First polar body morphology affects potential development of porcine parthenogenetic embryo in vitro

Zygote ◽  
2014 ◽  
Vol 23 (4) ◽  
pp. 615-621 ◽  
Author(s):  
Junhe Hu ◽  
Chenzhong Jin ◽  
Hui Zheng ◽  
Qinyan Liu ◽  
Wenbing Zhu ◽  
...  
Keyword(s):  
Embryo Development ◽  
Polar Body ◽  
Cell Number ◽  
Cortical Granules ◽  
Potential Development ◽  
First Polar Body ◽  
Porcine Embryo ◽  
Parthenogenetic Embryo ◽  
The Mean

SummaryPrevious studies have reported that the first polar body (PB1) morphology reflects embryo development competence, but the effects of PB1 on porcine embryo development remain unknown. This study aims to determine whether the ability of porcine embryo development is related to oocytes’ PB1 in vitro. The distribution of type II cortical granules (CGs) of porcine matured oocytes in grade B PB1 is significantly greater compared with those in grades A and C PB1 (71.43% versus 52.46% and 50%; P < 0.05). The ratio of porcine parthenogenetic blastocysts and the mean cell number in each blastocyst in the group with grade B PB1 is significantly greater than that with grades A and C PB1 (30.81% vs. 19.02% and 15.15%; P < 0.05) and (36.67 versus 24.67, 28.67; P < 0.05), and no significant differences are found in the embryo cleavage for all groups (79.75%, 84.30%, and 78.18% in grades A, B, and C PB1; P > 0.05). The acetylation level of porcine embryos in the group with grade B PB1 is significantly greater compared with those in the other groups (P < 0.05), and is almost 2.5 times higher than that in grade A. Therefore, porcine oocytes with PB1 in grade B are more competitive in cytoplasmic maturation and further embryo development in vitro.

Embryo development and sex ratio of in vitro-produced porcine embryos are affected by the energy substrate and hyaluronic acid added to the culture medium

2014 ◽  
Vol 26 (4) ◽  
pp. 570 ◽  
Author(s):  
Eva Torner ◽  
Eva Bussalleu ◽  
M. Dolors Briz ◽  
Marc Yeste ◽  
Sergi Bonet
Keyword(s):  
Hyaluronic Acid ◽  
Sex Ratio ◽  
Embryo Development ◽  
Culture Medium ◽  
Cell Number ◽  
Total Cell ◽  
Total Cell Number ◽  
Porcine Embryo ◽  
Preferential Loss

In the present study, the effects of replacing glucose with pyruvate–lactate and supplementing these in vitro culture (IVC) media with hyaluronic acid (HA) on porcine embryo development and sex ratio were examined. The in vitro-produced (IVP) porcine embryos were cultured in NCSU-23 medium with 0.0, 0.5 or 1.0 mg mL–1 HA, and with either 5.55 mM glucose (IVC-Glu) or pyruvate (0.17 mM)–lactate (2.73 mM) from 0 to 48 h post insemination (h.p.i.) and then with glucose from 48 to 168 h.p.i. (IVC-PL). Those embryos cultured with IVC-PL had significantly higher blastocyst rates (23.7 ± 1.5%) than those cultured with IVC-Glu (14.27 ± 2.75%). At 1.0 mg mL–1, HA tended to skew the sex ratio of blastocysts towards males in those embryos cultured in IVC-PL, and led to a significant decrease in the blastocyst rate compared with embryos cultured in the presence of 0.5 and 0.0 mg mL–1 HA and IVC-Glu (4.28 ± 0.28% vs 11.01 ± 1.42% and 10.14 ± 2.77%, respectively) and IVC-PL (14.37 ± 1.35% vs 20.96 ± 2.85% and 22.99 ± 1.39%, respectively). In contrast, there were no significant differences in the total cell number per blastocyst or in apoptosis rates. In conclusion, pyruvate and lactate were the preferred energy substrates in the early stages of IVP porcine embryos. Moreover, 1.0 mg mL–1 HA significantly decreased the percentage of blastocyst rates in both the IVC-Glu and IVC-PL groups, but only by a preferential loss of female embryos for those cultured in IVC-PL.

