus Expansion ◽  
Vitro Fertilization ◽  
Nuclear Maturation ◽  
Porcine Oocyte ◽  
Cytoplasmic Maturation

To investigate the effects of tannins (TA) on porcine oocyte in vitro maturation (IVM), different concentrations of TA (0, 1, 10 and 100 μg/mL) were supplemented with a maturation medium and the COCs and subsequent embryonic development were examined. The results showed that 10 µg/mL TA significantly improved the cumulus expansion index (CEI), cumulus-expansion-related genes (PTGS1, PTGS2, PTX-3, TNFAIP6 and HAS2) expression and blastocyst formation rates after parthenogenetic activation (PA), in vitro fertilization (IVF) and somatic cell nuclear transfer (SCNT) compared to the control groups, but not oocyte nuclear maturation. Nevertheless, 10 µg/mL TA dramatically enhanced the mRNA expression of oocyte-development-related genes (BMP15, GDF9, CDC2 and CYCLIN B1), GSH, ATP, SOD1, PGC1α, BMP15, GDF9 and CDC2 levels and reduced intracellular ROS level in porcine oocytes. These results indicated that porcine oocyte cytoplasmic maturation was improved by 10 µg/mL TA treatment during IVM. In contrast, a high concentration of TA (100 μg/mL) significantly decreased the CEI and PTGS1, PTGS2, PTX-3 and HAS2 mRNA expressions in cumulus cells, and reduced oocyte nuclear maturation and the total cell numbers/blastocyst. In general, these data showed that 10 μg/mL TA supplementation has beneficial effects on oocyte cytoplasmic maturation and subsequent embryonic development in pigs.

scholarly journals Porcine follicular fluid derived from > 8 mm sized follicles improves oocyte maturation and embryo development during in vitro maturation of pigs

Zygote ◽  
2020 ◽  
pp. 1-6
Author(s):  
Ji-Eun Park ◽  
Sang-Hee Lee ◽  
Yong Hwangbo ◽  
Choon-Keun Park
Keyword(s):  
Embryonic Development ◽  
Follicular Fluid ◽  
In Vitro Maturation ◽  
Developmental Competence ◽  
Control Group ◽  
Cumulus Expansion ◽  
Treatment Groups ◽  
Nuclear Maturation ◽  
Cytoplasmic Maturation

Summary The aim of the present study was to investigate the effects of porcine follicular fluid (pFF) from large-sized (LFF; >8 mm in diameter) and medium-sized (MFF; 3–6 mm in diameter) follicles on the maturation and developmental competence of porcine oocytes. Cumulus–oocyte complexes (COCs) were collected from follicles 3–6 mm in diameter. The collected COCs were incubated for 22 h with LFF or MFF (in vitro maturation (IVM)-I stage) and were incubated subsequently for 22 h with LFF or MFF (IVM-II stage). Cumulus expansion was confirmed after the IVM-I stage and nuclear maturation was evaluated after the IVM-II stage. Intracellular glutathione (GSH) and reactive oxygen species (ROS) levels were measured and embryonic development was evaluated. Relative cumulus expansion and GSH levels were higher in the LFF group compared with in the MFF group after the IVM-I stage (P < 0.05). After the IVM-II stage, the numbers of oocytes in metaphase-II were increased in the LFF group and GSH content was higher in all of the LFF treatment groups compared with in the MFF treatment groups during both IVM stages (P < 0.05). ROS levels were reduced by LFF treatment regardless of IVM stage (P < 0.05). Blastocyst formation and the total numbers of cells in blastocysts were increased in all LFF treatment groups compared with the control group (P < 0.05). These results suggested that pFF from large follicles at the IVM stage could improve nucleic and cytoplasmic maturation status and further embryonic development through reducing ROS levels and enhancing responsiveness to gonadotropins.

scholarly journals Chemical manipulation of glucose metabolism in porcine oocytes: effects on nuclear and cytoplasmic maturation in vitro

