• Journal of Embryo Transfer
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• v.19 no.3
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• pp.229-237
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• 2004

The main objective of this study was to examine the effects of serum and gonadotropins supplement during in vitro maturation(IVM) of bovine oocytes on nuclear maturation and embryo development, and we also examine the cell number. 1 . The first polar body(PB) extrusion rates of Korean native cow(KNC) oocytes matured in medium with FBS or gonadotropins were similar among treatment groups. The development rate to the blastocyst stage was significantly higher in the group of both supplement FBS and gonadotropins(26.0％) than in the group of non-supplement(9.9％) and gonadotropins (12.0％). The numbers of inner cell mass (ICM) and trophectoderm (TE) cells and total cell numbers of blastocysts were highest in the group of both supplement FBS and gonadotropins, and the number of ICM cells was increased by FBS supplementation (p<0.05). 2. The PB extrusion rates of KNC oocytes matured in medium with FBS in the different duration of IVM was significantly higher in the 0-18hr(63.1％) and in the 9-18hr(63.4％) group than in the 0-9hr.(37.4％) group (p<0.05). The embryo development rates did not differ among treatment groups. The numbers of TE cells and total cell numbers of blastocysts were similar among treatment groups, but the number of ICM cells of the 0-18h. group were significantly higher than the other treatment groups (p<0.05). The results indicate that although TCM199 alone can support bovine oocyte maturation and development to the blastocyst stage, a high quality of blastocysts can be produced from oocytes matured in medium containing serum and gonadotropins.

• Reproductive and Developmental Biology
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• v.34 no.1
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• pp.21-25
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• 2010

The purpose of this study was to investigate the effects of the collection time, co-culture and sperm penetration of canine oocytes on in vitro maturation and fertilization. The oocytes were cultured in TCM-199 media containing hormonal supplements (10% FCS, 10 IU/ml HCG, 10 IU/ml PMSG) at 5% $CO_2$, 95% air, $38^{\circ}C$. The in vitro maturation rate to MII stage of in vitro oocytes recovered from ovaries that collected at follicular, luteal and inactive phases of the reproductive phase for 44~72 hrs were 19.2%, 12.2%, and 6.0%, respectively. Follicular phases oocytes had a significantly higher in vitro maturation rate than oocytes collected at luteal and anestrus stage (p<0.05). The in vitro maturation rates to the MII stage of canine oocytes after 48 hrs of culture with glutathione, pyruvate, or glutathione + pyruvate were 12.5%, 10.7%, and 17.5%, respectively. This was higher than that in both alone or the combination of the two compared to the control group (19.0%). The sperm penetration rates of in vitro matured oocytes by fresh and frozen semen were 29/80 (36.3%) and 18/80 (22.5%), respectively. Although there are limited reports about canine oocytes co-culture and in vitro fertilization, our results on in vitro maturation is comparable to the results from other researches.

• The Korean Journal of Malacology
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• v.25 no.2
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• pp.105-112
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• 2009

We investigated the degree of obesity, histological development stages of gonads and sexual maturation induction rates of comb pen shell, Atrina pectinata, per the type of micro-algae supplied. Terms of maturation by singular or mixed supply of microalgae, it was found that maturation of the female was the quickest at 60.0% by the Tetraselmis tetrathele (Tet). experiment group followed by 57.1% by the Chlorella ellipsoidea (Chl). experiment group and 16.7% by the Phaeodactylum tricornutum (Pha). experiment group. However, there were no significant differences between Tet. experiment group and Chl. experiment group. As for the male, maturation was the quickest at 60.0% by the Tet. experiment group followed by 16.7% by the Chl. experiment group and 14.3% by the Pha. experiment group. In light of these results, Tet. is concluded to be a very useful feed organism in breeding the mother comb pen shells. Upon completion of the experiment, the sexual maturation induction rate for the female was found to be the highest at 82.0% in the Tet. experiment group followed by 72.0% by the Chl. experiment group, 64.0% by the Pha. experiment group and 58.0% by the mixed micro-algae experiment group. During the period of experiment, the survival rate was the highest at 94.4% by the mixed micro-algae experiment group followed by 90.0% by the Pha. experiment group, 83.1% by the Tet. experiment group and 78.8% by the Chl. experiment group.

• Fisheries and Aquatic Sciences
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• v.17 no.1
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• pp.115-121
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• 2014

Eyestalk ablation (ESA) is commonly used in aquaculture to stimulate ovarian maturation in crustaceans, and methyl farnesoate (MF) affects crustacean molting and reproduction. To investigate the physiological effects of ESA and MF treatments on the shrimp Litopenaeus vannamei, we compared the effects of single eyestalk removal and MF injections. The ESA group had the lowest survival rate (50%), and individuals in the $0.1{\mu}g$ and $1.0{\mu}g$ MF-treated groups had survival rates of 80 and 73.3%, respectively. Conversely, molting numbers were highest in the ESA group, and similar to those of the 1.0-${\mu}g$ MF group. To investigate shrimp growth, we measured body weight during the experimental period and found that individuals in the ESA and $1.0{\mu}g$ MF groups showed significant increases in body weight. Furthermore, to investigate the effects of ESA and MF treatments on gonadal maturation, the gonad somatic index (GSI) was calculated after the experiment. All treated groups (ESA and MF) had higher GSI values than the control group, but the ESA and $1.0{\mu}g$ MF groups were not significantly different. Using histological ovary analysis, we determined that all treated groups showed indications of the previtellogenic stage, unlike the control group (immature stage). These results suggest that the high-MF-concentration treatment produced effects similar to those of ESA with respect to molting number, growth, and ovarian maturation.