53 EFFECT OF THE NUCLEAR REPROGRAMMING TIME ON IN VITRO DEVELOPMENT OF EQUINE AGGREGATED CLONED EMBRYOS

2013 ◽  
Vol 25 (1) ◽  
pp. 174
Author(s):  
R. Olivera ◽  
C. Alvarez ◽  
I. Stumpo ◽  
G. Vichera
Keyword(s):  
Embryo Development ◽  
Polar Body ◽  
Nuclear Reprogramming ◽  
Chi Square ◽  
In Vitro Development ◽  
First Polar Body ◽  
Group 3 ◽  
Group 2 ◽  
Vitro Development

The time allowed for nuclear reprogramming is considered an essential factor for the efficiency of cloning and has not been evaluated in equine aggregated cloned embryos. The aim of our work was to assess the effect of different timing of activation stimulus after fusion of adult equine fibroblast cells to enucleated equine oocytes on embryo development and embryo quality. We processed a total of 1874 equine ovaries, recovering 3948 oocytes, of which 1914 (48.5%) had extruded the first polar body after 24 h of maturation. Oocyte collection, maturation, and the NT procedure were performed as described by Lagutina et al. (2007 Theriogenology 67, 90–98). Reconstructed oocytes (RO) were activated at 3 different times after cell fusion: (1) 1 h, (2) 1.5 h, and (3) 2 h. Activation was performed using 8.7 µM ionomycin for 4 min, followed by a 4-h culture in a combination of 1 mM DMAP and 5 mg mL–1 of cycloheximide. The RO were cultured in the well of the well system, aggregating 3 RO per well. The RO were cultured in DMEM-F12 with 5% fetal bovine serum (FBS) and antibiotics. Cleavage (48 h after activation), blastocyst, and expanded blastocyst rates (8–9 days) were assessed. In vitro development was compared using the chi-square test (P < 0.05). A total of 1608 RO were cultured. Cleavage was significantly lower in group 3 with respect to the other 2 groups [(1): 396/450, 88%; (2): 540/639, 84.5%; (3): 365/519, 70.3%]. There were no significant differences in blastocyst rates within the 3 groups considering the number of total RO [(1): 19/450, 4.2%; (2): 23/639, 3.6%; (3): 15/519, 2.9%] or aggregated RO per well [(1): 12.7%; (2): 10.8%; (3): 8.7%]. However, the rate of blastocyst expansion was higher (P < 0.05) in group 2 than in group 3 [(1): 17/19, 89.5%; (2): 23/23, 100%; (3): 11/15, 73.3%]. In conclusion, the timing of nuclear reprogramming did not affect blastocyst rates but affected cleavage rates and blastocyst quality. This indicates that 1 h before activation stimulus is enough for embryo development of equine aggregated cloned embryos.

scholarly journals Ultrastructure of in vitro oocyte maturation in buffalo (Bubalus bubalis)

Zygote ◽  
2010 ◽  
Vol 18 (4) ◽  
pp. 309-314 ◽  
Author(s):  
Rafael Gianella Mondadori ◽  
Tiago Rollemberg Santin ◽  
Andrei Antonioni Guedes Fidelis ◽  
Khesller Patrícia Olázia Name ◽  
Juliana Souza da Silva ◽  
...  
Keyword(s):  
Polar Body ◽  
Bubalus Bubalis ◽  
Ultrastructural Changes ◽  
Oocyte Nucleus ◽  
Cortical Granules ◽  
Immature Oocytes ◽  
Transmission Electron ◽  
High Lipid ◽  
First Polar Body

SummaryThe objective of the present study was to describe ultrastructural changes in the nucleus and cytoplasmic organelles during in vitro maturation (IVM) of buffalo cumulus–oocyte complexes (COCs). The structures were collected by ovum pick-up (OPU). Some COCs, removed from maturation medium at 0, 6, 12, 18 and 24 h, were processed for transmission electron microscopy. The average number of COCs collected by OPU/animal/session was 6.4, and 44% of them were viable. Immature oocytes had a peripherally located nucleus, Golgi complex and mitochondrial clusters, as well as a large number of coalescent lipid vacuoles. After 6 h of IVM, the oocyte nucleus morphology changed from round to a flatter shape, and the granulosa cells (GC) lost most of their contact with zona pellucida (ZP). At 12 h the first polar body was extruded and the aspect of lipid droplet changed to dark, probably denoting lipid oxidation. Cortical granules were clearly visible at 18 h of maturation, always located along the oocyte periphery. At 24 h of IVM the number of cortical granules increased. Ultrastructure studies revealed that: (1) immature oocytes have a high lipid content; (2) the perivitelline space (PS) increases during IVM; (3) Golgi complexes and mitochondrial clusters migrate to oocyte periphery during IVM; (4) 6 h of IVM are enough to lose contact between GC and ZP; (5) the oocyte lipid droplets’ appearance changes between 6 and 12 h of IVM.

