Clinical and Experimental Reproductive Medicine

  • 제35권4호
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  • Pages.247-265
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  • 2008
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  • 2233-8233(pISSN)
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  • 2233-8241(eISSN)
대한생식의학회 (The Korean Society for Reproductive Medicine)
권호 표지

사람의 제대 및 양막유래 줄기세포의 간세포로의 분화

Hepatogenic Potential of Umbilical Cord Derived-Stem Cells and Human Amnion Derived-Stem Cells

  • 김지영 (서울여자대학교 자연과학대학 생명공학과) ;
  • 이윤정 (수원제일산부인과) ;
  • 박세아 (서울여자대학교 자연과학대학 생명공학과) ;
  • 강현미 (서울여자대학교 자연과학대학 생명공학과) ;
  • 김경식 (연세대학교 의과대학 외과학교실) ;
  • 조동제 (연세대학교 의과대학 산부인과학교실) ;
  • 김해권 (서울여자대학교 자연과학대학 생명공학과)
  • Kim, Ji-Young (Department of Biotechnology, College of Natural Sciences, Seoul Women's University) ;
  • Lee, Yoon-Jung (Suwonjeil Women's Hospital) ;
  • Park, Se-Ah (Department of Biotechnology, College of Natural Sciences, Seoul Women's University) ;
  • Kang, Hyun-Mi (Department of Biotechnology, College of Natural Sciences, Seoul Women's University) ;
  • Kim, Kyung-Sik (Department of Surgery, College of Medicine, Yonsei University) ;
  • Cho, Dong-Jae (Department of OB/GYN, College of Medicine, Yonsei University) ;
  • Kim, Hae-Kwon (Department of Biotechnology, College of Natural Sciences, Seoul Women's University)
  • 발행 : 2008.12.30
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초록

목 적: 사람의 제대유래 줄기세포 (HUC)와 양막유래 줄기세포 (HAM)를 간유사세포로 분화 시키고자 하였다. 연구방법: 산모의 동의아래 만삭 정상 산모로부터 양막 및 제대를 얻고 줄기세포를 분리, 배양하였다. 분리된 세포의 줄기세포로서의 특성을 규명하기 위하여 RT-PCR, 세포면역화학 분석 그리고 중배엽성세포로의 분화능 실험을 하였다. 또한 이들 줄기세포를 분화 배양액에서 간유사세포로 분화 유도 후, 간세포 특이 유전자 발현, 알부민 ELISA, 알부민에 대한 면역블로팅 및 세포면역화학염색 그리고 PAS 염색을 시행하였다. 결 과: 사람의 양막 및 제대로부터 세포를 분리하여 줄기세포로서의 특성을 조사하고, 이들 세포의 지방세포, 연골세포, 골아세포로의 분화능력을 확인하였다. 동일한 분화 배양액에서 HUC과 HAM을 간유사세포로 분화시킨 결과 HUC이 모든 조건에서 HAM보다 알부민과 요소를 더 많이 합성 분비하였다. 간유사세포로의 분화능력이 더 좋은 것으로 나타난 HUC을 가지고 최적의 분화를 유도한 결과 $1.2{\pm}0.8\;{\mu}g/mL$의 알부민을 합성 분비하였고, $8.9{\pm}1.2\;mg/dL$의 요소를 합성 분비하였다. 결 론: HUC과 HAM 모두 알부민을 분비하는 간유사세포로 분화할 수 있었으며, HUC이 더 잘 유도되는 것으로 관찰되었다. 이러한 결과로써 HUC과 HAM을 간질환 환자의 간세포 이식 치료에 필요한 세포치료제로써 이용할 수 있을 것으로 생각된다.

Objectives: Many types of liver diseases can damage regenerative potential of mature hepatocytes, hepatic progenitor cells or oval cells. In such cases, a stem cell-based therapy can be an alternative therapeutic option. We examined whether human amnion-derived mesenchymal stem cells (HAM) and human umbilical cord-derived stem cells (HUC) could differentiate into hepatocyte-like cells as therapeutic cells for the liver diseases. Methods: HAM and HUC were isolated from the amnion and umbilical cord of the volunteers after a caesarean section with informed consent. In order to differentiate these cells into hepatocyte-like cells, cells were cultivated in hepatogenic medium using culture plates coated with fibronectin. Effects of hepatocyte growth factor, L-ascorbic acid 2-phosphate, insulin premixture fibroblast growth gactor 4, dimethylsulfoxide, oncostatin M and/or dexamethasone were examined on the hepatic differentiation. After differentiation, the cells were analyzed by RT-PCR, immunocytochemistry, immunoblotting, albumin ELISA, urea assay and periodic acid-schiffs staining. Results: Initial fibroblast-like appearance of HAM and HUC changed to a round shape during culture in the hepatogenic medium. However, in all hepatogenic conditions examined, HUC secreted more amounts of albumin or urea into medium than HAM. Expression of some of hepatocyte-specific genes increased and expression of new genes were observed in HUC following cultivation in hepatogenic medium. Results of immunocytochemistry and immunoblotting analyses demonstrated that HUC secreted albumin into the culture medium. PAS staining further demonstrated that HUC could store glycogen inside of the cells. Conclusions: Both HUC and HAM could differentiate into albumin-secreting, hepatocyte-like cells. Under the same hepatogenic conditions examined, HUC more efficiently differentiated into hepatocyte-like cells compared with the HAM. The results suggest that HUC and HAM could be used as sources of stem cells for the cell-based therapeutics such as in liver diseases.

키워드

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