생쥐배의 생체외 배양에서 일어나는 2-Cell Block에 관한 연구

우복희,유한기,왕영미,배인하 대한산부인과학회 1992 Obstetrics & Gynecology Science Vol.35 No.12

본 연구에서 인산, 포도당, 비타민 및 EDTA(ethylenediaminetetraacetic acid) 등을 수정란 및 초기 2-세포배 배양액에서 제거 혹은 첨가함으로써 다음과 같은 결론을 얻었다. 1. 배양액의 성분중 인산성분은 2-세포 발생에 억제적으로 작용하여 2-cell block의 원인이 되어 33-38%의 극복 효과가 있었다. 2. 배양액 성분중 포도당은 햄스터의 경우와 달라 억제적인 작용을 하지 않으며 촉진의 효과도 없었다. 3. 비타민중 panthotheic acid 및 inositol 은 2-cell block을 극복시키는 작용은 없으나 일단 극복된 배의 발생을 촉진시켜 배발생속도를 빠르게 하였다. 4. Fe++(Fe2(NO3))성분은 Ca++-inhibitor로 작용하며 8-세포기에 일어나는 compaction과정을 방해하는 것으로 추정되어 상실기 이상의 발생을 억제시켰다. 5. EDTA 성분 추가로 2-cell block이 67% 극복되었다. EDTA의 chelating 효과로 인한 Ca++의 기능을 조절함으로써 야기되는 것으로 추정되어 이에 대한 연구가 절실히 요망되고 있다. In most mammals, there is a block stage when fertilized ova(zygotes) are cultured and these zygotes can not develop to the blastocyst in vitro system except when cultured in ampulla portion of the oviduct. In mouse, this block stage is in a early 2-cell embryo. Its reason is that in vitro culture system can not mimic the in vivo system and can not satisfy all the conditions of dynamic environment and all the physiological conditions of embryos development of the in vivo system. Most basic reason of this 2-cell block is the inactivation of cdc genes of the nuclei, which is caused by unknown factor(s) of the in vitro system. In the present study, it is aimed to investigate the causing factor of mouse 2-cell block by the addition or removal of the components of the culture medium. Female mice of ICR strain (4-6 weeks old) were used. Superovulation was induced with PMSG 51U and HCG 51U at 48-hour interval. Then, immediately the female mice were mated with fertile males. Zygotes or early 2-cell embryos were collected by flushing the oviducts 18-30 hours after HCG injection. M16 was the basic medium used in the present study from which phosphate or glucose was omitted and vitamins (inositol, panthotheic acid), Fe+++, insulin, amino acids(glutamine, methionine, phenylalanine and proline) and EDTA were added depending on the experimental groups. In these different medium components, zygotes or 2-cell embryos were cultured for 24, 72 and 96 hours. Results as followings were obtained from this study. 1. Phosphate of the medium was strongly ingibitory to 2-cell embryo development, and 33-38% of 2-cell block were overcome by emission of phosphate from the cultrue medium. 2. Glucose seems to have neutral effect differenty from those of hamster. It did not show any inhibition or stimulation of 2-cell embryo development. 3. Panthotheic acid and inositol show some stimulatory effect of development of the 2-cell embryos which were already overcome by the emission of phosphate from the culture medium. 4. Fe+++ seems to be inhibitory to the compaction of 8-cell embryos by replacing CA++ of the culture medium. In the presence of Fe+++, all the embryos could not develop to 8-cell stage. 5. Overcoming effect of EDTA of 2-cell block was 67%. This seems to be caused by regulation of Ca++ function which is the major divalent lon in the culture dedium. It is further needed for the study of Ca++ function in mouse 2-cell block.