• Journal of Embryo Transfer
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• v.32 no.3
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• pp.111-122
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• 2017

The objective of this experiment was to explore the effects of Roscovitine (Rosco) prior to in vitro maturation (IVM) of immature pig oocyte. Brilliant cresyl blue test has been used to select the good quality of oocyte. Specifically, the effects of Rosco exposure on nuclear and cytoplasmic maturation, diameter, intracellular glutathione (GSH) and reactive oxygen species (ROS), and embryonic development after parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT), and gene expression levels in SCNT embryos have been measured. Cumulus oocyte complexes (COCs) have been exposed in $75{\mu}M$ of Rosco for 22 and 44 h. The COCs that were matured in the IVM for 44 h without Rosco used as control group. Diameter of matured porcine oocytes 44 h culture with Rosco was significantly lower than 22 h culture with Rosco and control groups. GSH was higher in control group than 22 h and 44 h with Rosco but reduction of ROS in 22 h than 44 h with Rosco. In PA, exposure with Rosco 44 h oocytes group has been significantly lower than 22 h and control group in rates of maturation, cleavage and blastocyst formation. Similarly, in SCNT embryos rates of maturation, cleavage and formation of blastocyst have been also significantly lower in 44 h Rosco treated group than other two groups. SCNT embryos treated with Rosco 22 h showed greater expression levels of POU5F1, DPPA2 and NDP52Il mRNA compared with other two groups. Our results demonstrate that Rosco treatment with 22 h prior to IVM improves the development competence of porcine oocyte.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.10
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• pp.1360-1366
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• 2001

The aims of this study were first to evaluate the effects of different levels (20, 40 and 100%) and sources (follicular size: large, >7 mm; medium, >5-7 mm; small, 3-5 mm) of porcine follicular fluid (pFF) on the in vitro maturation (IVM) of porcine oocytes, and the effects of fertilization treatments and different culture conditions on development of fertilized oocytes were also investigated. No differences in the maturation (63.6-76.6%) and cleavage (24.8-34.3%) rates were observed among the 20,40 and 100% pFF groups (p>0.05). The cleavage rates of oocytes cultured and fertilized in 40% and 100% pFF maturation media were significantly higher than those fertilized in m199-NBCS (51.0-61.2% vs. 12.8-31.8%. p<0.05), regardless of sources of the pFF. When oocytes were fertilized in m199-NBCS followed by culture in rabbit oviducts for 4 days, the cleavage rate in 40% pFF group was better than that in 100% pFF group (46.9% vs. 32.5%, p<0.05). Two oocytes recovered from the oviducts in the 40% pFF group developed to blastocysts after IVC. However, none developed to blastocysts when fertilized in the IVM medium after being transferred to rabbit oviducts. In conclusion, addition of pFF accompanied with gonadotropins (FSH, LH) in IVM medium enhanced maturation and cleavage rates of porcine oocytes. Direct addition of sperm suspension to IVM medium may be an alternative to simplify the fertilization procedures and to reduce the mechanical lesion during manipulation. Furthermore, rabbit oviducts provide a better environment for the in vitro fertilized oocyte developing to the morula and blastocyst stages.

• Korean Journal of Animal Reproduction
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• v.24 no.4
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• pp.375-383
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• 2000

The nuclear maturation and developmental competence of immature, oocytes collected from donors at various morphology of corpus luteum (CL) and fertilized in vitro was investigated by comparing the meiotic activity and the yields of embryos. Ovaries were divided and classified into 4 groups as the following criteria : Group 1 ; ovaries showed evidence of recent ovulation (corpus hemorragicum). Group 2 ; apex of CL was red or brown. Vasculization was limited to periphery of CL. Group 3 ; apex of CL was orange or tan. Vasculization was covered over apex of CL. Group 4 ; CL was light yellow to white and firm in texture and the vascular network on the surface of CL had disappeared. Modified TCM 199 was used for maturation in vitro of immature oocytes and development was induced by using TLP-PVA as a basic medium. When oocytes collected from each group of donors had been matured for 4, 14, and 24 hours in vitro, the proportion of oocytes reaching metaphase I and metaphase II were not different among oocytes from 4 group of ovaries. Mature metaphase II stage of oocytes in each group was first observed at 14 hours, whereas completion of maturation of. oocytes in each group was at 24 hours. Luteal morphology of ovaries had little effect on the proportion of embryos reached 2 cells and 8 cell stage. However, the proportion of embryos cleaved to morula and blastocyst stage was significantly higher in the oocytes obtained from group 1 and 3 than in the oocytes from group 2 and 4 (p＜0.05). This data suggest that reproductive status of the donor significantly influence the yield of in vitro embryos.

