심층신경망 기반의 스케일러블 이미지 전송 최적화

류성미(Sung Mi Ryu),유시환(See-Hwan Yoo),장석호(Seok-Ho Chang) 대한전자공학회 2021 대한전자공학회 학술대회 Vol.2021 No.6

생쥐배아의 부화와 탈각에 미치는 Pronase의 영향

문신용,최성미,김희선,류범용,오선경,서창석,김석현,최영민,김정구,최규홍,이진용,Moon, Shin-Yong,Choi, Sung-Mi,Kim, Hee-Sun,Ryu, Buom-Yong,Oh, Sun-Kyung,Suh, Chang-Suk,Kim, Seok-Hyun,Choi, Young-Min,Kim, Jung-Gu,Choi, Kyu-Hong,Lee, Jin-Y 대한생식의학회 2000 Clinical and Experimental Reproductive Medicine Vol.27 No.4

Objective: Hatching of the blastocyst from the zona pellucida (ZP) is a key event in mammalian implantation. In vivo, two factors have been identified as possible mediators of hatching: lysis of the ZP by substances elaborated either from the embryo or female reproductive tract and pressure exerted on the zona by expansion of the blastocyst. Two methods of zona manipulation were already in use to enhance the ability of embryos to hatch: mechanical PZD and chemical ZD by acidic Tyrode's solution. But several controversies of each method have been reported. The purpose of this study was to investigate the effect of pronase for mouse embryo hatching. Methods: Mouse embryos were obtained following ovulation induction of $F_1$ animals. Fresh and cryo-thawed morula embryos were exposed to 0.5, 1.0, 2.0, 5.0 ${\mu}g/ml$ pronase in Ham's F10 for 72 hrs. Main outcome measures were the rates of partial hatching and completely hatched blastocysts, and cell number of it. Results: In fresh and cryo-thawed group, the rates of completely hatched blastocyst were significantly higher in 5 ${\mu}g/ml$ pronase treatment group than control group. There was no difference in completely hatched blastocyst total cell number between pronase treatment group and control group. This suggest that pronase treatment did not harmful in mouse embryo development. In pronase treatment group, zona pellucida were thinner than control group. Conclusion: The addition of pronase to culture media may accelerate the hatching of embryo. So, enzymatic treatment of the zona may provide a valuable and effective assisted hatching technique for human in-vitro fertilization-embryo transfer.

착상 전 유전진단 기술 개발의 동물실험 모델로서 할구 생검된 생쥐 배아에서 동결보존 융해 후 배아 발생 양상과 공배양 효과에 관한 연구

김석현,김희선,류범용,최성미,방명걸,오선경,지병철,서창석,최영민,김정구,문신용,이진용,채희동,김정훈,Kim, Seok-Hyun,Kim, Hee-Sun,Ryu, Buom-Yong,Choi, Sung-Mi,Pang, Myung-Geol,Oh, Sun-Kyung,Jee, Byung-Chul,Suh, Chang-Suk,Choi, Young-Min,Kim, Jung 대한생식의학회 2000 Clinical and Experimental Reproductive Medicine Vol.27 No.1

Objective: The effects of cryopreservation with or without coculture on the in vitro development of blastomere-biopsied 8-cell mouse embryos were investigated. This experimental study was originally designed for the setup of a preclinical mouse model for the preimplantation genetic diagnosis (PGD) in human. Methods: Eight-cell embryos were obtained after in vitro fertilization (IVF) from F1 hybrid mice (C57BL(표현불가)/CBA(표현불가)). Using micromanipulation, one to four blastomeres were aspirated through a hole made in the zona pellucida by zona drilling (ZD) with acid Tyrode's solution (ATS). A slow-freezing and rapid-thawing protocol with 1.5M dimethyl sulfoxide (DMSO) and 0.1M sucrose as cryoprotectant was used for the cryopreservation of blastomere- biopsied 8-cell mouse embryos. After thawing, embryos were cultured for 110 hours in Ham's F-10 supplemented with 0.4% bovine serum albumin (BSA). In the coculture group, embryos were cultured for 110 hours on the monolayer of Vero cells in the same medium. The blastocyst formation was recorded, and the embryos developed beyond blastocyst stage were stained with 10% Giemsa to count the total number of nuclei in each embryo. Results: The survival rate of embryos after cryopreservation was significantly lower in the blastomere-biopsied (7/8, 6/8, 5/8, and 4/8 embryos) groups than in the non-biopsied, zona intact (ZI) group. Without the coculture, the blastocyst formation rate of embryos after cryopreservation was not significantly different among ZI, the zona drilling only (ZD), and the balstomere-biopsied groups, but it was significantly lower than in the non-cryopreserved control group. The mean number of cells in embryos beyond blastocyst stage was significantly higher in the control group ($50.2{\pm}14.0$) than in 6/8 ($26.5{\pm}6.2$), 5/8 ($25.0{\pm}5.5$), and 4/8 ($17.8{\pm}7.8$) groups. With the coculture using Vero cells, the blastocyst formation rate of embryos after cryopreservation was significantly lower in 5/8 and 4/8 groups, compared with the control, 7/8, and 6/8 groups. The mean number of cells in embryos beyond blastocyst stage was also significantly lower in 4/8 group ($25.9{\pm}10.2$), compared with the control ($50.2{\pm}14.0$), 7/8 ($56.0{\pm}22.2$), and 6/8 ($55.3{\pm}25.5$) groups. Conclusion: After cryopreservation, blastomere-biopsied mouse embryos have a significantly impaired developmental competence in vitro, but this detrimental effect might be prevented by the coculture with Vero cells in 8-cell mouse embryos biopsied one or two blastomeres. Biopsy of mouse embryos after ZD with ATS is a safe and highly efficient preclinical model for PGD of human embryos.