생쥐 배아의 냉동 및 해빙방법에 따른 발생률에 대한 비교 연구

우복희,손영수,김향미,장병우 대한산부인과학회 1996 Obstetrics & Gynecology Science Vol.39 No.2

ICR 계통의 제 2대 잡종 생쥐(CBA*C75BL)를 과배란 유도하여 2-세포기, 4-세포기 생쥐배아를 회수하였다. 회수된 배아를 propanediol과 DMSO를 각각 동결보존제로 사용하여 완만냉동-급속해빙, 급속냉동-급속해빙, 초급속냉동-급속해빙방법으로 냉동 보존 후 해빙을 실시하여 아래와 같은 결론을 얻었다. 1. Propanediol을 이용하여 2-세포기 배아를 냉동-해빙한 결과 완만냉동 방법에서 4-세포기 이상으로의 성장발달율이 84.0%, 부화율이 43.3%로 유의하게 높았다. 2. DMSO를 이용한 2-세포기 배아의 냉동-해빙의 결과는 급속냉동 방법에서 배아의 성장발달율이 84.2%, 부화율이 30.8%로 유의하게 높았다. 3. Propanediol을 이용한 4-세포기 배아를 냉동-해빙한 결과는 완만냉동 방법에서 배아의 성장발달율이 74.0%, 부화율은 54.8%로 유의하게 높았다. 4. DMSO를 이용한 4-세포기 배아의 냉동-해빙의 결과 급속냉동 방법에서 배아의 성장율은 84.4%, 부화율은 43.8%으로 유의하게 높았다. 따라서 냉동-해빙의 방법은 각각의 냉동보존제와 배아의 세포기에 따라 다르지만 propanediol을 사용한 경우 2-세포기와 4-세포기 배아 모두 완만냉동-급속해빙 방법이 가장 적합한 방법이며 DMSO를 사용한 경우는 급속냉동-급속해빙 방법이 가장 적절한 방법으로 생각된다. Obsectives: In order to investigate the best cooling-thawing method, according to cell stage and cryoprotectants. Material & Method: 2-cell and 4-cell stage of mouse embryos were cryopreved by slow, rapid and ultrarapid cooling method using the Propanediol and Dimethyl Sulfoxide, each other. After then, the cyopreserved embryos were thawed by rapid thawing method. Teh recovered embryos, developed embryos were observed and compared among three cooling methods. Result: The results were as followes: 1. When 2-cell mouse embryos with Propanediol were cyropserved by Slow cooling-Rapid thawing method, the cleavage rate(84.0%) and the hatching rate (43.3%) are significantly higher than the other methods. 2. When 2-cell mouse embryos with DMSO were cryopreserved by Rapid cooling-Rapid thawing method, the cleavage rate(84.2%) and the hatching rate (54.8%) are significantly higher than the other methods. 3. When 4-cell mouse embryos with Propanediol were cryopreserved by Slow cooling-Rapid thawing method, the cleavage rate (74.0%) and the hatching rate (54.8%) are significantly higher than the other methods. 4. When 4-cell mouseembryos with DMSO were cryopreserved by Rapid cooling-Rapid thawing method, the cleavage rate(84.4%) and the hatching rate (43.8%) are significantly higher than the other methods. Conclusion : We concluded that the mostly adequate cryopreserved method of 2-cell and 4-cell stage of mouse embryos using propanediol is the slow coolin-rapid thawing method and using DMSO is rapid cooling-rapid thawing method.

노령산모의 최근 임신 및 출산추세에 관한 연구

우복희,손영수,김승철,김향미,장병우,윤수정 대한산부인과학회 1996 Obstetrics & Gynecology Science Vol.39 No.3

We have compared the obstetric performances of elderly gravida with those of young gravida and assessed the recent tendency of pregnancy and delivery in elderly gravida. The data presented are based on 421 cases of mothers with advanced age, 35 years or more among a total of 9351 deliveries ween in Ewha Womans University Hospital in the past 4 years. (from 1991 to 1994). As the control subject 309 cases of age less 35 years were randomly selected from the repective period. The results were as follows. 1. The rate of elderly gravida was increased from 2.0% in 1991 to 6.1% in 1994. 2. The age distrivution of elderly gravida was from 35 years to 45 years. 3. The parity of elderly gravida was that primigravida was 22.8% and mulitgravida was 77.2%. 4. Vertex presentation was seen in 383 cases(91.0%), breech presentation in 35 cases(8.3%), and transeverse lie in 3 cases(0.7%). The frequency of abnormal presentation was significantly higher in the elderly gravida group than control group. 5. The preterm pregnancy was seen in 6.9% compared with 7.8% in control group. The postterm pregnancy was observed in 1.4% compared with 3.6% in control group. 6. In the elderly gravida, the rate of casarean section was ween in 58.2% compared with 35.6% in control group. Among the reasons for cesarean section, the highest incidence was previous C/S(54.3%). Other indications were elderly primigravida, CPD and abnormal presentation in decreasing order. 7. Concerning the prenatal complication, the incidence the hypertesive disorder was seen in 5.9% compared with 4.5% in control group. 8. Incidence of low birth weight (4,000 gm) was 13.1%, compared with 1.9% in control group. The rates of low apgar score, perinatal mortality, neonatal death and congenital anomaly were not significantly different from control group. 9. The male-to-female sex ratio was 1.25: 1, compared with 1: 1.9 in control group. The delivery rate of male baby was significantly higher in the elderly group than in control group. In conclusion, cesarean section rate is increased because doctors prefer cesarean section for elderly primigravida. So, if operation is performed only in selected patients who have the proper indicatins, we can decrease the rate of cesarean section. Prenatal complications and perinatal mortality rate show no significant differences between elderly gravida and younger gravida. Thus, provided that careful antenatal and intrapartal care is taken for elderly gravida, women have no reason to fear delayed childbirth. Since artificial abortion adversely affects maternal health and disproportionately high male-to-female sex ratio is expected to cause numerous social problems in the future, sex determination and artificial abortion by educating the patients to abandon the idea of preferential of male baby must be prevented.