• Clinical and Experimental Reproductive Medicine
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• v.46 no.4
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• pp.166-172
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• 2019

Objective: In vitro maturation (IVM) of immature oocytes can be useful for some infertile patients. In IVM programs, the rates of embryo formation and pregnancy are low. Therefore, it is essential to recognize the main factors involved in regulating oocyte maturation in vitro. The purpose of this study was to investigate the effects of growth differentiation factor 9 (GDF9) and cumulus cell (CC) supplementation in IVM medium on the rates of embryo formation and viability of human blastocysts. Methods: A total of 80 germinal vesicle oocytes from stimulated cycles underwent an IVM program. The oocytes were divided into four groups, where group I consisted of IVM media only and served as the control, group II consisted of IVM+CCs, group III consisted of IVM+GDF9 (200 ng/mL), and group IV consisted of IVM+CCs+GDF9 (200 ng/mL). Intracytoplasmic sperm injection was performed on the IVM oocytes, and the cleavage embryos that were generated were vitrified. Following thawing, the embryos were cultured for 3 additional days, and the viability rates of the developed blastocysts were determined. Results: The maturation rate of the oocytes did not differ significantly across the four groups. The fertilization rate in group II was significantly higher than that in the control group (76.5% vs. 46.2%). Embryo formation was significantly more frequent in all experimental groups than in the control group, while blastocyst formation did not show significant differences in the three experimental groups compared to the control. The mean viability rates in groups II, III, and IV were 58.16%, 55.91%, and 55.95%, respectively, versus 37.78% in the control group (p< 0.05). Conclusion: Supplementation of IVM culture media with GDF9 and CCs enhanced the fertilization, embryo formation, and viability rates of blastocysts generated from vitrified cleavage embryos.

• Development and Reproduction
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• v.20 no.2
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• pp.127-133
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• 2016

Purine metabolism is known factor for nuclear maturation of oocytes through both follicle cells and oocyte itself. However, it is largely unknown the roles of purine metabolism in the oocyte competence for fertilization and early development. In this study, the effects of adenosine in oocyte competence for development were examined using adenosine and its synthetic inhibitors. Adenosine treatment from GV intact stage for 18 hr (fGV) caused of decrease the fertilization rate but of increase the cleavage rate compared from the other stage treatment groups. Hadacidin did not effect on fertilization rate but increased cleavage rate without stage specificity. Adenosine did not block the effects of hadacidin with the exception of fGV group. Inhibition of purine synthetic pathways the fertilization rate was decreased in the fGV and fGVB groups but increased in the fMII group. Exogenous adenosine caused of decrease fertilization rate in the fGVB group but increase in the fMII group. Cleavage rate was dramatically increased in the adenosine treatment with synthetic inhibitors. It means that the metabolism of purine has stage specific effects on fertilization and cleavage. Exogenous adenosine had only can improve oocyte developmental competence when it treated at GV intact stage. On the other hand, endogenous synthesis in all maturation stage cause of increase the cleavage rate without effects on fertilization. These data suggest that adenosine at GV stage as a paracrine fashion and inhibitions of endogenous adenosine in all stage improve oocyte developmental competence.

• Journal of Animal Science and Technology
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• v.57 no.8
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• pp.27.1-27.6
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• 2015

This study was designed to investigate the effects of ${\alpha}$-tocopherol and granulosa cells monolayer on nuclear maturation and cleavage rates of ovine cumulus-oocyte complexes (COCs). The COCs (n = 2814) were matured in maturation medium supplemented with various concentration of ${\alpha}$-tocopherol (0, 5, 10, $15{\mu}g/ml$), oocytes were incubated at $39^{\circ}C$ with 5 % $CO_2$ for 24 h in three culture systems: (a) maturation medium (MM; n = 884), (b) co-cultured with granulosa cells (CG; n = 982) and (c) co-cultured with granulosa cells and cells were further cultured in MM for 12 h (CG + 12hMM; n = 948). Our results showed that ${\alpha}$-tocopherol had no effect on GVBD and MII as compared to control group, but when ${\alpha}$-tocopherol added to maturation medium the rate of cleavage decreased. This indicates interaction of above mentioned factors in any of the treatments showed no significant differences on the rate of maturation and cleavage stages (MII, GVBD and cleavage) (p > 0.05). The oocytes co-cultured with granulosa cells for 24 h had beneficial effects on cleavage rate. The maximum MII and cleavage rates were achieved when oocytes had extra 12 h culture in the maturation medium without granulosa cells. Results also showed our modified co-culture system (CG + 12hMM), improved rates of MII and the cleavage in comparison with other studied maturation systems.