ICSI 프로그램에서 생쥐 투명대를 이용한 고환조직내 정자의 동결

서태광,전병균,류은경,이은숙,류재웅,손시환,문진수,김광철,Suh, Tae-Kwang,Jeon, Byeong-Gyun,Ryu, Eun-Kyung,Lee, Eun-Sook,Ryoo, Zae-Yoong,Sohn, Sea-Hwan,Moon, Jin-Soo,Kim, Kwang-Chull 대한생식의학회 1997 Clinical and Experimental Reproductive Medicine Vol.24 No.2

The survival rate and motility recovered after cryopreservation of testicular spermatozoa in testicular sperm extraction (TESE)-ICSI program is low. The purpose of this study was to assess the availability and efficiency of mouse empty zona pellucida in cryopreserving human TESE spermatozoa. Mouse empty zonae pellucidae were obtained by extraction of cytoplasm with or without cytochalasin B treatment. Motile sperm from proven-fertile donor and two azoospermic patients after TESE were individually inserted into empty zona pellucida and cryopreserved. Two to five days after cyropreservation, the frozen sperm were thawed and the rates of recovery and motility were observed. The ooplasmic extraction rates of control (N=80) and cytochalasin B treated oocytes (N=80) were 94.0% and 96.2%, respectively (p>0.05). The post-thaw recovery rates of spermatozoa and rates of motility recovery of ejaculate (N=70) and testicular (N=70) sperm were 97.1%, 97.1% and 95.7%, 94.3%, respectively (p>0.05). The results of this study showed that the mouse zone pellucida is useful for cryostorage of single testicular spermatozoa.

Efficiency of the Establishment of Sheep Embryonic Stem Cell Lines from Whole Blastocysts and Isolated Inner Cell Masses

Zae-Yoong Ryoo(류재웅),Tae-Kwang Suh(서태광),Kyu-Shik Jeong(정규식),Sun-Uk Kim(김선옥),Kyung-Kwang Lee(이경광),Byoung-Gil Mheen(민병길),Byung-Hwa Hyun(현병화) 한국실험동물학회 1996 Laboratory Animal Research Vol.12 No.2

Establishment of Pluripotent Cell Lines Derived from Bovine Embryo Fertilized In vitro

Zae-Yoong Ryoo(류재웅),Byung-Hwa Hyun(현병화),Tae-Kwang Suh(서태광),Kyu-Shik Jeong(정규식),Sun-Uk Kim(김선옥),Byoung-Gil Mheen(민병길),Kyung-Kwang Lee(이경광) 한국실험동물학회 1996 Laboratory Animal Research Vol.12 No.2

세포질내 정자주입법(ICSI)에 있어서 정자흡입 및 난자내 주입방법에 관한 연구

이택후 ( Taek Hoo Lee ),김항진 ( Hang Jin Kim ),송건호 ( Gun Ho Song ),김대근 ( Dae Geun Kim ),전상식 ( Sang Sik Chun ),박윤규 ( Yoon Kyu Park ),서태광 ( Tae Kwang Suh ),전병균 ( Byeong Gyun Jeon ),류은경 ( Eun Kyung Ryu ),이은숙 대한산부인과학회 1997 Obstetrics & Gynecology Science Vol.40 No.12

Immobilization of spermatozoa prior to intracytoplasmic sperm injection(ICSI) sometimes results in crooked tail and this makes it difficult to aspirate sperm into an injection pipette tail first Head-first sperm aspiration into an injection pipette avoid this problem due to the bigger size of the sperm head. The effect of head or tail-first sperm injection into an oocyte on fertilization, cleavage, percentage of grade 1 embryos and development to blastocyst stage in ICSI program has been studied. A single living immobilized spermatozoa from oligoasthenozoospermic patient was injected into an oocyte head-first or tail-first according to the treatment. Eighteen hours after microinjection, oocytes were inspected for survival and fertilization Fertilized oocytes with two pronuclei were cultured in 30 μ I drop of mHTF supplemented with 10 % heat-inactivated follicular fluid(FF) at 37℃. On day 2, embryo transfer was performed with cleaved embryos. The remaining 2-8 cell stage embryos were co-cultured with BRL cells in mHTF+10 % FF for 72 hours and the developmental stage was observed. The data were analyzed by Analysis of Variance. A total of 164 oocytes from 36 cycles were assigned to each treatment and ICSI was performed(88 head-first, 76 tail-first). The rates of normal fertilization were 81.8 % and 76.3% for head-first and tail-first, respectively. Of the fertilized oocytes, the percentage of cleaved embryos and the percentage of grade 1 embryo among cleaved embryos were 88.9 % and 68.8 %, 93.1 % and 74.1 % for head-first and tail-first, respectively. Of the 2-8 cell embryos cultured, 44.4 %(16/36) and 50.0%(10/20) for head first and tail first, respectively developed to blastocyst stage. There were no differences in fertilization, cleavage, rates of grade 1 embryos, and development to blastocyst stage. In conclusion, head-first or tail-first sperm injection into an oocyte in ICSI program does not affect fertilization and subsequent embryo development to blastocyst stage in vitro.