Reproduction ◽  
2006 ◽  
Vol 131 (2) ◽  
pp. 289-298 ◽  
Author(s):  
Jason R Herrick ◽  
Amber M Brad ◽  
Rebecca L Krisher
Keyword(s):  
Embryonic Development ◽  
Cumulus Cells ◽  
Blastocyst Stage ◽  
Pentose Phosphate ◽  
The Other ◽  
Nuclear Maturation ◽  
Maturation In Vitro ◽  
Intracellular Content ◽  
Cytoplasmic Maturation

The objectives of this study were to manipulate metabolism of glucose through glycolysis and the pentose phosphate pathway (PPP) in porcine oocytes during in vitro maturation, and determine the effects of this manipulation on meiotic progression, intracellular glutathione (GSX) concentrations and embryonic development. Cumulus-oocyte complexes isolated from abattoir ovaries were matured (40–44 h) in Purdue Porcine Medium for maturation alone (control) or supplemented with pyrroline-5 carboxylate (PC, 0.1 μM; PPP stimulator), diphenyleneiodonium (DPI, 0.1 μM; PPP inhibitor), dinitrophenol (DNP, 10 μM; glycolytic stimulator), hexametaphosphate (HMP, 100 μM; glycolytic inhibitor), PC + HMP or DNP + DPI. At the conclusion of in vitro maturation, cumulus cells were removed and oocytes were randomly allocated for analysis of GSX, metabolism and nuclear maturation, or in vitro fertilization and embryo culture. Both DPI and DNP + DPI decreased (P ≤ 0.05) the activity of glycolysis and the PPP, increased (P ≤ 0.05) the percentage of immature oocytes, and decreased (P ≤ 0.05) the proportion of mature oocytes compared with control oocytes and oocytes from the other treatments. Embryonic development (cleavage and blastocyst stage) and the intracellular content of GSX were also decreased (P ≤ 0.05) following exposure to DPI or DNP + DPI compared with control oocytes and oocytes from the other treatments. Oocyte metabolism, nuclear maturation, GSX content and embryonic development were unaffected (P > 0.05) following exposure to PC, DNP, HMP or PC + HMP. Our results suggest that metabolism of glucose through the PPP and/or glycolysis plays a key role in the control of nuclear and cytoplasmic maturation of porcine oocytes in vitro.

333 EFFECTS OF BOVINE OOCYTE QUALITY ON KINETICS OF NUCLEAR MATURATION AND EMBRYONIC DEVELOPMENT AFTER IN VITRO FERTILIZATION

2010 ◽  
Vol 22 (1) ◽  
pp. 322
Author(s):  
I. Carvalhais ◽  
M. Faheem ◽  
A. Habibi ◽  
A. Geraldo ◽  
R. Agrícola ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Embryonic Development ◽  
Cumulus Cells ◽  
Oocyte Quality ◽  
Blastocyst Stage ◽  
Morphological Aspect ◽  
Vitro Fertilization ◽  
Nuclear Maturation ◽  
Group I

Many factors act together to prepare the immature oocyte for successful development to a competent embryo after fertilization. Defects in oocyte maturation and further development can possibly be caused by the oocyte quality or an inadequate nuclear maturation or even by a failure of both. In the present study, the effect of COCs quality on meiotic development and further embryo-development after in vitro fertilization was evaluated. A total of 3604 COCs was separated according to their morphological aspect and were classified as A, B, and C categories. Briefly, in class A, oocytes possessed compact layers of cumulus cells, being difficult to evaluate their number having a homogenous ooplasm with uniform color. In class B, oocytes show more or equal to five layers of cumulus cells, easily identifiable under a stereomicroscope and/or granulations in the ooplasm. In class C, some granulation was observed in oocytes with about three layers of cumulus cells. The total number of oocytes was divided into two groups (I and II) in which in the group I, COCs (n = 540) were fixed 0, 6, 12, 18, 24, and 30 h following ovarian aspiration, DNA was stained with aceto-orcein, and the nucleus were observed under a phase contrast microscope. In the Group II, COCs (n = 3064) were fertilized with frozen/thawed bull semen after 24 h of maturation, which was made in M199 medium (Sigma, St, Louis, MO, USA). The development of the embryos was evaluated on the third and seventh day after fertilization. Embryos were co-cultured with monolayers of granulosa cells in 45 μL droplets of B2 medium (CCD Laboratory, Paris, France), supplemented with 10% serum under mineral oil, at 39°C and 5% CO2 in air. It was observed that, other than the oocytes achieved metaphase II at 24 h was greater for the oocytes classified as A (65.4%), and B (61.0%) greater than C (51.2%), no statistical difference was observed between oocyte quality and capability to maturation. As far as the embryonic development is concerned, the same tendency was observed for the cleavage and for the morulae/blastocyst stage after 7 days after fertilization (P < 0.001). The percentages of cleaved oocytes classified as A, B, and C, were respectively 65.2%, 58.4%, and 48.0%. The development to the morulae/blastocyst stage of the cleaved embryos was A = 38.5%/27.4%, B = 33.6%/25.0%, and C = 30.9%/17.2% (Table 1). The results of our study clearly demonstrate that the morphology of the oocytes plays an important role on the in vitro embryonic developmental competence after fertilization. Table 1.Development of oocytes according to COCs quality, evaluated 3 and 7 days after fertilization The first author is supported by the Regional Foundation for Science and Technology of the Azores Government. This study was supported by the IBBA Institute grant number M2.1.2/I/022/2008 CITA-A is fully acknowledged.

