• Journal of Embryo Transfer
• /
• v.19 no.2
• /
• pp.165-172
• /
• 2004

This study was carried out to examine the effects of maturation media on in vitro maturation (IVM) of porcine immature oocytes, and on subsequent in vitro fertilization (IVF) and development (IVD). Cumulus-oocyte complexes (COCs) were collected from antral follicles of porcine ovaries collected from abattoir, and were matured in vitro in modified NCSU-37 (mNCSU-37), modified NCSU-23 (mNCSU-23), or TCM-199 supplemented with 10% porcine follicular fluid (pFF). Oocytes matured in vitro, were fertilized in vitro in modified Tris-buffered medium(mTBM) with the final motile sperm concentration of 1${\times}$105 sperm/mL, and subsequently putative embryos were developed in vitro in NCSU-23. The results are as follows. 1. In the result of IVM, the rate of germinal vesicle breakdown (GVBD) and of nuclear maturation were not significantly different among the media, though numeric value of them were slightly lower in TCM-199 than in mNCSU-37 or in mNCSU-23. 2. In the result of IVF, though the rate of sperm penetration was not significantly different among the maturation media, the percentage of oocytes with male pronucleus (MPN) of ones matured in mNCSU-37 (88.0%) was significantly higher than in TCM-199 (71.1%) (p<0.05). 3. In the result of IVD, the percentage of cleaved oocytes of ones matured in mNCSU-37 (52.3%) or in mNCSU-23 (53.7%) was significantly higher than in TCM-199 (43.1%) (p<0.05), but the rate of blastocysts at day 6 was not significantly different among the maturation media, though putative embryos from oocytes matured in mNCSU-37 or in mNCSU-23 were developed more than in TCM-199. These results suggested that mNCSU-37 or mNCSU-23 was more appropriate than TCM-199 as IVM medium for porcine immature oocytes.

• Development and Reproduction
• /
• v.7 no.1
• /
• pp.23-28
• /
• 2003

The seasonal reproductive cycle of the female honeycomb grouper, Epinephelus merra, inhabiting Chuuk was examined by histological observations of the ovaries. The gonadosomatic index (GSI) began to increase in February and peaked in March. Histological observations revealed many oocytes laden with yolk in the ovaries from March to April. From June to January, the ovaries were occupied by immature oocytes. These results suggest that the reproductive season of E. merra in Chuuk is from March through April.

• Clinical and Experimental Reproductive Medicine
• /
• v.41 no.2
• /
• pp.41-46
• /
• 2014

IVM refers to the maturation of immature oocytes in culture after their recovery from small antral follicles at the stage prior to selection and dominance. IVM requires little or no FSH in vivo and has been proposed as an alternative to conventional IVF, since it reduces the primary adverse effects caused by controlled ovarian stimulation, including the ovarian hyperstimulation syndrome. Moreover, IVM is a promising option for cases for which no standard protocol is suitable, such as FSH resistance, contraindications for ovarian stimulatory drugs, and the need for urgent fertility preservation. Recently, IVM has been used in women with regular cycles and normal ovaries. However, the pregnancy rate following IVM is suboptimal compared with that of conventional IVF, indicating that further studies to optimize the protocol and the culture conditions are warranted.

• Journal of Embryo Transfer
• /
• v.19 no.2
• /
• pp.185-189
• /
• 2004

The purpose of this study was to determine the possibility of in vitro maturation of tiger oocytes. Immature oocytes were recovered from a pair of ovaries. A total of 78 oocytes was collected, of which forty three were classified as good oocytes with compact cumulus cells and uniform cytoplasm. Forty three COCs were in vitro matured at $39^{\circ}C$, 5% CO2 in air atmosphere for 48 h in a IVM medium (TCM-199 supplement with 10% FBS, 0.6 mM cysteine, 0.2 mM pyruvic acid and 10 IU/mL HMG). Experiment I: the morphologic evaluation was conducted by measuring the diameter of oocytes with or without ZP, the thickness of ZP and the diameter of cytoplasm by microeyepiece at the same magnification (${\times}$100). Experiment II: the evaluation of meiotic development was conducted of the nuclear development stage of tiger oocytes. The results were summarized as follows: 1. The diameter of tiger oocytes $(176.5\pm6.1{\mu}m)$ with ZP was significantly (p<0.05) bigger than that of bovine oocytes $(150.7\pm4.9{\mu}m).$ The ZP thickness of tiger oocytes $(20.4\pm2.9{\mu}m)$ was significantly (p<0.05) bigger than that of bovine oocytes $(12.0\pm2.6{\mu}m;$ p<0.05). However, there was no significant difference in the diameter of cytoplasm (without ZP) between tiger $(122.1\pm9.7{\mu}m)$ and bovine oocytes $(118.7\pm7.5{\mu}m).$ 2. The rates of meiotic development of tiger oocytes were achieved GV (12.5 %) and MII (50.0%), respectively. These results indicated that tiger oocytes could be developed to MII in in vitro culture system.