생쥐에 있어서 兩分分割球 移植에 의한 새끼生産에 관한 硏究

徐泰光,朴恒均 慶北大學校 農業科學技術硏究所 1988 慶北大農學誌 Vol.6 No.-

本 硏究는 생쥐에 대한 過批卵反應, 回收된 受精卵의 兩分率, 兩分된 分割球의 體外培養 및 移植後 産仔로의 發達率을 조사하기 위하여 CBA 및 C57BL 系統의 供試싱쥐에서 얻어진 受精卵을 0.5% protease로 투명대제거 및 미세유리봉으로 兩分하여 얻어진 分割球를 blastocyst로 培養하여 BALB/C 系統의 受卵생쥐에 移植하여 다음과 같은 結果를 얻었다. hCG 注射後 36∼42時間, 48∼54時間, 62∼66時間, 72∼78時間에 受精卵을 回收한 結果 各各 2細胞期, 4細胞期, 8細胞期 및 桑實胚期의 受精卵이 대부분 回收되었으며 首當 平均 17.0個의 卵子가 回收되었다. 回收後 兩分한 8細胞期 受精卵의 兩分率은 83.6%로서 桑實胚의 65.5%에 比해 유의적으로 높았으며 分割球의 發達率은 桑實胚에서 얻어진 分割球가 76.5%로서 8細胞期 分割球의 60.9%에 比해 유의적으로 높았다. 한편 培養된 分割球와 intact embryo에 있어서 細胞期에 따라 各各 受卵생쥐當 2個, 10個, 15個씩 移植하였을 때 共히 15個씩 移植한 경우 受胎率이 가장 높았으나 細胞期에 따른 受胎率에 있어서 유의적인 차이는 없었다. 培養된 分割球와 intact embryo를, 桑實胚에서도 intact embryo를 移植하였을때 새끼 生産率은 各各 37.6% 및 43.4%로서 各各 培養된 分割球를 移植하였을때 보다 새끼生産率이 유의적으로 높았다. This study was conducted to investigate the effect of embryo stage on bisection rates of embryos, on development of separated demi-embryos and on subsequent development to full term following transfer of demi-embryos to recipients. The results obtained in this study are summarized as follows. The 2-, 4-, 8-cell and morula embryos were mostly obtained at 36-42, 48-54, 62-66 and 72-78 hours after injection of hCG, respectively, and the average number of embryos recovered per head was 17.0. The bisection rate of 8-cell embryos was 83.6%, which was significantly higher than that of morula embryos, 65.5%. But the development of morula demi-embryos to blastocyst after in-vitro culture was better than that of 8-cell demi-embryos and the rate was 76.5% and 60.9%, respectively. When the 2, 10 and 15 cultured demi-embryos or intact embryos were transferred to each recipient mouse, respectively. the highest pregnancy rate was obtained when 15 embryos were transferred. And the overall offspring production rate of intact embryos was higher than that of demi-embryos.

산양 배 분할구의 체내배양에 관한 연구

최광수,박항균,서태광 경북대학교 유전공학연구소 1987 遺傳工學硏究所報 Vol.2 No.1

In order to obtain basic informations and to develop basic techniques for the production of monozygotic twins of Korean native goats, the study of superovulation, ova recovery, bisection of embryos and in vivo culture of separated blastomeres was carried out. For the study, 19 goats were treated with FSH to induce superovulation and, as a result, 16 goats were responded. Ova or embryos were recovered from the uterus by surgical method after 4.0-7.0 days from the first mating. The mean number of corpora lutea and recovered ova were 10.1±3.3 and 5.9±2.5, respectively. 53 morula were recovered after 5.0-6.0 days from the first mating. Of these recovered morula, 48 were bisected by glass needle or razor blade and 61 available demi-embryos were obtained among them. 61 available demi-embryos were cultured in vivo and after culture for 48 hours, low rate of the demi-embryos was recovered. However, some of the recovered demi-embryos were developed to blastocyst.