scholarly journals A Brief Incubation of Cumulus-Enclosed Mouse Eggs in a Calcium-Free Medium Containing a High Concentration of Calcium-Chelator Markedly Improves Preimplantation Development

2020 ◽  
Vol 17 (10) ◽  
pp. 3505
Author(s):  
Valeria Merico ◽  
Silvia Garagna ◽  
Maurizio Zuccotti
Keyword(s):  
In Vitro Fertilization ◽  
Mammalian Species ◽  
Developmental Rate ◽  
Cumulus Cells ◽  
Fertilization Rate ◽  
Preimplantation Development ◽  
High Concentration ◽  
Vitro Fertilization ◽  
Positive Effects

The presence of cumulus cells (CCs) surrounding ovulated eggs is beneficial to in vitro fertilization and preimplantation development outcomes in several mammalian species. In the mouse, this contribution has a negligible effect on the fertilization rate; however, it is not yet clear whether it has positive effects on preimplantation development. Here, we compared the rates of in vitro fertilization and preimplantation development of ovulated B6C3F1 CC-enclosed vs. CC-free eggs, the latter obtained either after a 5 min treatment in M2 medium containing hyaluronidase or after 5–25 min in M2 medium supplemented with 34.2 mM EDTA (M2-EDTA). We found that, although the maintenance of CCs around ovulated eggs does not increment their developmental rate to blastocyst, the quality of the latter is significantly enhanced. Most importantly, for the first time, we describe a further quantitative and qualitative improvement, on preimplantation development, when CC-enclosed eggs are isolated from the oviducts in M2-EDTA and left in this medium for a total of 5 min prior to sperm insemination. Altogether, our results establish an important advancement in mouse IVF procedures that would be now interesting to test on other mammalian species.

336 MEIOTIC DEVELOPMENT AND CORTICAL GRANULES DISTRIBUTION IN CANINE OOCYTES DURING IN VITRO MATURATION

2010 ◽  
Vol 22 (1) ◽  
pp. 324 ◽  
Author(s):  
M. De los Reyes ◽  
D. Luna ◽  
J. Palomino
Keyword(s):  
In Vitro Maturation ◽  
Germinal Vesicle ◽  
Fluorescein Isothiocyanate ◽  
Cumulus Cells ◽  
Homogeneous Distribution ◽  
Cortical Granules ◽  
Dapi Staining ◽  
Nuclear Maturation ◽  
Cytoplasmic Maturation

