• Clinical and Experimental Reproductive Medicine
• /
• v.32 no.4
• /
• pp.315-324
• /
• 2005

Objectives: To compare the efficacy of GnRH antagonist multiple dose protocol (MDP) with that of GnRH agonist long protocol (LP) in controlled ovarian hyperstimulation for in vitro fertilization in patients with high basal FSH (follicle stimulating hormone) level or old age, a retrospective analysis was done. Methods: Two hundred ninety four infertile women (328 cycles) who were older than 41 years of age or had elevated basal FSH level (> 8.5 mIU/mL) were enrolled in this study. The patients had undergone IVF-ET after controlled ovarian hyperstimulation using GnRH antagonist multiple dose protocol (n=108, 118 cycles) or GnRH agonist long protocol (n=186, 210 cycles). The main outcome measurements were cycle cancellation rate, consumption of gonadotropins, the number of follicles recruited and total oocytes retrieved. The number of fertilized oocytes and transferred embryos, the clinical pregnancy rates, and the implantation rates were also reviewed. And enrolled patients were divided into three groups according to their age and basal FSH levels; Group A - those who were older than 41 years of age, Group B - those with elevated basal FSH level (> 8.5 mIU/mL) and Group C - those who were older than 41 years of age and with elevated basal FSH level (> 8.5 mIU/mL). Poor responders were classified as patients who had less than 4 retrieved oocytes, or those with $E_2$ level <500 pg/mL on the day of hCG injection or those who required more than 45 ampules of exogenous gonadotropin for stimulation. Results: The cancellation rate was lower in the GnRH antagonist group than in GnRH agonist group, but not statistically significant (6.8% vs. 9.5%, p=NS). The amount of used gonadotropins was significantly lower in GnRH antagonist group than in agonist group ($34.8{\pm}11.3$ ampules vs. $44.1{\pm}13.4$ ampules, p<0.001). The number of follicles > 14 mm in diameter was significantly higher in agonist group than in antagonist group ($6.7{\pm}4.6$ vs. $5.0{\pm}3.4$, p<0.01). But, there were no significant differences in clinical pregnancy rate (24.5% in antagonist group vs. 27.4% in agonist group, p=NS) and implantation rate (11.4% in antagonist group vs. 12.0% in agonist group, p=NS) between two groups. Mean number of retrieved oocytes was significantly higher in GnRH agonist LP group than in GnRH antagonist MDP group ($5.4{\pm}3.5$ vs. $6.6{\pm}5.0$, p<0.0001). But, the number of mature and fertilized oocytes, and the number of good quality (grade I and II) and transferred embryos were not different between two groups. In each group A, B, and C, the rate of poor response did not differ according to stimulation protocols. Conclusions: In conclusion, for infertile women expected poor ovarian response such as who are old age or has elevated basal FSH level, a protocol including a controlled ovarian hyperstimulation using GnRH antagonist appears at least as effective as that using a GnRH agonist, and may offer the advantage of reducing gonadotropin consumption and treatment period. However, much work remains to be done in optimizing the GnRH antagonist protocols and individualizing these to different cycle characteristics.

• Clinical and Experimental Reproductive Medicine
• /
• v.28 no.2
• /
• pp.95-103
• /
• 2001

