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Formation and Differentiation of Human Fetal Ovarian Follicles
도병록,이창주,송강원,윤현수,노성일,윤용달 한국발생생물학회 2000 발생과 생식 Vol.4 No.2
포유류 원시난포의 분화에는 뇌하수체에서 분비되는 gonadotropins 외에도 다양한 성장인자들 뿐 아니라 스테로이드호르몬 등이 관여하며, 난황낭에서 기원된 생식세포들과 중신에서부터 유입되는 기질세포들의 복잡한 상호작용에 의해 이루어진다. 특히 사람의 경우 태아기에 분화가 시작된 원시난포들은 성장을 개시한 후 배란이 되거나 혹은 폐쇄되어 난소에서 제거되는데, 일부 원시난포는 성장이 개시되기까지 50년 이상 원시난포의 상태로 유지된다. 그러나 원시난포의 The regulatory mechanisms of the initiation and the formation of ovarian follicles during fetal stage of mammals are largely unknown. In addition to the gonadotropins secreted from pituitary, various growth factors, and steroid hormones are believed to be involved in the differentiation and initiation of growth of primordial follicles consisting of primordial germ cells migrated from yolk sac and streamed cells from mesonephric somatic cells. In human, primordial follicles that have already initiated differentiation at fetal stage undergo either folliculogenesis to ovulate or atresia after growth. Some of primordial follicles remain without growth for 50 years or longer. The objective of this paper is to review the mechanism of the formation, growth arrest, and initiation of primordial follicles in human fetal and neonatal ovaries.
사람 다수정난자의 체외배양시 Fragmented Embryo와 Non-fragmented Embryo에서의 Methionine 유입량 및 미토콘드리아 분포양상의 비교
도병록,정미경,장미경,이경아,고정재,윤태기,차광열,Do, B.R.,Chung, M.K.,Chang, M.K.,Lee, K.A.,Ko, J.J.,Yoon, T.K.,Cha, K.Y. 대한생식의학회 1995 Clinical and Experimental Reproductive Medicine Vol.22 No.3
Despite the frequent incidence of embryo fragmentation in early human embryos, the reason of the embryo fragmentation has not been known yet. This study was conducted to investigate the histological difference(s) between fragmented (FR) and non-fragmented (NFR) human embryos focusing on comparison of mitochondrial distribution and protein synthesis. Multi-pronuclei zygotes (MPZ) such as three or more pronuclei containing in human in vitro fertilization and embryo transfer (IVF-ET) program were used for this study. MPZ were cultured in TCM-199 supplemented with 10% of human fetal cord serum (hFCS) in 5% $CO_2$ incubator at $37^{\circ}C$ for 24 hours. The cleaved embryos to 2-4 cells after 24 hours were grouped by their grade of fragmentation. Embryos were stained with Rhodamine123 (Rh123) and fluorescence was evaluated under the fluorescence microscope through PB 450-490 filter (Leitz). Regarding to protein synthesis during early human embryogenesis, there is no significant difference in the amount of synthetic proteins between FR and NFR embryos. Distribution of cytoplasmic organelles in embryos was evaluated by transmission electron microscope (TEM). The cytoplasmic distribution of mitochondria was different between FR and NFR embryos. The mitochondrial distribution was even in NFR, whereas severely aggregated in FR. It is not able to clarify in the present study whether this uneven mitochondrial distribution in FR embryo is the reason for embryo fragmentation or is the result from fragmentation. Physiological disparity related to the mitochondrial distribution may be one of the reasons for embryo fragmentation. Further studies should be addressed to investigate the physiological differences between FR and NFR embryos.
황체호르몬 수용체의 발현이 저반응 환자군의 임신에 미치는 영향
이정복,도병록,김은수,김명희,천은경,정현정,노성일,김문규,윤현수,Lee, Jung-Bok,Do, Byung-Rok,Kim, Eun-Soo,Kim, Myung-Hee,Chun, Eun-Kyung,Jeong, Hyeon-Jeong,Roh, Sung-Il,Kim, Moon-Kyoo,Yoon, Hyun-Soo 대한생식의학회 2004 Clinical and Experimental Reproductive Medicine Vol.31 No.2
Objectives: To investigate the effect of LH receptor in folliculogenesis, we confirm the expression level of LH receptor (LH-R) mRNA in human granulosa cells (GCs) and its expression levels were analyzed by comparison to embryo developmental rate and pregnancy rate. Materials and Methods: GCs were obtained at the time of oocyte retrieval from the patients undergoing IVF-ET program. The patients were divided into two groups: Group I (n=20) is poor responder (retrieved oocyte(s)$\leq$3ea), Group II (n=80) is normal responder (retrieved oocytes>3ea). After the extraction of total RNA, semiquantitative RT-PCR was performed and the expression level of LH-R mRNA was normalized by $\beta$-actin. Statistical analysis was performed by using $X^2$ test, Student's t-test and Pearson correlation. Results: In Group II, the relative values of LH-R mRNA (0.680 vs. 0.463, p<0.005) and pregnancy rate (54.7% vs. 23.1%, p<0.05) were significantly higher than in Group I. Number of retrieved oocyte(s) was gradually increased when the expression of LH-R mRNA was increased (p<0.05). But the quality of retrieved oocyte and transferred embryo were not related with the expression of LH-R mRNA. When the pregnancy rate was compared with FSH only group and FSH combined with hMG group in the ovarian stimulation protocol, FSH combined with hMG group was significantly higher than FSH only group in Group I (37.5% vs. 0%), and the expression of LH-R mRNA was significantly higher in hMG combined group than FSH only group (p<0.05). Conclusion: Expression level of LH-R mRNA has important role in ovarian function related with the response to gonadotrophin in human folliculogenesis. Furthermore these data might provide the evidence that additional use of hMG is helpful to poor responders.