Low development of IVM canine oocytes could be in part attributed to an impaired cytoplasmic maturation. In mammalian oocytes, migration and the redistribution of cortical granules (CGs) around the periphery of the oocyte contribute to the inhibition of polyspermy and it is an important criterion to evaluate cytoplasmic maturation. The state of nuclear maturation and the distribution of CGs were evaluated in canine oocytes cultured for different periods in order to compare the synchrony of nuclear and cytoplasmic maturation during in vitro maturation. Bitch ovaries at different stages of the estrous cycle were obtained following ovariectomy. COCs with compact cumulus cells showing a homogeneous cytoplasm were selected for experiments. Thirty-six COCs were processed at immature stage, placed in PBS medium until evaluation. A total of 275 COCs were matured in vitro for 48, 72, and 96 h in TCM-199 with Earle’s salt supplemented with 25 mM Hepes, 10% FCS, 0.25 mM pyruvate, 10 IU mL-1 of hCG, 300 IU mL-1 penicillin, and 20 mg mL-1 streptomycin, at 38.5°C and 5% CO2. At each culture period, the oocytes were stained with Lens culinaris agglutinin (LCA), labeled with fluorescein isothiocyanate, and the CGs distributions were examined under a fluorescent microscope. The nuclear status of the denuded oocytes was determined by DAPI staining under a fluorescence microscope. For each treatment, at least four replicates were performed and the data was analyzed by ANOVA using Tukey’s test to determine the differences P < 0.05. Three types of CGs distribution were distinguished during canine oocyte maturation: (1) homogeneous distribution throughout the cytoplasm including the cortex; (2) heterogeneous (clusters) within the cytoplasm and (3) densely distributed beneath the oolemma. Nuclear stages were classified as immature or germinal vesicle (GV) stage; resumption of meiosis or germinal vesicle break down (GVBD); metaphase I to telophase I (MI toTel I); and mature or second metaphase (MII). The distribution patterns of GCs were different (P < 0.05) among oocytes cultured for different periods and the nuclear maturation status also differed between oocytes cultured for different intervals (P < 0.05). Most (>84%) of the immature oocytes at GV showed a uniform distribution of CGs throughout the cytoplasm. At 48 h of culture, CGs distribution was mainly Type 2 (25%) and 3 (61%) and the oocytes were at GVBD (33%) and MI-Tel I (33%) stages. Most nuclei of the type 3 oocytes were in the MI (40%) and MII (11%) stages, corresponding to those oocytes matured for 72 (88%) or 96 h (71%). These results indicate that canine oocytes migrate to the cortex during IVM and this process is not finished before 72 h of culture. In addition, although the re-distribution of the CGs occurred in parallel with nuclear maturation, the oocytes cannot always proceed to the MII stage; however, in such oocytes the CGs are distributed beneath the oolemma. Supported by Grant FONDECYT 1080618.

172 DETERMINING THE REQUIREMENTS FOR LH AND FSH DURING SHEEP IN VITRO OOCYTE MATURATION

2014 ◽  
Vol 26 (1) ◽  
pp. 200 ◽  
Author(s):  
C. de Frutos ◽  
R. Vicente-Perez ◽  
P. J. Ross
Keyword(s):  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Mixed Logit ◽  
Cumulus Cells ◽  
Pituitary Hormones ◽  
Developmental Competence ◽  
Cumulus Expansion ◽  
Nuclear Maturation ◽  
Grade 3

In vitro maturation (IVM) of oocytes in domestic animals is a widespread practice of research and commercial relevance. Gonadotropic hormones are typically supplemented to the IVM medium to stimulate resumption of meiosis, progression to metaphase II (MII), and oocyte developmental competence. The common use of pituitary-derived products presents 2 problems: contamination from other pituitary hormones and inconsistences from batch-to-batch variation. Recombinant hormones can help circumvent these issues and identify specific gonadotropin requirements for in vitro maturation. The aim of the present study was to determine the effect of supplementing recombinant bovine LH and/or FSH (AspenBio) to the maturation of ovine oocytes in terms of cumulus expansion and progression to the MII stage. Abattoir-derived sheep cumulus–oocyte complexes (COC) were obtained from 1- to 5-mm-diameter antral follicles by ovary slicing. Oocytes with a homogeneous cytoplasm surrounded by at least 3 layers of cumulus cells were selected and cultured in serum-free IVM medium (Cotterill et al. 2012 Reproduction 144, 195–207) at 38.5°C and 5% CO2. The COC obtained from 8 replicates were allocated into 4 experimental groups: (1) no hormones; (2) 1.5 μg mL–1 recombinant bovine LH (rbLH); (3) 1.5 μg mL–1 recombinant bovine FSH (rbFSH); and (4) rbLH and rbFSH. The expansion of cumulus cells was recorded in each group after 24 h of IVM and COC classified as (1) very poor or no cumulus expansion (grade 1); (2) limited cumulus expansion (grade 2); and (3) full cumulus expansion (grade 3). Nuclear maturation in the 4 treatments was evaluated by assessing progression to the MII stage via DNA staining with Hoechst 33342 and fluorescence imaging. The effect of treatment on the observed proportion of MII oocytes was evaluated using a mixed logit model including treatment and replicate as fixed and random effects, respectively. Culture in IVM medium in the absence of gonadotropins or in the presence of rbLH resulted in poor cumulus expansion (grade 1). The supplementation of IVM medium with rbFSH (with or without rbLH) yielded a high degree of cumulus expansion (grades 2–3). Likewise, addition of rbFSH enhanced progression of oocytes to the MII stage, whereas use of rbLH, although it had an effect on progression to MII, did not augment the effect of rbFSH (Table 1). These results indicate that rbFSH is necessary and sufficient to induce sheep oocyte maturation in a high proportion of oocytes. Table 1.Cumulus expansion and oocyte nuclear stage after IVM