연구목적: 본 연구는 생쥐 preantral follicles의 체외 배양 조건을 확립하고 이를 기초로 높은 체외 발달률 그리고 산자 생산률을 얻고자 하였다. 연구재료 및 방법 : Preantral follicles의 oocyte-granulosa cell complexes (OGCs)는 생후 12일된 FI ($C57BL{\times}CBA$)으로부터 난소를 적출하여 효소를 이용한 방법을 통해 획득하였다. 회수된 complexes는 10일 또는 12일 동안 체외 성장을 위해 Transwell-COL membrane insert로 옮겨졌고 5% FBS, 100 mIU/ml FSH, 100 mIU/ml hMC가 첨가된 ${\alpha}MEM$에서 배양되었다. 체외 성숙을 위해 1.5 IU/ml hCG가 첨가된 ${\alpha}MEM$에서 18 hrs 배양을 실시하였다. 그 후 M16에서 수정능력이 획득된 정자와 수정을 하여 4 hrs, 7 hts, 9 hrs 후에 10% FBS가 첨가된 modified M16 배양액에서 4일간 배양하거나 또는 bovine cumulus cell과 co-culture를 실시하였다. 그리고 형태적으로 정상적인 22개의 상실배와 포배를 2마리의 위임신 대리모 (ICR)의 자궁에 이식하여 산자 생산을 유도하였다. 결과: 1) OGCs 크기가 mouse preantral follicles의 핵 및 세포질 성숙에 미치는 영향을 조사하였을 때 $120{\sim}150{\mu}m$의 preantral follicles (MII: 33.0%, 난할률: 36.7%, 상실배 이상; 20.9%)은 핵 및 세포질 성숙에 있어서 $70{\sim}110{\mu}m$ (MII: 12.2%, 난할률: 10.2%, 상실배 이상: 4.8%)보다 더 높았다(p<0.001). 2)체외 성장기간의 연장이 mouse preantral follicles의 핵 및 세포질 성숙에 미치는 영향을 조사하였을 때 10일 (난할률: 38.2%)은 12일 (난할률: 20.0%)보다 난할률에서만 더 높았다 (p<0.01). 3) 체외 수정 시간의 연장이 mouse preantral follicle의 세포질 성숙에 미치는 영향을 조사하였을 때 9 hrs (난할룰 31.5%, 상실배 이상: 14.3%)은 4 hrs (난할률: 17.5%, 상실배 이상: 4.8%), 7 hrs (난할률: 20.4%, 상실배 이상: 6.1%) 보다 세포질성숙에 있어서 유의하게 높은 발달률을 나타냈다 (p<0.01). 4) 공배양이 mouse preantral follicle의 세포질성숙에 미치는 영향을 조사하였을 때 공배양 (상실배 이상: 17.4%)을 실시했을 때와 M16 (상실배 이상17.4%)에서 배양되었을 때는 차이가 없었다. 5)preantral follicle의 크기 ($120{\sim}150{\mu}m$), 체외 성장기간 (10일), 체외 수정 기간 (9시간), 배양 환경 (단지 medium만 존재)의 적절한 결과들을 종합하여 수행하였을 때 MII 성숙률, 난할률, 상실배 이상의 발달률은 30.2%, 39.3%, 22.5%이었고 총 22개의 상실배 및 포배를 2마리의 대리모에 이식했을 때 1마리가 임신했고 1마리의 산자를 생산했다. 결론: 따라서, 본 실험은 preantral follicle을 이용한 체외 배양 시스템이 생쥐 oocyte를 공급하는 또 다른 방법으로 효과적으로 이용될 수 있다는 것을 시사한다.

• Clinical and Experimental Reproductive Medicine
• /
• v.15 no.1
• /
• pp.40-52
• /
• 1988