Evaluation of Bisphenol a Induced Apoptosis in Sertoli Cell-lines
김지향,김진규,도병록,이창주,윤용달 한국발생생물학회 2005 발생과 생식 Vol.9 No.2
본 연구는 Leydig 세포주와 Sertoli 세포주상에 bisphenol A(BPA)와 diethylstilbestrol(DES)의 영향을 알아보고자 수행하였다. 세포 종류에 따른 BPA의 영향을 알아보기 위해, BPA의 농도별로 두 세포주에 처리하여 세포생존율을 비교하였다. Sertoli 세포주가 Leydig 세포주에 비해서 저농도의 BPA에서 생존율이 유의하게 감소되는 것을 확인할 수 있어, Sertoli 세포가 Leydig 세포주에 비해 BPA에 The present report aimed at evaluating the effect of bisphenol A(BPA) and diethylstilbestrol(DES) on Leydig or Sertoli cell-lines. To identify the differences in the susceptibility to BPA upon different cell-types, assay of the cell viability was done on TM3(Leydig cells) and TM4(Sertoli cells) cell-lines. The result indicates that Sertoli cells are more sensitive to low dose of BPA than Leydig cells. Also, the BPA- or DES-treated Sertoli cells showed a reduction of phospholipase D(PLD) activity identically. According to the confirmation of the mRNA expression of fas receptor and fas ligand in the BPA-treated cells, fas/fasL system activated by BPA will deliver the apoptosis signal onto Sertoli TM4 cells. However, Fas/FasL system was not activated in the DES-treated cells unlike the BPA-treated cells.
Bioceramic-Poly D,L-Lactic-co-Glycolic Acid(PLGA) Scaffold에접종한 인간지방조직-유래중간엽줄기세포의골형성
강유미,홍순갑,도병록,김해권,이준영 한국발생생물학회 2011 발생과 생식 Vol.15 No.2
본실험은bioceramic을첨가하여만든다공성poly D,L-lactic-co-glycolic acid(PLGA)-scaffold가인간지방조직에 서유래된중간엽줄기세포(human adipose tissue derived mesenchymal stem cells, ATMSCs)의골형성과정에효과적인지를알아보고자수행하였다. ATMSCs를well plate에접종하여골형성유도(osteogenic induction, OI) 배양액으로28일동 안배양하였다. OI배양액군의증식률은세포접종후14일까지는세포증식이활발하게진행됐지만, 21일이후세포의 증식이둔화되는양상을보였다. 반면, 기본배양액군은 꾸준한세포증식을보이며21일이후에는OI배양액군 보다더높은증식을나타냈다. OI배양액군의 alkaline phosphatase(ALP) 활성은 세포배양 21일까지는 증가했지만, 8일에는 감소한 반면에 기본배양액군은 계속 감소하는 양상을 띠었다. OI배양액군의 세포는 배양 21일에는 뚜렷한 nodule의 형성을 관찰할 수 있었고, nodule에 칼슘의 축적이 일어남을 확인하였다. ATMSCs를 scaffold에 접종하여 OI배양액으로 배양하였다. Scaffold 내 골아세포 분화에 따른 ALP 활성은 PLGA scaffold와 Bioceramic-PLGA scaffold 모두에서 세포 배양 21일에 급격히 증가하였고, Bioceramic-PLGA scaffold의 ALP 활성이 PLGA scaffold보다 크게 증가하였다. 칼슘과 인의 함량 역시 Bioceramic-PLGA scaffold에서 높게 나타났으며, Bioceramic-PLGA scaffold의 Ca/P ratio가 PLGA scaffold보다 높게 나타났다. 생체내에 이식된 scaffold의 생분해성과 광물화는 bioceramic-PLGA scaffold에서 더욱 뚜렷하게 관찰되었다. 본 실험의 결과들을 종합해 볼 때 ATMSCs의 골형성능은 well plate보다 scaffold가 더 효과적이며, bioceramic이 scaffold의 세포 부착률과 ALP 활성을 증가시켜 골형성능에 효과적으로 작용하는 것으로 생각된다. 또한 bioceramic이 ATMSCs의 골형성 분화에 따른 광물화단계의 scaffold 내 칼슘과 인의 함량을 증가시키는 것으로 사료된다. 생체 내 scaffold의 생분해성은 PLGA scaffold보다 Bioceramic-PLGA scaffold가 빠른 분해를 나타내며, 광물화에 따른 칼슘 침착이 더 활발한 것은 scaffold에 포함된 bioceramic이 생체 내 세포의 부착, 증식, 분화를 증가시켜 골형성을 촉진시키는 물질로 작용한 것으로 사료된다.