• 한국생물공학회:학술대회논문집
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• 2003.10a
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• pp.365-368
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• 2003

Hair follicles develop as a result of epithelial-mesenchymal interactions between epidermal keratinocytes and dermal cells. Moreover hair follicles constitute multiple cells that influence hair follicle development and cyclic activity. We isolated some cells using explantation and enzymatic digestion method from human scalp hair follicles. So we could culture some follicular cells, such as outer root sheath (ORS) cells, dermal papilla (DP) cells, dermal sheath (DS) cells, matrix cells and melanocyte.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.118-118
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• 2003

The aim of this study was to investigate the effect of glucose on embryonic development of mouse embryos. Two cell embryos were recovered from ICR female mice(3-4weeks) at 46~50 hrs after hCG 5 IU injection (mated just after hCG injection) and cultured in 50 $\mu m$ DMEM droplets supplemented with nothing (control: n=46), glucose 0.5mM (Group A; n=46) or glucose 3.15 mM(Group B; n=46) under mineral oil. All experimental media were supplemented with 20% human follicular fluid. Total blastocyst formation rates was lower (NS) in glucose groups (group A: 52.2% : B. 47.8%) than control group (60.9%). ZiB rates was the highest (P<0.05) in control (47.8%) than those in group A (21.7%) and B (28.3%). ZeB rates were the highest (NS) in group A (30.4%) than those in control (13.0%) and group B (19.6%). Blastocysts, cultured in group B (50.5), had the highest (NS) mean cell number compared with the others (control: 39.2 ; group A: (45.6). The ICM proportion (% ICM of total cells) in blastocysts cultured in group A (20.6%) was the highest (NS) than those of other tested groups (control: 15.2 ; group B: 13.9%). This study shows that a low dose of glucose added to culture medium increases the ICM proportion of blastocysts in mice.

• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.95-102
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• 1997

The studies on the carried out to investigate the effective concentration of cryoprotectant agents and sucrose by one-step straw method of bovine embryos. The follicular oocytes were cultured in TCM-199 medium containing 10 IU/ml PMSG(Sigma, USA), 10 IU/ml hCG(Sigma, USA), 1$\mu\textrm{g}$/ml $\beta$-estradiol(Sigma, USA) and 10% FCS for 24~48 hrs in incubator with 5% CO2 in air at 38.5$^{\circ}C$ and then matured oocytes were again cultured for 12~18 hrs with motile capacitated sperm by preincubation of heparin. The bovine embryos following dehydration by cryoprotective agents and various concentrations of sucrose were directly plunged into liquid nitrogen and thawed in 3$0^{\circ}C$ water. Survival and in vitro developmental rate was defined as devellpmental rate on in vitro culture or FDA-test. The results are smmarized as followes : 1. The high in vitro developmental rates of bovine frozen embryos after rapidly thawed in freezing medium was attained 2.0M glycerol, 2.0M DMSO, 1M or 2.0M propanediol. 2. The high in vitro developmental rates of bovine frozen embryos after rapidly thawed in freezing medium was obtained single cryoprotectant(6.7~17.4%) than mixed cryoprotectants(6.7~16.7%). 3. In vitro developmental rate of bovine embryos after rapid frozen-thawing in the freezing medium added 0.25M and 0.50M sucrose were higher cleavage rate than those of sucrose concentration of 0.75M and 1.00M. 4. The freezing methods on in vitro developemental rates of bovine embryos was attained slow freezing method(9.70~15.6%) higher than rapid freezing method(9.4~13.3%).

• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.103-109
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• 1997

The studies on the carried out to investigate to determine the optimum thawing temperature and equilibration time and 1 step straw method of frozen bovine embryos. The follicular oocytes were cultured in TCM-199 medium containing 10 IU/ml PMSG(Sigma, USA), 10 IU/ml hCG(Sigma, USA), 1 $\mu\textrm{g}$/ml $\beta$-estradiol(Sigma, USA) and 10% FCS for 24~48 hrs in incubator with 5% CO2 in air at 38.5$^{\circ}C$ and then matured oocytes were again cultured for 12~18 hrs with motile capacitated sperm by preincubation of heparin. The bovine embryos following dehydration by cryoprotective agents and various concentration of sucrose were directly plunged into liquid nitrogen and thawed in 3$0^{\circ}C$ water. Survival and in vitro developmental rate was defined as developmental rate on in vitro culture or FDA-test. The results are summarized as followes : 1. The equilibration time on in vitro developmental rates of bovine embryos was attained after short period of time(2.5~5 min.) in the freezing medium higher than long period of time (10~20 min.). 2. The temperature thawed at 3$0^{\circ}C$ after rapid freezing of bovine embryos resulted in a significantly higher in vitro developmental rate than did at 2$0^{\circ}C$ and 35$^{\circ}C$. 3. The thawing time on in vitro developmental rates of bovine embryos was attained after short period of time(1~5 min.) in the freezing mediuim higher than long period of time(10min.). 4. The in vitro developmental rates of bovine embryos after rapid frozen-thawing by 1 step straw method in the freezing medium added 1.5M, 2.0M glycerol, DMSO, propanediol and 0.25M, 0.50M, 0.75M, 1.00M sucrose were 12.5~19.4%, 10.0~15.6%, 9.1~13.8% and 6.7~12.9%, respectively.

• Journal of Veterinary Clinics
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• v.16 no.1
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• pp.138-144
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• 1999

To establish the differential diagnosis and treatment method in bovine ovarian cysts, specially ovarian cysts with corpus luteum, serum progesterone (P$_4$) concentration and ultrasonography for measuring the cystic wall thickness and diameter of cyst and corpus luteum were investigated from slaughtered cows with ovarian cysts. Ovarian cysts were classified 8 types by the number of cyst, cystic wall thickness and present of corpus luteum. Ovarian cysts with corpus luteum were 11 (13.6%) of 81 cows and ovarian cysts without corpus luteum were 70 (86.4%) cows. The incidence rates of 8 various types of ovarian cysts were as follows; 2Ba 33.3%, 2Aa 25.9% and 2Bb 14.8%, respectively. The incidence rates of ovarian cysts without corpus luteum were follicular cyst 59.2% and luteal cyst 27.2%. The cystic wall thickness were 2Ab 3.7 mm and 2Bb 3.5 mm, and the serum P4 concentrations were above 2.0 ng/ml in 1Aa, 1Ab, 1Ba, 2Ab and 2Bb, respectively. In ovarian cysts with corpus luteum, the correlation coefficients between corpus luteum area and serum P$_4$ concentration were 0.45. In ovarian cysts without corpus luteum, there was significantly positive correlations between cystic wall thickness and serum P$_4$ concentration ($r^2$= 0.54, p＜0.01). These results indicate that PGF$_2$$\alpha$ analogues can be choice for treatment of ovarian cysts with corpus luteum and above 3 mm the cystic wall thickness because serum P$_4$ concentrations were above 2.0 ng/ml in ovarian cysts with corpus luteum and thickened cystic wall. In conclusion, it is suggest that ultrasonography is useful diagnostic tool for diagnosis and selection of treatment remedy in cystic ovaries of bovine.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.9
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• pp.1204-1210
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• 2011