兩分한 생쥐 胚의 體外 및 體內 發生에 관한 硏究

崔光洙,全益秀,朴修奉,徐泰光,朴恒均 慶北大學校農業科學技術硏究所 1990 慶北大農學誌 Vol.8 No.-

本 연구는 micromanipultor를 이용하여 생쥐의 8세포기배와 상실배 그리고 배반포기배를 兩分후 생존성을 검토하고, 또한 배반포기배를 양분후, 選別 및 배양 과정없이 암컷생쥐에 移植하는 경우 새끼쥐 생산의 가능성을 검토하고자 수행된 것이다. 그 결과를 요약해 보면 다음과 같다. 1. 생쥐의 8細胞期胚와 桑實胚를 兩分하여 M2에 배양한결과 각각 64%, 81%가 胚盤胞期胚까지 發生하였다. 2. M2 배양액에서 발생시킨 배반포기배를 Ham's F-10에서 배양한 결과 8세포기배에서는 86%, 상실배에서는 90%가 정상적으로 outgrowth 되었다. 3. 배반포기배를 배양 과정없이 바로 兩分하여 Ham's F-10에서 배양한 결과 97%가 정상적으로 outgrowth되었으나 암컷생쥐에 이식한 결과 산차는 얻지 못하였다. The study was carried out to investigate the viability of mouse embryos bisected at the stages of 8-cell, morula and blastocyst and, also, to find out the feasibility of offspring production by transfer of the bisected blastocysts with Quick-splitting. The results obtained are summarized as follows: 1. The mouse embryos were bisected with bio-cut blade at the stages of 8-cell and morula, and cultured in Medium 2. The bisected embryos were developed to blastocyst stage by 64% and 81%, respectively. 2. The blastocysts which were cultured in Medium 2 after being bisected at the stages of 8-cell and morula were observed normal outgrowth in Ham's F-10 medium by 86% and 90%, respectively. 3. The blastocysts which were Quick-splitted in Medium 2 were observed normal outgrowth by 97%, however, no offspring was obtained by transfer of Quick-splitted blastocysts.

세포질내 정자주입법(ICSI)에 있어서 정자흡입 및 난자내 주입방법에 관한 연구

이택후,김항진,송건호,김대근,전상식,박윤규,서태광,전병균,류은경,이은숙,문진수,김광철 경북대학교 의학연구소 2000 경북대학교병원의학연구소논문집 Vol.4 No.1

Study on Method of Sperm Aspiration and Injection into an Oocyte in Intracytoplasmic Sperm Injection(ICSI) Immobilization of spermatozoa prior to intracytoplasmic sperm iniection(ICSI) sometimes results in crooked tail and this makes it difficult to aspirate sperm into an injection pipette tail first. Head-first sperm aspiration into an injection pipette avoid this problem due to the bigger size of the sperm head. The effect of head or tail-first sperm injection into an oocyte on fertilization cleavage, percentage of grade I embryos and development to blastocyst stage in ICSI program has been studied. A single living immobilized spermatozoa from oligoasthenozoospermic patient was injected into an oocyte head-first or tail-first according to the treatment. Eighteen hours after microinjection, oocytes ware inspected for survival and fertilization Fertilized oocytes with two pronuclei were cultured in 30μl drop of mHTF supplemented with 10% heat-inactivated follicular fluid(FF) at 37℃. On day 2. embryo transfer was performed with cleaved embryos. The remaining 2-8 cell stage embryos were co-cultured with BRL cells in mHTF + 10% FF for 72 hours and the developmental stage was observed. The data were analyzed by Analysis of Variance. A total of 164 oocytes from 36 cycles were assigned to earth treatment and ICSI was performed(88 head-first, tail-first). The rates of normal fertilization were 81.8% and 76.3% for head-first and tail-first, respectively. Of the fertilized oocytes, the percentage of cleaved embryos and the percentage of grade 1 embryo among cleaved embryos were 88.9% and 68.8%, 93.1% and 74.1% for head-first and tail-first, respectively. Of the 2-8 cell embryos cultured, 44.4%(16/36) and 50.0%(10/20) for head first and tail first, respectively developed to blastocyst stage. There were no differences in fertilization, cleavage, rates of grade 1 embryos, and development to blastocyst stage. In conclusion, head-first or tail-first sperm injection into an oocyte in ICSI program does not affect fertilization and subsequent embryo development to blastocyst stage in vitro.