scholarly journals Luteolin Orchestrates Porcine Oocyte Meiotic Progression by Maintaining Organelle Dynamics Under Oxidative Stress

2021 ◽  
Vol 9 ◽  
Author(s):  
Soo-Hyun Park ◽  
Pil-Soo Jeong ◽  
Ye Eun Joo ◽  
Hyo-Gu Kang ◽  
Min Ju Kim ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Oocyte Maturation ◽  
Polar Body ◽  
Blastocyst Formation ◽  
Cortical Granules ◽  
Expression Levels ◽  
Meiotic Progression ◽  
First Polar Body ◽  
Underlying Mechanisms

Increasing evidence has demonstrated that oxidative stress impairs oocyte maturation, but the underlying mechanisms remain largely unknown. Here, for the first time, we examined the antioxidant role of luteolin in meiotic progression and the underlying mechanisms. Supplementation of 5 μM luteolin increased the rates of first polar body extrusion and blastocyst formation after parthenogenetic activation, and the expression levels of oocyte competence (BMP15 and GDF9)-, mitogen-activated protein kinase (MOS)-, and maturation promoting factor (CDK1 and Cyclin B)-related genes were also improved. Luteolin supplementation decreased intracellular reactive oxygen species levels and increased the expression levels of oxidative stress-related genes (SOD1, SOD2, and CAT). Interestingly, luteolin alleviated defects in cell organelles, including actin filaments, the spindle, mitochondria, the endoplasmic reticulum, and cortical granules, caused by H2O2 exposure. Moreover, luteolin significantly improved the developmental competence of in vitro-fertilized embryos in terms of the cleavage rate, blastocyst formation rate, cell number, cellular survival rate, and gene expression and markedly restored the competencies decreased by H2O2 treatment. These findings revealed that luteolin supplementation during in vitro maturation improves porcine meiotic progression and subsequent embryonic development by protecting various organelle dynamics against oxidative stress, potentially increasing our understanding of the underlying mechanisms governing the relationship between oxidative stress and the meiotic events required for successful oocyte maturation.

44 EFFECT OF CYTOPLAST VOLUME ON FERTILIZATION AND EMBRYO DEVELOPMENT IN PORCINE M-II OOCYTES RECONSTRUCTED WITH KARYOPLASTS AND CYTOPLASTS OBTAINED BY THE 'CENTRI-FUSION' METHOD

2008 ◽  
Vol 20 (1) ◽  
pp. 102
Author(s):  
N. Maedomari ◽  
K. Kikuchi ◽  
M. Fahrudin ◽  
N. Nakai ◽  
M. Ozawa ◽  
...  
Keyword(s):  
Polar Body ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Developmental Competence ◽  
First Polar Body ◽  
Developmental Capacity ◽  
Sperm Penetration ◽  
Pronuclear Formation ◽  
And Control

