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형광직접보합법을 이용한 미배양 양수세포에서 산전 이수배수체 확인
설혜원,고희정,송남희,김숙령,이화진,오선경,박중신,전종관,윤보현,신희철,문신용,Seol, Hye-Won,Ko, Hee-Jung,Song, Nam-Hee,Kim, Sook-Ryoung,Lee, Hwa-Jin,Oh, Sun-Kyung,Park, Joong-Shin,Jun, Jong-Kwan,Yoon, Bo-Hyun,Syn, Hee-Chul,Moon, Shin- 대한생식의학회 2003 Clinical and Experimental Reproductive Medicine Vol.30 No.3
Objective: The aim of the present study was to evaluate the clinical efficiency of fluorescent in situ hybridization (FISH) in the prenatal diagnosis of chromosomal aneuploidy. Methods: We reviewed data of 268 cases to identify women undergoing genetic amniocentesis at cytogenetic laboratory, from January 2000 to December 2002. Amniotic fluid was submitted for both rapid FISH on uncultured interphase amniocytes using a commercially available DNA probe for chromosome 13, 18, 21, X, Y and standard karyotyping on cultured metaphase amniocytes. Results from FISH and full karyotype were compared. Results: There were 251 cases (84%) normal and 17 cases (16%) abnormal in FISH results. All 17 cases of trisomy 13, 18, 21 including two cases of mosaicism and sex chromosome aneuploidies which are detected by FISH were confirmed with conventional cytogenetics and there was no false positive result. Twenty two cases had karyotypically proven abnormalities that could not have been detected by the targeted FISH. Conclusion: Interphase FISH analysis of uncultured amniotic fluid cells has been shown to be an effective and reliable technique for rapid fetal aneuploidy screening during pregnancy as an adjunctive test to conventional cytogenetics.
정상과 비정상핵형을 가진 인간 배아줄기세포의 비교 분석
설혜원 ( Hye Won Seol ),백진아 ( Jin Ah Baek ),정주원 ( Ju Won Jung ),윤보애 ( Bo Ae Yoon ),김희선 ( Hee Sun Kim ),오선경 ( Sun Kyung Oh ),구승엽 ( Seung Yup Ku ),김석현 ( Seok Hyun Kim ),문신용 ( Shin Yong Moon ),최영민 ( Young M 서울대학교 인구의학연구소 2011 人口醫學硏究論集 Vol.24 No.-
Human embryonic stem cells (hESCs) are a useful source for biological research fields and applications in regenerative medicine. The maintenance and distribution of the undifferentiated hESCs of normal karyotypes are very important. The changes of colony morphology were observed during the culturing of hESCs, and chromosomal analysis was performed in this study. Abnormal karyotypes were identified in the cells which have the changes of the colony morphology. Attachment ratio and the spontaneous differentiation ratio of abnormal cells were compared to those of normal cells. High cell attachment ratio and low spontaneous differentiation ratio were confirmed in karyotypically abnormal cells. HESCs showed chromosomal anomaly if the colony morphology changed, and high attachment ratio and low spontaneous differentiation ratio have been observed during the cell culture. Our results suggest that the morphology of the colonies would play a ro1e as a criterion for chromosomal anomaly.
김희선,설혜원,안희진,오선경,구승엽,김석현,최영민,김정구,문신용,Kim, Hee-Sun,Seol, Hye-Won,Ahn, Hee-Jin,Oh, Sun-Kyung,Ku, Seung-Yup,Kim, Seok-Hyun,Choi, Young-Min,Kim, Jung-Gu,Moon, Shin-Yong 대한생식의학회 2004 Clinical and Experimental Reproductive Medicine Vol.31 No.4
Objective: This study was performed to evaluate the possibility of prolonged culture of human embryonic stem cells (hESC; SNUhES2) on human amniotic fluid cells (hAFC), which had been storaged after karyotyping. Method: The hAFC was prepared for feeder layer in the presence of Chang's medium and STO medium (90% DMEM, 10% FBS) at $37^{circ}C$ in a 5% $CO_2$ in air atmosphere. Prior to use as a feeder layer, hAFC was mitotically inactivated by mitomycin C. The hESCs on hAFC were passaged mechanically every seven days with ES culture medium (80% DMEM/F12, 20% SR, bFGF). Results: The hAFC feeder layer support the growth of undifferentiated state of SNUhES2 for at least 59 passages thus far. SNUhES2 colonies on hAFC feeder appeared slightly angular and flatter shape as compared with circular and thicker colonies observed with STO feeder layer and showed higher level with complete undifferentiation in seven days. Like hESC cultured on STO feeders, SNUhES2 grown on hAFC expressed normal karyotype, positive for alkaline phosphatase activity, high telomerase activity, Oct-4, SSEA-3, SSEA-4, Tra-1-60 and Tra-1-81 and formed embryoid bodies (EBs). Conclusion: The hAFC supports undifferentiated growth of hESC. Therefore, these results may help to provide a clinically practicable method for expansion of hESC for cell therapies.