Pentoxifylline이 생쥐 정상정자의 체외수정 및 초기배아의 성장발달에 미치는 영향에 관한 연구

우복희,유한기,안정자,손영수,김향미 대한산부인과학회 1997 Obstetrics & Gynecology Science Vol.40 No.1

Object: This study was carried out to investigate the effect of pentoxifylline on in vitro fertillization and developmen of preimplantation stage of mouse embryos. Matrial and Methods:F1 hybrid mice was superovulated with PMSG/hCG and mouse oocytes were recruited. After the normal sperms were incubated with PTX before in vitro fertilization, it was observed whether the fertilization and embryo development was affected or not by the sperm preparation(washing, dilution and no washing or no dilution). And after 1-cell and 2-cell stage of mouse embryos were incubated with PTX, the development to hatching blastocyst was also observed. Results: When in vitro fertilization was revealed by using the washed normal sperms after 0, 3.6 and 7.2 mM PTX incubation, the fertilization rates were 92.5%, 48.8%, 36.8%, respectively. So 3.6 and 7.2 mM groups presented significantly low fertilizatin rate, but the development rates were 93.9%, 85.0%, 95.2%, respectively. Therefore, there were no significant difference between each group. When in vitro fertilization was revealed by using the diluted normal sperms after 0, 3.6, and 7.2 mM PTX incubation, the fertilization rates were 58.6%, 5.4%, 9.4%, respectively. So 3.6 and 7.2 mM groups presented significantly low fertilization rate. The developmental rates were 88.2%, 100%, 100%. And there were no significant difference between each group. When in vitro fertilization was revealed by using the not washed and not diluted normal sperms after 0, 3.6 and 7.2 mM PTX incubation, the fertilizatin rates were 61.2%, 5.7%, 3.8%, respectively. 3.6 and 7.2 mM group presented significantly low fertilization rate. The development rates were 73.3%, 0%, 0%, respectively. So 3.6, 7.2 mM group presented significantly low developmental rate. After 1-cell stage of mouse embryos were incubated in 0, 5, 10, 50 μM of PTX, the development rates were not significantly different among them. After 2-cell stage of mouse embryos were incubated in 0, 5, 10, 50 μM of PTX, the development rates were not significantly different among them. Conclusion: In conclusion, when PTX is used in in vitro fertilization program with normal sperms, it may affect the fertilization and embryo development in high concentration. And if PTX concentration is very low, the developmental rate would not be affected. So PTX must not be used to normal sperms and where use of PTX is indicated, it is recommended that remainder PTX must be removed as completely as possible.

말기 정상단태임신에 있어서 모체혈청 알파 - 태아단백농도에 영향을 미치는 요인에 관한 연구

우복희,안정자,김승철,박미혜 대한산부인과학회 1997 Obstetrics & Gynecology Science Vol.40 No.1

This study was purosed to investigate maternal and fetal clinical parameters effecting on maternal serum alpha-fetoprotein(MSAFP) levels at late normal singleton pregnancies. The subjects of this study were 171 pregnant women with gestational age of 36 to 42 weeks, and didn`t have any medical or gynecologic diseases. They delivered fetuses within three days after blood test of MSAFP. MSAFP levels were measured by enzyme- immunoassay. The analysed clinical parameters included fetal sex, fetal weight, gestatioal age, maternal age, gravidity, parity, maternal weight and maternal total weight gain during pregnancy. The results were as follows: At uncomplicated late pregnancies, 1. Male fetus bearers had higher MSAFP(mean: 191.8 ng/ml, SD: 80.8 ng/ml, n=79) than female-fetus bearers(mean 153.6 ng/ml, SD 73.0 ng/ml, n=92)(p=0.0014). 2. Multiparas had higher MSAFP(mean: 192.7 ng/ml, SD: 77.9 ng/ml, n=80) than nulliparas(mean: 152.3 ng/ml, SD 75.1 ng/ml, n=91)(p=0.0007). 3. MSAFP did not have correlation with maternal age, maternal weight, maternal total wight gain during pregnancy, gestational age, fetal weight. According to the above results, fetal sex and parity are the factors that influences MSAFP levels at uncomplicated late pregnancies. So MSAFP values should be interpreted with cautions.