Metaphase-II chromosome transfer (M-II transfer) of oocytes is considered to be one of the advanced procedures to improve fertilization and developmental abilities of oocytes with poor cytoplasmic maturation. The aim of this study was to investigate the developmental capacity after IVF and IVC of porcine oocytes reconstructed from karyoplasts and cytoplasts produced by centri-fusion (Fahrudin et al. 2007 Cloning Stem Cells 9, 216–228). In brief, IVM oocytes (Kikuchi et al. 2002 Biol. Reprod. 66, 1033–1041) with a visible first polar body were centrifuged at 13 000g for 9 min to stratify the cytoplasm. Then the zonae pellucidae were removed with pronase treatment. Zona-free oocytes were layered on a 300-µL discontinuous gradient of Percoll in TCM-HEPES with 5 µg mL–1 of cytochalasin B. After centrifugation at 6000g for 4 s, fragmented cytoplasms with approximately equal volumes were obtained, stained with Hoechst-33342, and classified into cytoplasm with (K; karyoplast) or without (C; cytoplast) chromosomes. One karyoplast was fused with 0, 1, 2, 3, and 4 cytoplasts (K, K + 1C, K + 2C, K + 3C, and K + 4C, respectively) by an electric stimulation with a single DC pulse (1.5 kV cm–1 for 20 µs) and cultured for 1 h. Zona-free oocytes without any reconstruction served as control oocytes. The diameters of the reconstructed and control oocytes were measured. All specimens were fertilized in vitro with frozen–thawed boar sperm, and cultured using the well of the well (WOW) system (Vajta et al. 2000 Mol. Reprod. Dev. 55, 256–264). Their fertilization status and developmental competence were examined. Data were analyzed by ANOVA followed by Duncan's multiple range tests. The diameter differed significantly among K to K + 4C oocytes (75.0–127.1 µm; P < 0.05), whereas the diameter of K + 2C oocytes was similar to that of the control oocytes (110.5 µm). Regardless of the cytoplast volume, sperm penetration rates (73.1–93.8%) for K to K + 4C oocytes were not significantly different compared to control oocytes (78.0%). Male pronuclear formation rates of K to K + 4C oocytes (92.3–97.1%) were also not different significantly different compared to control oocytes (96.6%). However, monospermy rates of K oocytes was significantly higher (61.6%; P < 0.05) than those of the reconstructed (K + 1C to K + 4C; 18.2–34.9%) and control oocytes (32.9%). The blastocyst formation rates in K, K + 1C, K + 2C, and K + 3C groups (0.0–9.8%; P < 0.05) were significantly lower than those in the control and K + 4C groups (17.8% and 15.3%, respectively; P < 0.05). The total cell numbers per blastocyst in K + 1C and K + 2C groups (7.5 and 8.3 cells, respectively) were significantly lower than in the control, K + 3C, and K + 4C groups (15.3–26.2 cells; P < 0.05). These results suggest that the cytoplast volume of porcine M-II transferred oocytes, produced by reconstruction from a karyoplast and cytoplast(s) and centri-fusion, is important for their ability to develop to the blastocyst stage and influences cell number.

332 EFFECT OF DIFFERENT PIG OOCYTE ACTIVATION PROTOCOLS ON EMBRYO DEVELOPMENT IN SOF AND NCSU-23

2007 ◽  
Vol 19 (1) ◽  
pp. 281 ◽  
Author(s):  
I. Lagutina ◽  
G. Lazzari ◽  
C. Galli
Keyword(s):  
Embryo Development ◽  
Embryo Culture ◽  
Cell Number ◽  
Additional Treatment ◽  
Oocyte Activation ◽  
Blastocyst Development ◽  
Chi Square ◽  
Parthenogenetic Embryo ◽  
Pig Oocyte

Several factors affect nuclear transfer success. These include efficient parthenogenetic activation and embryo culture medium that should efficiently support pre-implantation development of good quality blastocysts. We investigated pig oocyte activation and embryo development in SOFaa in response to ionomycin (Io = 5 µM Io for 4 min; Io° = 15 µM Io for 20 min) and electric impulse (EL; one 30-µs pulse of DC 1.5 kV cm−1 in the presence of 50 µM Ca) in combination with 2 mM 6-DMAP or 10 µg mL−1 cycloheximide (CHX) +5 µg mL−1 cytochalasin B (CB) for 4 h. In addition, we studied the effect of elevated (1 mM) (Cheong et al. 2002 Mol. Reprod. Dev. 61, 488) in comparison with 50 µM Ca during EL activation on embryo development in SOFaa and NCSUaa-23. Porcine oocytes were recovered from slaughtered donors and matured in vitro for 44 h in DMEM-F12 supplemented with 10% FCS, 0.05 IU LH and FSH (Menogon®, Ferring, Milan, Italy), 0.3 mM cystine, 0.5 mM cysteamine, 50 ng mL−1 long-EGF, 100 ng mL−1 long-IGF1, 5 ng mL−1 bFGF (Sigma-Aldrich, Milan, Italy) in 5% CO2 at 38.5°C. The rates of cleavage, blastocyst formation (BL) and BL cell number on Day 7 (BL-D7) were recorded. All experiments were done with 3 replicates. The data were compared by chi-square test. There was no difference in the ability of Io (all groups) and EL + CB activated oocytes to cleave, whereas the additional treatment of EL-activated oocytes with DMAP and CHX + CB significantly increased cleavage. Io activation resulted in poor blastocyst development in comparison with all EL-activated groups (see Table 1). When calcium levels were elevated during EL activation, significantly more embryos developed in SOFaa (35.6%, n = 191 vs. 26%, n = 192; P &lt; 0.05), but no differences were observed with culture in NCSUaa-23 (about 56%). The BL rate was significantly higher in NCSUaa-23 vs. SOFaa (55.9%, n = 68 vs. 34.8%, n = 69, respectively); however, the BL total cell number was significantly higher in SOFaa (58 ± 18, n = 40 vs. 86 ± 35, n = 56, respectively; P &lt; 0.05). In conclusion, we have found that SOFaa and NCSUaa-23 differ in ability to support pig parthenogenetic embryo development. EL activation combined with elevated Ca significantly increased the embryo developmental capacity in SOFaa but not in NCSUaa-23. NCSUaa-23 was more efficient for embryo culture, whereas SOF produced BLs of higher quality. Table 1.Effect of activation protocol on the development of pig parthenogenetic embryos in SOFaa This work was supported by grants ISS-CS11 and Fondazione Cariplo.