초기 계대 인간 배아줄기세포의 해동 후 효율적인 배양 방법
백진아,김희선,설혜원,서진,정주원,윤보애,박용빈,오선경,구승엽,김석현,최영민,문신용,Baek, Jin-Ah,Kim, Hee-Sun,Seol, Hye-Won,Seo, Jin,Jung, Ju-Won,Yoon, Bo-Ae,Park, Yong-Bin,Oh, Sun-Kyung,Ku, Seung-Yup,Kim, Seok-Hyun,Choi, Young-Min,Moon, 대한생식의학회 2009 Clinical and Experimental Reproductive Medicine Vol.36 No.4
목 적: 인간 배아줄기세포 (human embryonic stem cells; hESCs)는 미분화 상태로 무한 증식할 수 있는 자가 증식(self-renewal) 능력과 인체의 모든 세포로 분화할 수 있는 전분화능 (pluripotency)의 특징을 가진 세포로, 손상된 세포를 건강한 세포로 대체하고자 하는 세포치료 (cell therapy) 연구에 활용하기 위한 세포 공급원 (cell source)으로 제시되고 있다. 그러나 인간 배아줄기세포는 확립된 초기에 세포를 안정적으로 배양하고 유지하는 과정이 쉽지 않으며, 특히 동결보존되어 있던 세포를 해동한 후 배양할 때 자연 발생적 분화가 높기 때문에 세포주의 유지에 많은 어려움이 따른다. 본 연구에서는 동결보존되어 있던 초기 계대의 인간 배아줄기세포를 해동하여 다시 배양할 때 자연 발생적 분화 부분을 기계적 분리 방법으로 제거하여 미분화 상태의 세포를 보다 빠르게 확보하기 위한 효율적인 방법에 대해 알아보고자 하였다. 연구방법: 인간 배아줄기세포를 계대 배양한지 4일이 되는 날, 50% 이상의 자연 발생적 분화가 나타난 세포군에서 분화된 부분만을 절개용 유리 피펫 (drawn-out dissecting pasture pipette)을 사용하여 기계적인 방법으로 제거하였다. 이후 지속적으로 배양액을 교환해 주며 세포군의 제거된 부분을 7일째 되는 날까지 관찰하였다. 결 과: 기계적 분리 방법을 사용하여 인간 배아줄기세포의 자연 발생적인 분화 부분을 제거한 빈 공간에 미분화상태의 인간 배아줄기세포가 분열하여 채워지는 것을 관찰하였다. 또한, 이 실험 방법을 연속 두 번 적용하여 배양했을 때 미분화 세포로 회복되는 세포군의 비율이 조금 더 높아지는 것을 확인할 수 있었다. 결 론: 동결되어 있던 초기 계대 인간 배아줄기세포의 해동 후, 자연 발생적 분화에 의해 미분화 상태를 유지하는 세포의 수가 적어 계대를 유지 하기가 어려울 때 이와 같은 기계적 분리 방법을 사용하여 자연 발생적 분화 부분을 제거한 후 배양을 지속하는 것이 단기간 내에 미분화 상태를 유지하는 인간 배아줄기세포의 양적 확보를 위한 효율적인 방법이라고 사료된다. Objective: Human embryonic stem cells (hESCs) have the capacity to differentiate into all of the cell types and therefore hold promise for cell therapeutic applications. In order to utilize this important potential of hESCs, enhancement of currently used technologies for handling and manipulating the cells is required. The cryopreservation of hESC colonies was successfully performed using the vitrification and slow freezing-rapid thawing method. However, most of the hESC colonies were showed extremely spontaneous differentiation after freezing and thawing. In this study, we were performed to rapidly collect of early passage hESCs, which was thawed and had high rate of spontaneously differentiation of SNUhES11 cell line. Methods: Four days after plating, partially spontaneously differentiated parts of hESC colony were cut off using finely drawn-out dissecting pipette, which is mechanical separation method. Results: After separating of spontaneously differentiated cells, we observed that removed parts were recovered by undifferentiated cells. Furthermore, mechanical separation method was more efficient for hESCs expansion after thawing when we repeated this method. The recovery rate after removing differentiated parts of hESC colonies were 55.0%, 74.5%, and 71.1% when we have applied this method to three passages. Conclusion: Mechanical separation method is highly effective for rapidly collecting and large volumes of undifferentiated cells after thawing of cryopreserved early passage hESCs.