남성인자 불임환자의 ICSI 시술 후 발생한 배아에서 Multicolor FISH 의 임상 적용을 이용한 염색체 이상의 착상 전 유전진단

김석현(Seok Hyun Kim),최성미(Sung Mi Choi),김희선(Hee Sun Kim),류범용(Buom Yong Ryu),방명걸(Myung Geol Pang),오선경(Sun Kyung Oh),구승엽(Seung Yup Ku),지병철(Byung Chul Jee),서창석(Chang Suk Suh),최영민(Young Min Choi),김정구(Jung Gu 대한산부인과학회 2000 Obstetrics & Gynecology Science Vol.43 No.9

Objective : The genetic defects in human gametes and embryos can cause the adverse effects on overall reproductive events. Biopsy of embryos for preimplantation genetic diagnosis(PGD) offers a new possibility of having children free of the genetic disease. This study was performed for the clinical application of Multicolor fluorescent in situ hybridization(FISH) to make a diagnosis of chromosomal aneuploidy, one of the most frequent cytogenetic abnormalities, in the biopsied blastomeres of human embryos. Materials and Methods : Clinical PGD of chromosomal aneuploidy with the Multicolor FISH technique which was recently developed and optimized in our laboratory was performed in the biopsied blastomeres of human embryos obtained from in vitro fertilization(IVF) with intracytoplasmic sperm injection(ICSI) using the ejaculated and testicular sperms in the severe male factor infertility couples. Results : Normal pregnancy was achieved after intrauterine transfer of the chromosomally normal embryos excluding the embryos with aneuploidy diagnosed successfully by PGD with the multicolor FISH technique, and confirmed by karyotyping of the cultured amniocytes after amniocentesis. As a result, the clinical efficacy of multicolor FISH directly applied to the chromosome analysis of embryos obtained from IVF was confirmed. Conclusion : PGD with Multicolor FISH can be an efficacious diagnostic technique in the establishment of normal pregnancy, especially when it is applied to the male factor infertility couples who are at high risk of having aneuploid offsprings. In addition, PGD makes it possible to achieve a higher pregnancy rate with the selection and transfer of normal embryos, compared with the conventional IVF with ICSI.

인간 배아에서 착상 전 유전진단을 위한 Multicolor Fluorescence In Situ Hybridization 기술의 개발

김석현(Suk Hyun Kim),최성미(Sung Mi Choi),김희선(Hee Sun Kim),류범용(Bum Yong Ryu),방명걸(Myung Geol Bang),오선경(Sun Gyung Oh),지병철(Byung Chul Jee),서창석(Chang Suk Seo),최영민(Young Min Choi),배광범(Gwang Bum Bae),김정구(Jung Goo 대한산부인과학회 2000 Obstetrics & Gynecology Science Vol.43 No.12

Objective : The genetic defects in human gametes and embryos can cause the adverse effects on overall reproductive events. Biopsy of embryos for preimplantation genetic diagnosis(PGD) offers a new possibility of having children free of the genetic disease. In this study, we attempted to attain the optimized methodology of Multicolor fluorescent in situ hybridization(FISH) technique, and then to apply the developed technique for the diagnosis of chromosomal abnormality such as aneuploidy in biopsied blastomeres of human embryos in a shorter time. Methods : In the first-year study, various experimental conditions for FISH applied to the chromosomes of metaphase and interphase nuclei were performed to get an accurate and rapid result of FISH, and the simultaneous denaturation technique of both target DNA and DNA probe was developed. In the second-year study, the experimental conditions for FISH established appropriately in the first year were advancingly improved and optimized utilizing the abnormally fertilized and developmentally arrested human embryos to establish the clinical efficacy of PGD in human embryos. Results : In the first-year study, the direct method of FISH with the sex chromosome-specific DNA probes could show the results 2.5 hours earlier than the indirect method, and there were no false negative results in either method. The next step was to develop the simultaneous denaturation technique of both target DNA and DNA probe, and the duration time to get the results could be shortened by an hour without affecting the results of FISH, compared with the conventional method. Accordingly, the most appropriate methodology of FISH was developed and established through reducing the required time for FISH by 3.5 hours, compared with the conventional method. Human blastomere biopsy technique was also acquired utilizing the abnormally fertilized embryos in IVF-ET program, and the application of newly developed FISH technique resulted in 70% of success rate in our laboratory. In the second-year study, the conditions for blastomere preparation prior to FISH were improved, and Multicolor FISH technique was developed for the simultaneous availability of information on many chromosomes. Multicolor FISH was applied to the blastomeres biopsied from the abnormally fertilized and developmentally arrested human embryos, and the fluorescent signals could be observed successfully in more than 90% of blastomeres. With the application of 6 color FISH to 118 blastomeres biopsied from 37 abnormally fertilized embryos in 23 infertile patients undergoing IVF-ET, various chromosomal abnormalities were analyzed in 114 blastomeres(96.6%). Conclusions : The most appropriately optimized Multicolor FISH technique was developed and established for the practical use of clinical application. The final goal of this project on the development of PGD using Multicolor FISH technique for the chromosomal abnormality in human embryos was successfully achieved by establishment of the prevention measures on the birth of infants with aneuploidy which is one of the most frequent chromosomal anomalies.