• Development and Reproduction
• /
• v.8 no.1
• /
• pp.35-42
• /
• 2004

Oocyte maturation refers to the process that prophase I arrested germinal vesicle(GV) drives the progression of meiosis to metaphase II(MII) to have the capacity for fertilization and embryo development. To better understand the molecular mechanism(s) involved in oocyte maturation, we identified differentially expressed genes(DEGs) between GV and MII mouse oocytes using a new innovative annealing control primer (ACP) technology. Using 20 ACPs, we successfully cloned 32 DEGs between GV and Mll oocytes, and 26 out of these 32 DEGs were functionally known genes. Four genes including Pscd2 were GV-specific, 10 genes including PKD2 and CSN3 were highly expressed in GV oocytes(GV-selective), and 12 genes including Diva were highly expressed in MII oocytes (MII-selective). Ail of the genes identified in this study were first reported in the oocyte expression using ACP system and especially, we could characterize the existence of PKD-CSW signaling pathwayin the mouse oocytes. Results of the present study would provide insight for studying molecular mechanisms regulating oocyte maturation.

• Journal of Embryo Transfer
• /
• v.29 no.4
• /
• pp.361-368
• /
• 2014

This study was designed to evaluate the effect of bovine serum albumin (BSA) in a maturation medium on oocyte maturation and embryonic development in pigs. Immature pig oocytes were matured for 44 h in a medium supplemented with 0.4% (w/v) BSA, 0.1% (w/v) polyvinyl alcohol (PVA), or 10% (v/v) pig follicular fluid (PFF). After IVM, oocytes reached metaphase II stage were activated for parthenogenesis (PA) or used as cytoplasts for somatic cell nuclear transfer (SCNT). Nuclear maturation (89.5%, 90.7% and 91.3% for BSA, PVA and PFF, respectively) and intraoocyte glutathione contents (1.20, 1.16 and 1.00 pixels/oocyte for BSA, PVA and PFF, respectively) were not altered by the macromolecules added to maturation medium. IVM of oocytes in a medium containing BSA (21.4%) and PVA (20.7%) showed significantly lower blastocyst formation after PA than culture in medium with PFF (39.2%). After SCNT, oocytes matured in medium with BSA showed decreased embryonic development to the blastocyst stage (9.2%) compared to those matured in medium with PFF (28.9%), while 23.6% of SCNT oocytes matured in medium with PVA developed to the blastocyst stage. When the effect of BSA in a maturation medium during the first 22 h and the second 22 h of IVM in combination with PFF or PVA was examined, PVA-BSA showed a higher nuclear maturation (94.1%) than BSA-PFF (84.5%). However, there was no significant difference in the blastocyst formation among tested combinations (47.3, 52.2, 50.0, 44.4 and 49.0% for PFF-PFF, PFF-BSA, PVA-BSA, BSA-PVA and BSA-PFF, respectively). Our results demonstrate that BSA and PVA added to maturation medium can support oocyte maturation comparable to PFF-supplemented medium. However, maturation of oocytes in a BSA-containing medium decreases embryonic development after PA and SCNT when compared with the medium supplemented with PFF.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.42 no.1
• /
• pp.48-55
• /
• 2009

The gonad maturation and spawning of blue crab, Portunus trituberculatus (Miers, 1876) using samples caught by trammer net in the western coast of Korea from January to December. The gonadosomatic index (GSI) showed the highest value of 15.0 in June and lowest value of 1.29 in August. Hepatosomatic index (HSI) showed the lowest in the spawning season from June to July. Thereafter, from August the value started to increase to 5.77 in December, showing an adverse trend against GSI. In the histological studies of the female reproductive organ, the gonad filled with primary oocytes from January to February and secondary oocytes in May. The female gonads mature from June, and mature and immature groups could be distinguished from December to February or March. In August, female developed another cycle of gonad development, showing new primary oocytes in the gonads again. Some of female crabs had receptaculum filled with already active sperm cells in it from March to May. In the time of August to September, almost all the females had the sperm sacs in the receptaculum. From the result, the coupling period estimated from August to September. Total RNA and DNA values of the crabs were highest in May (2.51, $0.57{\mu}g/mg$). The DNA value remained unchanged except for the value in May ($0.09-0.13{\mu}g/mg$). while RNA value was lowest in January ($1.08{\mu}g/mg$). The RAN/DNA ratio was the lowest (6.23%) in May and highest (18.05%) in July. The value of correlations coefficient between body weight and fecundity of the crabs was higher that of between carapace width and fecundity.