62 DIFFERENT EFFECT OF TRICHOSTATIN A ON IN VITRO DEVELOPMENT OF PORCINE TRANSGENIC SOMATIC CELL NUCLEAR TRANSFER AND PARTHENOGENETICALLY ACTIVATED EMBRYOS

2008 ◽  
Vol 20 (1) ◽  
pp. 112 ◽  
Author(s):  
H. X. Wei ◽  
K. Zhang ◽  
Y. F. Ma ◽  
Y. Li ◽  
Q. Y. Li ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Trichostatin A ◽  
Polar Body ◽  
Cell Number ◽  
Cell Numbers ◽  
First Polar Body ◽  
Electrical Fusion

Accumulating evidence suggests that trichostatin A (TSA), a histone deacetylase inhibitor, can increase the success rate of somatic cloning. The objective of this study was to investigate the effect of 50 nm TSA treatment on the development of porcine somatic cell nuclear transfer (SCNT) and parthenogenically activated (PA) embryos. Cumulus-oocyte complexes were matured in vitro. The oocytes with the first polar body (PB1) were chosen for SCNT, and the rest with PB1 or good morphology were selected for PA by a single 100-μs direct current pulse of 1.6 kV cm–1, the same parameter as for electrical fusion. GFP transgenic fetal fibroblast cells were used as nuclear donors. Data were analyzed using SPSS (13.0; SPSS, Inc., Chicago, IL, USA) with one-way ANOVA. In Experiment 1, immediately after electrical fusion and activation, the reconstructed embryos were randomly cultured in porcine zygote medium 3 (PZM3) with 10 μg mL–1 cytochalasin B (CB) and 10 μg mL–1 cycloheximide (CHX), with either 0 nm (control) or 50 nm TSA for the first 4 h, before being cultured for another 20 h in PZM3 without CB and CHX. After being washed, the embryos were cultured in PZM3 medium without TSA until Day 6 at 39.0°C, 5% CO2, 5%O2, 90% N2, and 100% humidity. The same experimental design was used for PA embryos concurrently. The results showed that there were no significant differences in blastocyst rates for SCNT or PA between control and TSA groups (23.0 ± 6.1% v. 27.9 ± 6.3%; 21.0 ± 1.0% v. 17.5 ± 3.2%, respectively). Neither were there differences in the cell numbers of blastocysts (38.3 ± 5.7 v. 32.2 ± 3.4; 42.2 ± 3.5 v. 39.0 ± 1.9, respectively). In Experiment 2, TSA treatment was prolonged to either 36 or 40 h. The blastocyst rates of SCNT were increased (7.3 ± 1.2% (0 h), 13.3 ± 2.6% (36 h), and 20.0 ± 3.3% (40 h)), whereas those of PA were decreased (46.7 ± 5.0% (0 h), 27.7 ± 6.5% (36 h), and 30.8 ± 6.3% (40 h)). The cell numbers of blastocysts from either SCNT or PA were also decreased (SCNT: 47.5 ± 3.8, 37.5 ± 2.0, and 37.1 ± 3.3; PA: 46.1 ± 1.9, 37.5 ± 1.9, and 39.3 ± 2.2; P < 0.05). In Experiment 3, the cell number and the apoptotic index of Day 5, 6, and 7 PA blastocysts treated with 0 or 50 nm TSA were determined by the terminal deoxynucleotide-mediated nick end labeling (TUNEL) assay (Table 1). The results suggested that TSA treatment probably delayed embryo development, which may be one of the reasons for the lower cell numbers in the TSA-treated group. Table 1. Cell apoptosis of PA blastocyst by TUNEL