조기진통 임신부의 양수내 Interleukin-6 및 양막 융모막염의 관련성에 관한 연구

우복희,안정자,김영주,박미혜,정화순 대한산부인과학회 1997 Obstetrics & Gynecology Science Vol.40 No.2

Objective: Our purpose was to determine whether amniotic amniotic fluid concentrations of interleukin-6 are valuable in the prediction of histologic chorioamnionitis and in the prediction of perinatal morbidity. Study Design: The relation among amniotic fluid imterleukin-6, histologic chorioamnion- itis, and perinatal outcome was examined in patients with preterm labor and intact membranes. Interleukin-6 was determined by ELISA method. Results: Patients with histologic chorioamnionitis had significantly higher amniotic fluld interleukin-6 concentrations than patient without histologic chrioamnionitis. Also, patient with histologic chorioamnioitis had significantly higher perinatal morbidity rate than patient without histologic chorioamnionitis. An ammiotic fluld interlerkin-6 concentration had a sensitivity of 75% and a specificity of 62.5% in the diagnosis of preterm delivery. Conclusion: Amniotic fluld interleukin-6 is a sensitive test for the prediction of histologic chorioamnionitis and of perinatal morbidity.

생쥐배아의 체외성장에서 Albumin 첨가에 의한 수질극복효과에 대한 연구

우복희,유한기,손영수,이미자,김향미 대한산부인과학회 1996 Obstetrics & Gynecology Science Vol.39 No.1

체외수정시술 프로그램의 정도관리의 방법으로써 생쥐 제 1대 잡종(C57BL*CBA)의 1-세포기 배아와 2-세포기 배아를 이용하여 배양액에 사용되는 수질에 따른 배아의 성장 및 발생 및 소혈청 알부민의 첨가에 의한 배아의 성장발육의 효과를 생물학적 검증을 시행후 비교분석하여 다음과 같은 결과를 얻었다. 1. 1-세포기 생쥐 배아는 단순 배양액과 복합 배양액에서 수질에 관계없이 체외 성장발육을 하였으며, 소혈청 알부민의 첨가로 체외 성장발육이 유의하게 촉진되었다. 2. 2-세포기 생쥐배아는 단순 배양액에서는 정제되지 않은 물에 민감하며, 고순도의 물에 비해 유의하게 저조한 체외 성장발육을 보였으나 이는 소혈청 알부민의 첨가로 극복되는 것으로 나타났다. 복합 배양액에서는 체외 성장 발육에 있어 수질에 따른 차이를 보이지 않았으나 소혈청 알부민의 첨가로 체외 성장 발육이 유의하게 촉진되었다. 이상의 결과로 미루어 보아 소혈청 알부민의 첨가로 인한 초기 생쥐배아의 체외 성장발육은 배아의 세포기나 배양액의 종류에 관계없이 촉진되며, 수질에 대한 영향은 2-세포기 배아에서 민감하게 나타내며, 또한 이는 단순 배양액에서 현저하게 나타나지만 이는 소혈청 알부민의 첨가로 극복되는 것으로 사료된다. This study was carried out to investigate the sensitivity of preimplantation mouse embryo to the influence of water quality and whether the influence of poor water quality is overcome by the addition of SBA. F1 hybrid mice was superovulated with PMSG/hCG and timely mated. 1-cell stage and 2-cell stage mouse embryos were recruited. The embryos were cultivated in m=KRB and Ham`s F-10 media; both media were produced by using highly purified water adn tap water and each media was divided into two groups: 0.4% BSA was added to one group and nothing was added to other group. 1-cell stage mouse embryo grew and developed in vitro in simple media and complex media regarless of water quality. And its in vitro growth and development was significantly stimulated by the addition of BSA. In simple media, in vitro growth and development of 2-cell mouse embryo was low in poor water quality but the retarded growth and development was overcome by the addition of BSA. In complex media, its in vitro growth and development was not changed by water quality but it was significantly stimulated by the addition of BSA. Conclusively, in vitro growth and development of preimplantation mouse embryo is stimulated by the addition of BSA regardless of embryonic cell stage or media type. 2-cell stage mouse embryo appears to be sensitive to the influence of poor quality, especially in simple media, but it would be overcome by the addition of BSA.