• Journal of Embryo Transfer
• /
• v.29 no.2
• /
• pp.133-139
• /
• 2014

$K^+$ channels are involved in the regulation of a variety of physiological functions, including proliferation, apoptosis and differentiation, in mammalian cells. Our previous study demonstrated that the blockage of $K^+$ channels inhibits mouse early embryonic development. This study was designed to identify the effect of $K^+$ channels during bovine embryonic development. $K^+$ channel blockers (tetraethylammonium (TEA), $BaCl_2$, quinine, ruthenium red and fluoxetine) were added to the culture medium during in vitro fertilization (IVF) for 6 h to first identify the short-term effect of these chemicals. Among $K^+$ channel blockers, fluoxetine, which is used as a selective serotonin reuptake inhibitor, significantly increased the blastocyst formation rate by approximately 6% when compared to control. During the in vitro maturation (IVM) of immature oocytes and the in vitro culture (IVC) of embryos, the oocytes and embryos were exposed to fluoxetine for either a short-term (6 h) or a long-term (24 h) to compare the embryonic development in response to exposure time. The 6 h exposure to fluoxetine during IVM did not affect the blastocyst formation rate, but the rate of blastocyst formation was reduced after the 24 h exposure. On the other hand, embryonic development increased approximately 10% in both groups of embryos exposed to fluoxetine for 6 and 24 h during IVC. Taken together, fluoxetine treatment during IVF and IVC, but not IVM, enhances bovine embryonic development. These results suggest that fluoxetine-modulated signals in oocytes and embryos could be an important factor towards enhancing bovine embryonic development.

• Journal of Embryo Transfer
• /
• v.32 no.3
• /
• pp.123-130
• /
• 2017

This study was conducted to investigate the effects of alpha-lipoic acid (aLA) as an antioxidant that decrease the reactive oxygen species (ROS) in bovine embryonic development. Slaughterhouse derived bovine immature oocytes were collected and 4 different concentrations (0, 5, 10 and 20 mM) of aLA was supplemented in bovine in vitro maturation (IVM) medium. After 20 hrs of IVM, maturation rates, levels of ROS and glutathione (GSH), and further embryonic development after parthenogenetic activation (PA) and in vitro fertilization (IVF) was investigated according to aLA concentrations. Maturation rate was significantly higher in 10 mM group than other groups (80.5% vs. 62.9, 73.9, 64.2%; P<0.05). In the levels of ROS and GSH in matured oocytes as an indicator of oocyte quality, significantly better results were shown in 5 and 10 mM groups compared with other 2 groups. After IVM, significantly higher rates of blastocyst formation were shown in 10 mM groups in both of PA (27.9% vs. 18.8, 22.3, 14.2%; P<0.05) and IVF (32.6% vs. 23.9, 27.3, 16.2%; P<0.05) embryos. In addition, significantly more cell total cell number and higher inner cell mass ratio in 10 mM PA and IVP blastocysts showed developmental competence in 10 uM groups. Therefore, based on the entire data from this study, using $10{\mu}M$ of aLA confirmed to be the optimal concentration for bovine oocyte maturation and embryonic development.

• Journal of Embryo Transfer
• /
• v.24 no.3
• /
• pp.213-220
• /
• 2009

The objective of this study was to examine the effect of eCG and various concentrations (20, 40, and 80 ${\mu}g/ml$) of porcine FSH on nuclear maturation and intracellular glutathione (GSH) level of oocytes, and embryonic development after parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT) in pigs. Immature pig oocytes were matured in TCM-199 supplemented with porcine follicular fluid, cysteine, pyruvate, EGF, insulin, and hormones (10 IU/ml hCG and 10 IU/ml eCG or $20{\sim}80{\mu}g/ml$ FSH) for the first 22 h and then further cultured in hormone-tree medium for an additional 22 h. Nuclear maturation of oocytes ($85{\sim}89%$) was not influencem foreCG and various concentrations FSH. Embryonic development to the cleavage stage ($86{\sim}94%$) and mean number of cells in blastocyst ($33{\sim}37$ cells) after PA were not altered but blastocyst formation e-treignificaddlor(p<0.05) improvem forthe supplementation eith 80 ${\mu}g/ml$ FSHr(64%) compared to 47%, io8%, iand 47% in oocytes that were treated with eCG, 20,i and 40 ${\mu}g/ml$ FSH,i numectivelo. In SCNT, fusion ($78{\sim}83%$) of cell-cytoplast couplets and siosequent embryo cleavage ($82{\sim}88%$) were not influencem fordifferent gonadotropins but blastocyst formation tended to increase forthe supplementation eith 80 ${\mu}g/ml$ FSHr(25% vs. $11{\sim}18%$). Our nuults demonstrated that oocyte maturation and embryonic development after PA and SCNT e-frinfluencem fortype of gcem fortype of gits concentration. In this study, supplementation of maturation medium eith 80 ${\mu}g/ml$ FSHrimproved preimplantation development of PA and SCNT pig embryos, probably by increasing intracellular GSH concentration of matured oocytes.