208 EFFECT OF GDF8 AND SB-431542 ON PORCINE OOCYTE DURING IN VITRO MATURATION AND SUBSEQUENT EMBRYONIC DEVELOPMENT

2016 ◽  
Vol 28 (2) ◽  
pp. 235
Author(s):  
J. D. Yoon ◽  
E. Lee ◽  
S.-H. Hyun
Keyword(s):  
Treatment Group ◽  
Embryonic Development ◽  
Follicular Fluid ◽  
In Vitro Maturation ◽  
Transforming Growth Factor ◽  
Formation Rate ◽  
Transforming Growth Factor Β ◽  
Nuclear Maturation ◽  
Porcine Oocyte

Growth differentiation factor-8 (GDF8) is a member of the transforming growth factor-β that has been identified as a strong physiological regulator. SB-431542 (SB) is a specific inhibitor of transforming growth factor-β superfamily type I activin receptor-like kinase (ALK) receptors such as ALK4, ALK5, and ALK7. The purpose of this study is investigation of the effects of GDF8 and SB on porcine oocytes during in vitro maturation and subsequent embryonic development. We first performed ELISA to detect GDF8 concentrations in follicular fluid for each size of follicle; sizes were as follows: small (<3 mm), medium (>3 mm and <6 mm), and large (>6 mm) follicle. After detection of the GDF8 concentration in follicular fluid, we investigated the effect of GDF8 and SB treatment during in vitro maturation (IVM) on nuclear maturation, intracellular glutathione (GSH), and reactive oxygen species (ROS) levels, and embryonic development after IVF and parthenogenetic activation (PA). Data were analysed by ANOVA followed by Duncan using SPSS (Statistical Package for Social Science, IBM, New York, NY, USA) mean ± SEM. The ELISA result showed different concentrations of GDF8 for each grade of follicular fluid: small, 0.479 ng mL–1; medium, 0.668 ng mL–1; and large, 1.318 ng mL–1. During the IVM process, 1.318 ng mL–1 of GDF8 and 5 ng mL–1 of SB were added to the maturation medium as control, SB, SB+GDF8, and GDF8 treatment groups. After 44 h of IVM, GDF8 group (90.4%) showed a significantly higher nuclear maturation rate than control and SB+GDF8 groups (85.4 and 81.7%). The SB group (78.9%) showed significantly reduced nuclear maturation rate compared with control (P < 0.05). The GDF8 treatment group showed a significant decreased intracellular ROS and increased GSH levels compared with other groups (P < 0.05). The SB+GBF8 treatment group showed a significantly better cytoplasmic maturation than the SB treatment group. In the PA embryonic development analysis, the GDF8 treatment group showed a significantly higher blastocyst formation rate compared with other groups (47.9, 37.2, 46.4, and 58.7% respectively; P < 0.05). In the IVF embryonic development analysis, the GDF8 treatment groups showed significantly higher blastocyst formation rate compared with the SB group (28.2 and 42.2%, respectively; P < 0.05). In conclusion, treatment with GDF8 during porcine oocyte IVM improved the embryonic developmental competence via increased cytoplasmic maturation and led to better oocyte maturation from the ALK receptor inhibition by SB.