Estradiol-17${\beta}$($E_2$) levels in the blood were estimated according to varying the time and amount of the administration of $Clomid^{(R)}$. $Clomid^{(R)}$ were administered on the 2nd, 3rd and 4th menstrual day corresponding to the recruitment period and on the 5th menstrual day corresponding to the selection period of the ovarian follicles, respectively. And $Clomid^{(R)}$ were administered 50 mg, 100 mg and 150 mg/day, repectively. The effects of the sequential HMG to $E_2$ levels in the blood were also estimated. The results were as following : 1. Blood $E_2$ levels according to the day and amount of administration of $Clomid^{(R)}$ were the highest in the group 3(D $2{\sim}6$, 150 mg/day, with HMG) and the lowest in the group 6(D $5{\sim]9$, 50 mg/day, without HMG). $E_2$ levels showed increasing tendency to 0 day. 2. In the cases of the administration of $Clomid^{(R)}$ during the $2nd{\sim}6th$ menstrual day, $E_2$ levels according to the amount were similar among groups and showed increasing tendency daily. 3. In the cases of administration of $Clomid^{(R)}$ during the $2nd{\sim}6th$ menstrual day, $E_2$ levels according to the sequential HMG independent of the amount of $Clomid^{(R)}$ were higher in the with HMG group than without HMG groups. 4. In the case of the administration of $Clomid^{(R)}$ during the $5th{\sim}9th$ menstrual day, $E_2$ levels according to the amount were the highest in the 100 mg/day group and the lowest in the 50mg/day group. 5. In the cases of administration of $Clomid^{(R)}$ independent of the amount during the 5th${\sim}$9th menstrual day, $E_2$ levels according to the sequential HMG were higher in the with HMG group than without HMG group. 6. $E_2$ levels according to the amount independent of the day of the administration of $Clomid^{(R)}$ were the highest in the 100 mg/day group and 150 mg/day, 50 mg/day group in low sequence. 7. $E_2$ levels according to the sequential HMG independent of the day and amount of the administration of $Clomid^{(R)}$ were higher in the with HMG group than the without HMG group. 8. $E_2$ levels according to the day of the administration of $Clomid^{(R)}$ independent of the amount of $Clomid^{(R)}$ and sequential HMG were the highest in the group D 2${\sim}$6 and the lowest in the group D 5${\sim}$9. According to the above results, there were higher $E_2$ levels in the group with sequential HMG than without HMG. Therefore, the hypothesis, postulated initially by the author, was not verified that sequential HMG would not affect the $E_2$ levels which were related to the process of the selection of the ovarian follicle in the connection with 'FSH window'. Because it may be the stimulation after the selection of later predominant follicle. And the highest level of $E_2$ was estimated in the $Clomid^{(R)}$ 150 mg/day group with sequential HMG on the 2nd${\sim}$6th day, and the higher levels were estimated in the 2nd${\sim}$6th day, 3rd${\sim}$7th day and 4th${\sim}$8th day groups than the 5th${\sim}$9th day group. The lower levels were estimated in the $Clomid^{(R)}$ 50 mg/day group without HMG than 100 mg/day and 150 mg/day on the 5th${\sim}$9th day. Therefore, further study will be needed that combines analyses of the E2 levels in the blood according to the various administration of $Clomid^{(R)}$ with or without sequential HMG and determination of the numbers and size of the ovarian follicles by ultrasonogram.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.33 no.6
• /
• pp.533-540
• /
• 2000

Gonadal development, gametogenesis, reproductive cycle, gonad index, flesh weight rate, and first sexual maturity of the turban shell, Lunella coronata coreensis were investigated by histological observation. The materials used were collected monthly from the rocky intertidal zone of Daehang-ri, Buan-gun, Jeollabuk-do, on the west coast of Korea, from July 1998 to June 1999. Sex of L coronata coreensis was separate. The gonad was widely located in the spirals of the visceral mass buried in the digestive gland. The ovary and testis were composed of a number of oogenic follicles and speymatogenic follicles, respectively. Monthly variations in the gonad index increased from March ($23.86{\pm}3.73$) when the water temperature increased and reached the maximun in July ($49.76{\pm}6.47$). And then, the gonad index sharply decreased in September ($15.58{\pm}2.33$). The flesh weight rate ranged from $25.2{\%}$ to $32.3{\%}$, and its variation showed a similar pattern to the gonad index. Individuals $<5.9 mm$ in shell height could not take part in reproduction in both sexes. Percentages of first sexual maturity of female and male specimens ranging from $7.0{\~}7.9 mm$ in shell heights were $84.6{\%}\;and\;91.7{\%}$, respectively, and $100{\%}$ in those over 8.0 mm in shell height in both sexes took part in reproduction. By studying the monthly changes of the morphological features and sizes of germ cells during gametogenesis in the gonad, the reproductive cycle of this species could be devided into five successive stages: early active (December to April), late active (January to July), ripe (May to August), spawning (July to September), and recovery (September to March). The spawning period of this species was once a year between July and September, and the main spawning occurred in July when the seawater temperature reached above $24.8^{\circ}C$. The fully ripe eggs were $150{\~}160\;{\mu}m$ in diameter.