정상아와 발육지연 태아에 있어서의 태아 중뇌동맥의 혈류속도파형의 비교

우복희,안정자,김영주,정혜원,김승철,이경순,문혜성 대한산부인과학회 1994 Obstetrics & Gynecology Science Vol.37 No.8

이화대학병원 산부인과를 내원한 임신 24주에서 40주까지의 정상 임신부의 태아 중뇌동맥에서 도플러 초음파를 시행하고 임신결과가 정상으로 확인된 70명의 분석결과 중뇌동맥의 혈류속도 파형은 이완기에도 지속적인 혈류를 나타내었고 임신이 진행됨에 따라 뇌혈관망 저항의 감소로 PI의 감소가 오게되며 따라서 혈류속도는 증가함을 알 수 있었다. 또 임신결과가 10 percentile이하의 저체중아로 나타난 경우에는 정상 임신에 비하여 brain sparing effect의 개념에 입각하여 더욱 낮은 PI 즉 뇌혈관 망 저항의 심한 감소 및 혈류속도의 증가를 보여주었다. 본 연구결과 정상 임신과 저체중아의 태아 중뇌동맥의 PI가 저체중아의 예후를 분석하는데 도움이 될 것으로 사려되며 앞으로 중뇌동맥에 대한 더 많은 연구가 필요할 것으로 생각된다. Doppler flow velocity waveforms of the middle cerebral arteries wer obtained in 70 normal pregnancies and 40 cases of intrauterine growth retardation(birth weight below the tenth percentile) between 24 and 40 weeks gestation. In normal pregnancies, there was a gestational age-related fall in pulsatility index for middle cerebral artevy. In growth retarded pregnancies, more reduced pulsatility index values in the middle artery were associated than normal prenancies, suggesting the presence of a $quot;brain-sparing$quot; effect. Because the fetus compensates for hypoxia by increasing blood flow to the brain, doppler middle cerebral artery waveforms may permit the evaluation of fetal compromise and hypoxia.

냉동보존 정자를 이용한 GIFT (Gamete Intrafallopian Transfer) 에 의한 임신성공 1 례

우복희,손영수,김향미,신경순 대한산부인과학회 1993 Obstetrics & Gynecology Science Vol.36 No.1

본 교실에서는 3년간 지속된 속발성 불임환자에 남편의 냉동보존정자를 이용한 GIFT를 시행한 결과 삼태임신에 성공하여 임신 302주에 조기진통으로 인해 제왕절개술로 3명의 조산아를 분만하였기에 간단한 문헌고찰과 함께 보고하는 바이다. In vitro fertilization-embryo transfer (IVF-ET) for treatment of irreparable tubal damage provides a means for otherwise hoplessly infertile couples to obtain offspring. Recently, gamete intrafallopian transfer (GIFT) was applied as an alternative to IVF-ET to patient with at least one patent fallopian tube which was simpler and less expensive. 30-years old woman who had suffered from secondary infertility for 3 years was pregnant by GIFT method using cryopreserved husband sperm due to his business trip. Primary c-section was performed at pregnancy 302 weeks due to triple pregnancy with transverse lie of the first baby and premature labor. This case was presented with brief review of the literature.

멕켈 - 그루버증후군

우복희,김영주,손영수,김향미 대한산부인과학회 1993 Obstetrics & Gynecology Science Vol.36 No.10

필자들은 최근 임신35주 4일에 복부초음파로 산전진찰하고 임신 종결후 확인된 메켈-그루버증후군 1례를 경험하였기에 간단한 문헌고찰과 함께 보고하는바이다. The main features of the meckel syndrome are postaxial polydactyly, large polycystic kidneys and occipital encephalocele, a variety of other malformations have also been described, including craniofacial anomalies, heart defects hepatomegaly and genital anomalies. The condition is a unique subgroup neural tube defects with autosomal recessive mode of inheritance. A case of severe congenital anomaly due to Meckel-Grber syndrome (polydactyly, polycystic kidney, cleft lip and palate, hypogonadism) diagnosed by ultrasonogram in the antenatal period is presented with a bredf review of the literature.