Objective of this study was to examine the effect of sodium nitroprusside (SNP), a nitric oxide (NO) donor on steroid synthesis, growth and apoptosis of buffalo granulosa cells (GCs) in vitro. Follicular fluid of antral follicles (3-5 mm diameter) was aspirated and GCs were cultured in 0 (control), $10^{-3}$, $10^{-5}$, $10^{-7}$, $10^{-9}\;M$ of SNP for 48 h. To evaluate whether this effect was reversible, GCs were cultured in presence of $10^{-5}\;M$ SNP+1.0 mM $N^{\omega}$-nitro-L-arginine methyl ester (L-NAME) a NO synthase (NOS) inhibitor or hemoglobin (Hb, $1.0{\mu}g$) as NO scavenger. Nitrate/nitrite concentration was evaluated by Griess method, progesterone and estradiol concentrations by RIA and apoptosis by TUNEL assay. SNP ($10^{-3}$, $10^{-5}$, $10^{-7}\;M$) significantly (p<0.05) inhibited estradiol and progesterone synthesis, growth, disorganized GCs aggregates and induced apoptosis in a dose dependent manner. However, $10^{-9}\;M$ SNP induced the progesterone synthesis and stimulated GCs to develop into a uniform monolayer. Combination of SNP $10^{-5}$ M+L-NAME strengthened the inhibitory effect while, SNP+Hb together reversed these inhibitory effects. In conclusion, SNP at greater concentrations ($10^{-3}$, $10^{-5}$ and $10^{-7}\;M$) has a cytotoxic effect and it may lead to cell death whereas, at a lower concentration ($10^{-9}\;M$) induced progesterone synthesis and growth of GCs. These findings have important implications that NOS derived NO are involved at physiological level during growth and development of buffalo GCs which regulates the steroidogenesis, growth and apoptosis.

• Development and Reproduction
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• v.1 no.1
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• pp.79-89
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• 1997

Recent studies have demonstrated that apoptotic cell death plays an important role in the mechanism underlying follicular atresia and luteolysis. However, the mechanisms responsible for initiating these processes have not been elucidated. In in vitro fertilization (IVF) programs, it is highly possible that continuous and repeated administration of FSH/hMG and GnRH agonists for the usage of ovarian hyperstimulation may induce apoptotic death of granulosa cells leading to atresia in the human ovarian follicles. The present study was performed to investigate whether FSH/hMG and GnRh agonists used for a longer period in controlled ovarian hyperstimulation has any effect on the apoptosis of granulosa-luteal (GL) cells obtained from hyperstimulated ovaries. To examine apoptotic cell death in the GL cells, cells were stained with acridie orange followed by observed in some of GL cells. Similar but distinct staining of apoptotic GL cells was observed when the cells were examined by using in situ TUNEL method. The healthy-looking cells with normal nuclear morphology were not stained, whereas cells with pyknotic nuclei or with apoptotic nuclei were intensively stained. After examining the ultrastructural features of GL cells by TEM, it was confirmed that the majority of cells seemed to have normal nuclei while GL cells undergoing apoptotic cel death were rarely found. The DNA extracted from GL cells showed a typical pattern of fragmentation following DNA electrophoretic analysis. We have confirmed that the apoptosis occurs in granulosa-luteal cells obtained from hyperstimulated ovaries. Technically, in situ apoptosis detection method is simple and reproducible and is well suited to identify the quality of oocytes retrieved from hyperstimulated ovaries.

• Clinical and Experimental Reproductive Medicine
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• v.16 no.1
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• pp.81-91
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• 1989

It has been suggested that the prognosis for fertility of the infertile patients with healed pelvic tuberculosis is very poor. Total 60 patients(77 cycles) with previous history of pelvic tuberculosis who underwent IVF-ET from January 1988 to March 1989 at SNUH were classified into three groups according to the principal histopathological lesions : tuberculous endometritis group(N=20, 28 cycles), tuberculous salpingitis group(N=32, 37 cycles) and pelvic peritoneal tuberculosis group(N=8, 12 cycles). To evaluate the effects of previous pelvic tuberculous lesions on ovarian follicular growth and development in controlled ovarian hyperstimulation for IVF-ET and its final outcome, serum E2 levels on the day of hCG administration(Day 0) and the day after hCG administration(Day +1), the number of ovarian follicles with mean diamete ${\geqq}$ 12 mm on Day 0, the number of oocytes retrieved by transvaginal aspiration, and pregnancy rate per cycle were measured and compared with control group(N=123, 161 cycles). There were no significant differences in cancellation rate during controlled ovarian hyperstimulation, total dosage of FSH and hMG administrated, menstrual cycle date(MCD) of hCG injection, serum E2 levels, the number of ovarian follicles with mean diameter ${\geqq}$ 15 mm, and the number of oocytes retrieved between pelvic tuberculosis group and control group. But in pelvic tuberculosis group, the number of ovarian follicles with mean diameter 12-14 mm, total number of ovarian follicles(${\geqq}$ 12 mm), and pregnancy rate per cycle were significantly decreased. These data suggest that previous pelvic tuberculous lesions have no significant adverse effects on the ovarian response to gonadotropin stimulation. IVF-ET proved to be an useful treatment modality for infertile patients with previous history of pelvic tuberculosis in spite of its relatively lowered pregnancy rate.