• Journal of Veterinary Clinics
• /
• v.31 no.5
• /
• pp.394-402
• /
• 2014

The objective of this study was to investigate the effect of chronic unpredictable stress on the reproductive function and ovarian luteinizing hormone receptor (LHR) expression. 9-week-old C57BL/6 female mice were randomly divided into two groups: control group and stressed group. Mice have been stressed twice a day for 35 days with 12 different stressors which were randomly selected. The results demonstrate that there is significant increase in the anxiety-related behaviors (P < 0.05), decrease body weight gain rate (P < 0.01) and decrease in the average of litter size in stressed mice compared with control group (P < 0.01). Furthermore, the rate of primary, secondary and early antral follicles in stressed mice significantly decreased (P < 0.05), whereas that of atretic follicles significantly increased compared with control mice (P < 0.01). The immunohistochemical analysis revealed that reduced LHR expression in granulosa cells of follicle and luteal cells of corpus luteum in response to chronic unpredictable stress. The western blot analysis revealed significantly decrease in LHR expression in the stressed mice ovaries compared with the control (P < 0.05). These results suggest that ovarian LHR expression affected by chronic unpredictable stress and the modulated ovarian LHR is responsible for ovarian follicular maldevelopment and reproductive dysfunction.

• Pediatric Gastroenterology, Hepatology & Nutrition
• /
• v.4 no.1
• /
• pp.18-27
• /
• 2001

Purpose: The aims of this study are to investigate the effect of the eradication of H. pylori on histological change of gastric mucosa in children with H. pylori gastritis and to determine whether the histological grading by the Sydney system is valuable in predicting the effect of treatment. Methods: 1) Histological scores by the Sydney system and the endoscopic characteristics were assessed before and at least four weeks after anti-H. pylori therapy in 42 children with H. pylori gastritis. 2) In 32 children treated with omeprazole, amoxicillin and clarithromycin (OAC), pretreatment histological scores and endoscopic findings were compared between the eradicated and the noneradicated to evaluate their predictive value for the successful eradication. Results: 1) In the eradicated (27 cases), nodular gastritis significantly decreased from 89% to 63% (p<0.05). There was an significant improvement in the mean activity score from 2.06 before treatment to 0.24 after treatment (p<0.01). The mean inflammatory score also improved from 2.61 before treatment to 1.89 after treatment (p<0.05). Lymphoid follicles significantly decreased from 48% to 15% (p<0.05). Epithelial damage improved in all 4 cases. But in the noneradicated (15 cases), there was no significant change in the frequency of nodular gastritis, the mean activity score, the mean inflammatory score and the frequency of the lymphoid follicles. 2) In 32 children treated with OAC, there was a tendency that the higher was the pretreatment score of the bacterial density, the lower was the eradication rate of H. pylori (p=0.072). Conclusion: The loss of the polymorphonuclear cell infiltration is the most prominent histological change after successful eradication. There may be negative correlation of the grade of the bacterial density with the success rate of the anti-H. pylori therapy.

• Applied Microscopy
• /
• v.41 no.1
• /
• pp.47-53
• /
• 2011

Hair is an appendage of skin which protects the body from outer physical and chemical stimuli. Hair is generated from the hair follicle lying on a sunken basal layer of epidermis. Hair cycling, which regenerates hair follicles throughout the life time of the organism. Numerous kinds of factors which exist at the hair follicle have been reported to regulate hair cycling, Human growth hormone secreted from pituitary gland, initially demonstrated to accelerate organ's growth, has been reported to play a role in the biology of organ size determination. We investigated the effect of 6-histidines residues tagged at amino-terminus of human growth hormone using light and electronmicroscopic methods. Human growth hormone encapsulated in nano-liposome (LhGH) was used to find how LhGH affects hair follicle cycling of mouse (C57BL6/CrN). Distilled water as a negative control, 3% Minoxidil as a positive control, and LhGH were applied to mouse for weeks. LhGH increased the number of exposed hairs per given areas ($1mm^2$). This result was also confirmed using a different breed of mice which show natural hair loss in an old age (about 17 months after birth). When LhGH was applied for 3 weeks after natural hair loss, natural hair loss on these mice was prevented, However, the control group mice on which LhGH was not applied showed further hair loss. This result indicates that LhGH may stimulate hair cycling of mouse. In clusion, it is cleat that the LhGH increased the number of hair on mice and help the depilated skin to grow new hair follicles again.