scholarly journals 141 BENEFICIAL EFFECT OF ETHYLENEDIAMINETETRAACETIC ACID COMBINED WITH HEMOGLOBIN ON PRE-IMPLANTATION DEVELOPMENT OF PORCINE IN VITRO PRODUCED EMBRYOS

2005 ◽  
Vol 17 (2) ◽  
pp. 221
Author(s):  
J.H. Kim ◽  
G.S. Lee ◽  
H.S. Kim ◽  
S.H. Lee ◽  
D.H. Nam ◽  
...  
Keyword(s):  
Beneficial Effect ◽  
Embryo Development ◽  
Ethylenediaminetetraacetic Acid ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Total Cell ◽  
Blastocyst Formation ◽  
Total Cell Number ◽  
Porcine Embryo

Developing a porcine embryo culture system is important for increasing the rates of implantation and pregnancy of somatic cell nuclear transfer (SCNT) embryos. Ethylenediaminetetraacetic acid (EDTA) was shown to inhibit glycolytic activity of cleavage stage embryos, thereby preventing the premature stimulation of glycolysis and enhancing development. However, EDTA should not be used for later-stage embryos as the inhibition of glycolysis reduces energy production at the blastocyst stage and significantly inhibits inner cell mass development. On the other hand, addition of a nitric oxide (NO) scavenger, hemoglobin (Hb), to the culture medium is known to promote embryo development to the blastocyst stage. This study was conducted to evaluate the beneficial effect of EDTA combined with Hb on pre-implantation development of porcine embryos in vitro. Porcine embryos produced by in vitro maturation and fertilization were cultured for 6 days in North Carolina State University (NCSU)-23 medium supplemented with EDTA or/and Hb. All data were subjected to one-way ANOVA and protected least significant difference (LSD) test using the general linear models (GLM) procedure of the statistical analysis system (SAS Institute, Inc., Cary, NC, USA) program to determine differences among experimental groups. Statistical significance was determined when the P value was less than 0.05. In Exp. 1, culturing porcine zygotes with 100 mM EDTA (n = 537) significantly increased cleavage rates (85.3%) at 48 h post-insemination compared to supplementing with 0, 1, or 10 mM EDTA (78.9, 79.7, or 78.2%, respectively). However, EDTA at these concentrations did not promote blastocyst formation compared to the control. In addition, no difference was observed in total cell numbers in blastocysts among the experimental groups (41.8, 42.6, 45.8, 44.5, respectively). In Exp. 2, in vitro-fertilized oocytes were cultured with 0, 1, or 10 mg/mL Hb. Culturing with Hb did not promote porcine embryo development, but significantly increased the total cell number of blastocysts obtained from 1 mg/mL Hb supplementation (n = 566) compared to that of the control (56.8 vs. 41.6). In Exp. 3, culturing embryos (n = 548) with 100 mM EDTA + 1 mg/mL Hb significantly improved rates of cleavage (84.0% vs. 75.2%) and blastocyst formation (19.2% vs. 12.7%), and the total number of cells in blastocysts compared to those of the control (58.4 vs. 42.3). In conclusion, our results demonstrated that EDTA or Hb have different roles in supporting in vitro pre-implantation development of porcine embryos; EDTA mainly stimulated early cleavage up to the 2- to 4-cell stage, and Hb promoted the total cell number of blastocysts. However, combined supplementation with these two chemicals improved cleavage, blastocyst formation, and total cell number in blastocysts. This study was supported by a grant from Korea Ministry of Science and Technology (Biodiscovery).