• Korean Journal of Ichthyology
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• v.29 no.4
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• pp.252-257
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• 2017

The oogenesis of the Microphysogobio yaluensis was investigated using light microscopy. Various developmental oocytes appeared in the ovary of the M. yaluensis. The oogenesis is largely divided into four stages: nuclear-chromatin stage, peri-nucleoli stage, vitellogenesis (yolk vesicle and yolk granule stages), and mature stage. The nuclear-chromatin is distributed in a large germinal vesicle as threads. The peri-nucleoli stage has many acidic nucleoli lining at the inner side of the nuclear membrane and an egg envelope just weakly starts. As the oogenesis gradually proceeds, they change to the vitellogenesis stage. The oocyte become to drastically increase and the marginal area of the ooplasm is covered with many vacuoles showing no negative reactions with hematoxylin and eosin staining, called the yolk vesicle stage. Many yolk vesicles-owned oocyte largely increase and as the development continues, its ooplasm is changed from the yolk vesicles to the yolk granules of eosinophilic. At the mature stage, lots of granules merged into a big yolk mass, acidophilic. Even at the mature stage, the egg envelope was still thin between the ooplasm and the follicular layer of the oocyte.

• Journal of Veterinary Clinics
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• v.5 no.1
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• pp.1-7
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• 1988

An observational study was conducted to evaluate the effect of anterior pituitary gonadtropin on super-ovulation in donors, embryo quality, development stage of embryo, transfer location, corpus luteum quality of recipient, ease of transfer and induced versus natural estrus on pregnancy rates. The data were collected from 451 non-surgical transfers of fresh and frozen bovine embryos. 1. The average number of CL, formed in ovaries which were stimulated with follicular stimulating hormone, and/or APG, were 11. 1${\pm}$1.21, 12.3${\pm}$1.84, respectively. The numbers of embryos recovered were 8.2${\pm}$1.35, 8.7${\pm}$ 1.39, and the numbers of transferable embryos were 6.4${\pm}$0.98. 6.6${\pm}$1.01. respectively. 2 There was no effect of spontaneous versus prostaglandin-induced estrus of recipients on pregnany rates of either fresh (61.5% vs. 59.5%) or previously frozen(54.5% vs. 58.5%) embryos. 3. There were significant differences(p<0.05) in pregnancy rates for embryo grade 1(63.6%), grade 2 (45.4%) and grade 3 (30.4％) in fresh group and for grade-1 (60.5%), grade-2 (47.8%) and grade-3 (18.1%) in previously frozen embryos. 4. The recipients transfered with embryo morulae were found to have a lower (p<0.05) pregnancy rates than those transfered with embryos of blastocyst in be th fresh and previously frozen embryos. 5. There was no significant difference between transfer locations in pregnancy rates using both fresh and frozen embryos. 6. Attempts to select recipients for high pregnancy rates on the basis of corpus luteum (CL) quality were performed. CL were classified into 3 quality grades, on the basis of size and prominence. Quality grades 1 and 2 group had higher (p<0.05) pregnancy rates than grade-3 group using both fresh and frozen embryos. 7 Ease of transfer was ranked to a scale of one to three on the basis of increasing difficulty (time required). Transfers ranted as ease score 1 and 2 had significantly higher (P<0.05) pregnancy pregnancy rates (45.8%, 66.6%) than ease score 3 (27.7%, 31.8%) using both fresh and frozen embryos respectively.