• Korean Journal of Fisheries and Aquatic Sciences
• /
• v.17 no.5
• /
• pp.423-435
• /
• 1984

The reproductive cycle of the small filefish, Rudarius ercodes was investigated based on the annual variations of gonadosomatic index(GSI) and hepatosomatic index(HSI) by electronic and photic microscophy. The specimens used were collected at the coastal area of Benden island, Sizuokagen, Japan, from September 1982 to August 1983. GSI began to increase from March, starting season of longer daylength and higher water temperature, and reached the maximum value between June and August. It began to decrease from September with the lowest value appearing between November and February without any evident variation. The annual variations of HSI were not distinct in male filefish and were negatively related to GSI in female : HSI decreased in the summer season when the ovary was getting mature and reached the maximum in the winter season when the ovary was getting retrogressive. The ovary consisted of a pair of saccular structure with numerous ovarian sacs branched toward the median cavity. Oogonia divided and proliferated along the germinal epithelium of the ovarian sac. Young oocytes with basophile cytoplasm showed several scattering nucleoli along the nuclear membrane. when the oocytes growing to about 300 ${\mu}m$, nuclear membrane to disappear with nucleus migrating toward the animal pole. The regions of protoplasm were extremely confined within the animal hemisphere in which most of cytoplasms were filled with yolk materials and oil drops. After ovulation, residual follicles and growing oocytes remaining in the ovarian sacs degenerated. But perinucleatic young oocytes without follicles formed were not degenerated, and growing continuously still in the next year. Mitochondria and endoplasmic reticula in the cytoplasm remarkably increased with oocytes maturing and yolk accumulating. Those were considered to be functionally related to the yolk accumulation. Five or six layers of possible vitellogenin, oval-shaped disc structures with high electron density, appeared in the apex of follicular processes stretching to the microvilli pits of mature oocytes. Testis consisting of a pair of lobular structures in the right and left were united in the posterior seminal vesicle, Cortex of testis was composed of several seminiferous tubules, and medulla consisting of many sperm ducts connected with tubules. Steroid hormone-secreting cells with numerous endoplasmic reticula and large mitochondria of well developed cristae were recognized in the interstitial cells of the growing testis. Axial filament of spermatozoon invaginated deeply in the central cavity of the nucleus and the head formed U-shape with acrosome severely lacking, mitochondria formed large globular paranuclei at the posterior head, and microtubular axoneme of the tail represented 9+9+2 type. The annual reproductive cycles could be divided into five successive stages : growth(March to July), maturation(May to September), Spawning(mid May to early October) and resting stages(October to February). The spawning peak occurred from June to August.

• Clinical and Experimental Reproductive Medicine
• /
• v.26 no.2
• /
• pp.171-178
• /
• 1999