Cytoplasmic maturation of the marsupial oocyte during the periovulatory period

1996 ◽  
Vol 8 (4) ◽  
pp. 509 ◽  
Author(s):  
KE Mate
Keyword(s):  
Zona Pellucida ◽  
Polar Body ◽  
Monodelphis Domestica ◽  
Cortical Granules ◽  
Nuclear Maturation ◽  
Bettongia Penicillata ◽  
Maturation In Vitro ◽  
First Polar Body

During the period immediately before ovulation, the oocytes of most eutherian and marsupial mammals complete the first meiotic maturation division and extrude the first polar body. In marsupials, this phase of nuclear maturation is accompanied by an increase in size of the egg and maturation of cytoplasmic components. Oocytes from at least four marsupial species, Trichosurus vulpecula, Macropus eugenii, Bettongia penicillata and Monodelphis domestica, continue to grow after formation of the follicular antrum and, although the rate of growth slows in larger follicles, it continues into the period immediately before ovulation. The basis of this growth is unknown but may include accumulation of fluid and/or yolk-like material. Maturational changes within the cytoplasm of the oocyte also occur during the periovulatory period, including the accumulation of cortical granules. Differences in the structure of the zona pellucida are also evident between follicular and ovulated eggs; these differences are suggestive of compression of the zona pellucida, but may involve the addition of extra material. These findings suggest that the marsupial oocyte may not achieve complete cytoplasmic maturity until after ovulation; however, their relevance to fertilization and embryonic development require further investigation. Like those of eutherian mammals, marsupial oocytes undergo spontaneous nuclear maturation once removed from the follicular environment, suggesting a basically similar control system. It is not known whether the preovulatory cytoplasmic changes seen in marsupial oocytes matured in vivo also occur during maturation in vitro.

129 IN VITRO PRODUCTION OF EQUINE EMBRYOS FROM YOUNG AND OLD MARES BY INTRACYTOPLASMIC SPERM INJECTION

2008 ◽  
Vol 20 (1) ◽  
pp. 145 ◽  
Author(s):  
C. Herrera ◽  
M. Revora ◽  
L. Vivani ◽  
M. H. Miragaya ◽  
C. Quintans ◽  
...  
Keyword(s):  
Embryo Development ◽  
Intracytoplasmic Sperm Injection ◽  
Polar Body ◽  
Age Groups ◽  
Reproductive Status ◽  
Developmental Competence ◽  
Fisher Exact Test ◽  
In Vitro Production ◽  
The Mean

High merit mares obtain their utmost productive value at the same time their reproductive soundness diminishes. The aim of our study was to compare the developmental competence of equine oocytes from young and old mares after intracytoplasmic sperm injection (ICSI) and in vitro culture. Ovaries from young and old mares were obtained from a pool of slaughterhouse animals that have been previously selected by overall good body condition, reproductive status, and age. Young mares were 3 to 8 years old, and old mares were more than 15 years old. The age of all mares was determined by teeth observation and reproductive status by ultrasonography. Oocytes were obtained from ovaries 1 h postmortem by individual dissection of follicles between 10 and 25 mm and scraping of the follicle wall with a bone curette. Recovered oocytes were matured in vitro for 24–30 h, and all oocytes with an intact cytoplasm and a visible polar body were subject to ICSI and cultured for 7.5 days in SOFm. The maturation rate, cleavage, and embryo development rate and mean number of blastomeres at 7.5 days of culture were compared between oocytes and embryos from young and old mares. Maturation, cleavage, and developmental rates were analyzed by Chi Square and Fisher exact test, whereas the mean number of blastomeres at 7.5 days of culture was compared by one way ANOVA and t-test. A total number of 54 oocytes from young mares and 37 oocytes from old mares were obtained. There were no significant differences between the maturation, cleavage, or embryo development rates between young (79.63, 56.41, and 18.18%) and old (91.89, 63.33, and 15.79%) mares. In addition, the mean number of cells on embryos from each group did not differ significantly (57.75 v. 81; young v. old). Our preliminary results show that similar in vitro rates are achieved when oocytes from young or old mares are matured and fertilized in vitro by ICSI. This does not correlate with the reproductive senescence in old mares and the result obtained with other reproductive techniques. Further studies will determine if pregnancies are equally achieved using in vitro produced embryos from both age groups.

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