160 Exposure to Cadmium Affects Oocyte and Embryo Competence in Cattle

2018 ◽  
Vol 30 (1) ◽  
pp. 219
Author(s):  
C. De Canditiis ◽  
N. Pagano ◽  
V. Franco ◽  
I. Paradiso ◽  
É. C. Dos Santos ◽  
...  
Keyword(s):  
Embryo Development ◽  
Cumulus Cells ◽  
Early Embryo ◽  
Dapi Staining ◽  
Vitro Fertilization ◽  
Nuclear Maturation ◽  
Significant Difference ◽  
Vitro Model ◽  
Hoechst Staining

There is a growing worldwide concern regarding the increased release of the heavy metal cadmium (Cd) in the environment, due to several industrial processes, as it is known to affect health. Among other heavy metals, Cd is widely recognised to influence the reproductive system at different levels, interfering with both gametes and embryo functions in several species (Thompson and Bannigan, 2008 Reprod. Toxicol. 25, 304-315). The in vitro model can be used to mimic environmental conditions allowing us to evaluate their effect on oocyte maturation and early embryo development. Therefore, the aim of this study was to evaluate the influence of different Cd concentrations on nuclear maturation, apoptosis in cumulus cells, and cleavage and blastocyst yields in cattle. For this purpose, abattoir-derived bovine oocytes were in vitro matured, fertilized, and cultured according to standard procedures (Rubessa et al. 2011 Theriogenology 76, 1347-1355). In particular, oocytes were matured with 0 (control; n = 126), 0.1 μM (n = 139), 1 μM (n = 134), and 10 μM of Cd (n = 135), at 39°C under humidified air with 5% CO2, 7% O2, and 88% N2. For each replicate, after 22 h of maturation, a representative sample of oocytes (n = 10 per each group) was used to evaluate nuclear maturation by 4′,6-diamidino-2-phenylindole (DAPI) staining and another sample (n = 10 per each group) to assess cumulus-cells complex apoptosis by TUNEL/Hoechst staining (Pocar et al. 2005 Reproduction 130, 857-868). The remaining oocytes were in vitro fertilized and cultured with 0 (n = 106), 0.1 μM (n = 119), 1 μM (n = 114), and 10 μM (n = 115) Cd. The experiment was repeated 3 times. On Day 8 post-IVF, the blastocyst yields were recorded. Differences among groups were analysed by ANOVA, with the least significant difference method used as a post hoc test. Data are presented as means ± SE. Unexpectedly, the exposure of oocytes to Cd during IVM did not affect the percentage of oocytes undergoing nuclear maturation (on average 96.3 ± 2.3). In contrast, concentrations of 1 and 10 μM Cd increased the percentage of apoptotic cumulus-cells in cumulus–oocyte complexes (COC) compared with the control (3.4 ± 0.4, 10.6 ± 1.8, 15.0 ± 0.9, 16.7 ± 4.0, respectively, with 0, 0.1, 1, and 10 μM; P < 0.05). It is worth pointing out that with the highest concentration, cumulus expansion did not occur and cumulus cells appeared detached from the oocyte. Likewise, 1 and 10 μM Cd decreased cleavage rates compared with the control (68.7 ± 1.8, 54.3 ± 5.0, 58.5 ± 4.2 and 2.8 ± 2.6, respectively, with 0, 0.1, 1, and 10 μM Cd; P < 0.01). Finally, blastocyst yields decreased when oocytes were treated with 0.1 μM Cd and no development to blastocyst was observed at the 2 higher concentrations (35.1 ± 1.7, 26.2 ± 3.1, 0, 0, respectively, with 0, 0.1, 1, and 10 μM; P < 0.01). In conclusion, exposure to Cd during maturation negatively affects bovine COC, as indicated by the increased apoptotic index in cumulus cells, without influencing the nuclear maturation process. Furthermore, the presence of Cd during in vitro fertilization and culture severely impairs both the fertilization and post-fertilization embryo development.

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