The follicular fluid (FF) of ovary contains various biological active products which affected on the growth of follicles and the fertilization of oocyte in physiological reproductive process of mammals. This study was designed to determine the effects of human FF on fertilization of oocyte and embryonal development in vitro culture. The FF was prepared as clear without blood contamination by needle aspiration from mature follicles of human at the time of oocytes retrieval for in vitro fertilization (IVF). As the medium for culture in vitro of embryonal cells, human tubal fluid (HTF) supplemented with follicular fluids at concentrations of 10%, 40% and pure FF were used. These effects were compared to control group of cultured embryos in HTF supplemented with 0.4% BSA (bovine serum albumin). For IVF, 64 eggs in control group, 67 eggs in 10% FF, 57 eggs in 40% FF and 64 eggs in pure FF were respectively allocated. And the rates of fertilization were almost similar in all groups as resulting 82.81% in control, 85.07% in 10% FF, 87.71% in 40% FF and 81.25% in pure FF. On the examination for embryonal cleavage from fertilized eggs, the rates of developing to 4 cell stage was similar in all groups, as results 98.11% in control, 98.27% in 10% FF and 98% in 40% FF but 78.84% in pure FF. And the rates of developing to 8-16 cell stage were significantly reduced as 44% in 40% FF and 44.23% in pure FF (p<0.05) compare to 71.69% in control media. As likewise, the rates of developing to morular stage were also significantly reduced to 36% (p<0.05) and 21.15% (p<0.01) respectively in 40% FF and pure FF. And the rates to blastocystic stage of embryo was lowest as 7.69% in pure FF (Table 1). The quality of embryonal cells on cleavage to the 8-16 cell stage was poorer, higher concentrations of FF. The rates of grade 1 in pure FF, as 23.07%, was lowest compare to those of other groups, in which the rates of grade 1 in control, 10% FF and 40% FF were 58.49%, 47.36% and 34% respectively. And on the contrary, the rate of grade 4 in pure FF was highest as 23.07%, while those were 5.66% in control, 8.77% in 10% FF and 20% in 40% FF (Table 2). On the viability of embryos, the rate of embryonal cell death was more rise, at the higher concentrations as well as longer exposure in the follicular fluid. At 48 hours after in vitro culture of embryos, the rate of survival embryos in pure FF was markedly lowered as 44.23%, compare to that of control (p<0.05). But there was not significant difference between the rates of survival embryos in each group beside the pure FF, which the rates were 77.35% in control, 70.17% in 10% FF and 60% in 40% FF respectively. And at 72 hours after in vitro culture, the rates of survival embryos were also significantly dropped to 21.15% in pure and 36% in 40% at concentration of FF compare to 62.26% in control (p<0.05, p<0.01). Finally, the rate of embryonal death at 96 hours after in vitro culture was highest as 82.69% in pure FF among all groups which those were 35.84 in control, 56.14% in 10% FF and 64% in 40% FF respectively (Fig. 1, 2, 3). In conclusion, this study suggests that the FF has no effects, in particular, to the in vitro fertilization of oocytes but exerted a bad effect to the cleavage, quality and viability of the embryonal cells during in vitro culture. However, the FF is harmful on embryonal development at conditions in higher concentration and especially on the embryos after $8{\sim}16$ cell stage.

• Journal of Embryo Transfer
• /
• v.26 no.1
• /
• pp.19-25
• /
• 2011

This study was carried out to evaluate the effect of early pregnant cow as donor for Ovum Pick-Up (OPU) derived oocyte aspiration and embryo production in Holstein heifers. Four non-pregnant and 2 pregnant Holstein heifers were used as donor and then carried out total 17 OPU session for 10 weeks (2 times per week). Recovered cumulus-oocyte-complexes (COCs) were classified into 4 grade by oocyte cytoplasm and cumulus cells and matured in vitro in TCM-199 supplemented with 10% FBS, 10 mg/ml FSH and 1 mg/ml estradiol in 5% $CO_2$ and over 99% humidity for 24 h. After 24 h co-incubation with post-thaw sperm, the presumed zygotes were cultured in CR1aa medium with 4 mg/ml BSA for 3 days and then changed CR1aa medium with 10% of FBS for another 3~4 days. The Mean number of aspirated follicles and collected oocytes in the early stage pregnant and non-pregnant heifers were $13.0{\pm}4.3$ and $10.6{\pm}3.9$, $5.4{\pm}3.4$ and $7.7{\pm}3.6$ per session, respectively. Rate of collected oocyte from aspirated follicles were 59.2% and 50.5%, respectively. The average number of good quality oocytes (Grade I and II) in the early stage pregnant and non-pregnant heifers was $3.7{\pm}2.7$ and $4.9{\pm}2.6$ (Mean${\pm}$SD). Cleavage and blastocyst developmental rates in Grade I and II were 22.2% and 25.5%, and then $1.7{\pm}0.9$ and $1.4{\pm}1.1$ blastocyst per session, respectively. In conclusion, OPU technology can be used in early stage pregnant and non-pregnant heifers without any problem and so applied OPU derived embryo production to maximize the ability of genetically valuable females.