난자 세포질내 정자 주입술을 이용한 체외수정시술시 누적임신율에 관한 연구

김석현(Suk Hyun Kim),심순섭(Soon Sub Shim),지병철(Byung Chul Jee),최성미(Sung Mi Choi),김희선(Hee Sun Kim),류범용(Bum Yong Ryu),오선경(Sun Gyung Oh),서창석(Chang Suk Seo),최영민(Young Min Choi),배광범(Gwang Bum Bae),김정구(Jung Goo K 대한산부인과학회 2001 Obstetrics & Gynecology Science Vol.44 No.1

Objective : To compare the clinical outcomes between Day 2 and Day 3 embryo transfer(ET) groups in in vitro fertilization and embryo transfer(IVF-ET) with intracytoplasmic sperm injection(ICSI). Methods : From May, 1997 to December, 1998, 174 cycles of IVF-ET with ICSI were performed and classified into two groups : Day 2 ET group(n=134) and Day 3 ET group (n=40). In Day 3 ET group, embryos fertilized after ICSI were cultured in vitro for further 24 hours in M3 media. Results : There were no significant differences in the age and BMI of patients, basal serum FSH level, protocol of controlled ovarian hyperstimulation(COH), indication of ICSI, and source of sperm for ICSI between two groups. Only the number of the previous failed IVF-ET cycles was significantly higher in Day 3 ET group(p<0.05). Serum E2 level on hCG day, the numbers of oocytes retrieved after COH, oocytes fertilized after ICSI, and embryos transferred, and the rates of fertilization, cleavage, and implantation showed no significant differences. However, cumulative embryo score(CES) was significantly higher in Day 3 ET group(p<0.05). Although there were no significant differences in the rates of pregnancy per ET, spontaneous abortion, and live birth, the rates of biochemical and multiple pregnancy were significantly higher in Day 3 ET group(p<0.05). Conclusions : In IVF-ET with ICSI, the relatively higher CES may contribute to the higher risk of multiple pregnancy in Day 3 ET group, compared with the conventional Day 2 ET group.

난자 세포질내 정자 주입술을 이용한 체외수정시술시 누적임신율에 관한 연구

김석현(Seok Hyun Kim),심순섭(Soon Sup Shim),지병철(Byung Chul Jee),최성미(Sung Mi Choi),김희선(Hee Sun Kim),류범용(Buom Yong Ryu),오선경(Sun Kyung Oh),서창석(Chang Suk Suh),최영민(Young Min Choi),배광범(Kwang Bum Bai),김정구(Jung Gu 대한산부인과학회 2001 Obstetrics & Gynecology Science Vol.44 No.3

Objective : To evaluate the cumulative pregnancy rate(CPR) of in vitro fertilization and embryo transfer(IVF-ET) with intracytoplasmic sperm injection(ICSI). Methods : Medical records of 260 infertile patients undergoing 519 cycles of IVF-ET with ICSI from January, 1994 to December, 1999 were retrospectively reviewed. The CPR beyond 12 weeks of gestation was estimated by Kaplan-Meier method. The CPRs were compared by log-rank test between groups divided by age of patients, indication of ICSI, and method of sperm retrieval for ICSI. Results : As 70 patients achieved an on-going pregnancy after IVF-ET with ICSI, the PR was 26.9% per patient and 13.5% per cycle. The overall CPR was 54.9% after 6 cycles of IVF-ET with ICSI. As expected, age had a significant strong effect on the CPR; CPRs afer 4 cycles of ICSI were 61.8% in the age group of 30 years(n=81), 43.7% in 31-35 years(n=106), and 15.3% in 36 years(n=73). There was no significant difference in the CPR between abnormal semen analysis group(n=184) and prior low fertilization rate group(n=66). In abnormal semen analysis group, the CPR of surgically retrieved sperm subgroup(n=60) was not significantly different from that of ejaculated sperm subgroup(n=124). Conclusions : The CPR of IVF-ET with ICSI was presented, and it could be of much help in the clinical counseling of IVF-ET patients. ICSI technique could be used successfully for IVF-ET in infertile couples who had the male factor infertility or the past history of low fertilization rate in the previous cycles.