Reproductive and Developmental Biology

The Korean Society of Animal Reproduction (한국동물번식학회)
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Efficient Gene Delivery into Hematopoietic Stem Cells by Intra-Bone Marrow Injection of Retrovirus

IBM 이식을 통한 골수 조혈 줄기 세포에의 효과적인 유전자 도입

  • Lee, Byun-Joo (Graduate School of Life Science and Biotechnology, College of Medicine, CHA Stem Cell Institute, Pochon CHA University) ;
  • Lee, Yong-Soo (Graduate School of Life Science and Biotechnology, College of Medicine, CHA Stem Cell Institute, Pochon CHA University) ;
  • Kim, Hye-Sun (Graduate School of Life Science and Biotechnology, College of Medicine, CHA Stem Cell Institute, Pochon CHA University) ;
  • Kim, Yu-Kyung (Graduate School of Life Science and Biotechnology, College of Medicine, CHA Stem Cell Institute, Pochon CHA University) ;
  • Kim, Jae-Hwan (Graduate School of Life Science and Biotechnology, College of Medicine, CHA Stem Cell Institute, Pochon CHA University) ;
  • Park, Jin-Ki (Animal Biotechnology Division, National Livestock Research Institute) ;
  • Chung, Hak-Jae (Animal Biotechnology Division, National Livestock Research Institute) ;
  • Chang, Won-Kyong (Animal Biotechnology Division, National Livestock Research Institute) ;
  • Kim, Dong-Ku (Graduate School of Life Science and Biotechnology, College of Medicine, CHA Stem Cell Institute, Pochon CHA University)
  • 이현주 (포천중문의과대학교 차 줄기세포 연구소) ;
  • 이용수 (포천중문의과대학교 차 줄기세포 연구소) ;
  • 김혜선 (포천중문의과대학교 차 줄기세포 연구소) ;
  • 김유경 (포천중문의과대학교 차 줄기세포 연구소) ;
  • 김재환 (포천중문의과대학교 차 줄기세포 연구소) ;
  • 박진기 (축산과학원 응용생명공학과) ;
  • 정학재 (축산과학원 응용생명공학과) ;
  • 장원경 (축산과학원 응용생명공학과) ;
  • 김동구 (포천중문의과대학교 차 줄기세포 연구소)
  • Published : 2008.03.31
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Abstract

Efficient gene transfer into hematopoietic stem cells is a great tool for gene therapy of hematopoietic disease. Retrovirus have been extensively used for gene delivery and gene therapy. However, current in vitro gene transfer has some obstacles suck as induction of differentiation loss of self-renewal capacity, and down-regulation of homing efficiency for in vitro hematopoietic stem cells transplantation. To overcome these problems, we developed efficient in vitro retroviral transfer technique by direct intra-bone marrow injection (IBM). We identified effective retrovirus gene transfer in bone marrow hematopoietic cells in vitro. Two weeks after retrovirus transfer via IBM injection, we observed stable EGFP gene expression in bone marrow, lymph node, spleen, and liver cells. In addition, $6.4{\pm}2.7%$ of hematopoietic stem/progenitor cells were expressed EGFP transgene from flow cytometry analysis. Our results demonstrate that in vitro retrovirus gene transfer via IBM injection can provide a viable alternative to current or moo gene transfer approach.

조혈 줄기 세포에의 효과적인 유전자 전달은 유전자 치료의 새로운 가능성을 제시할 수 있다. 레트로바이러스를 이용한 유전자 전달 기술은 많은 기초 연구와 임상 시도가 이루어진 대표적인 바이러스이다. 그러나 현재 사용되고 있는 in vitro에서의 조혈 줄기 세포에의 유전자 도입은 조혈 줄기 세포의 분화 유도, 자기 복제 능력과homing 능력의 저하 등 많은 문제점이 있다. 본 연구는 이러한 문제점을 극복하기 위한 방법으로서 마우스의 대퇴골에 직접 레트로바이러스를 이식하는 IBM (Intra-Bone Marrow) 방법을 이용하여 조혈 줄기 세포에의 효과적인 유전자 도입을 시도하였다. IBM 이식 2주 후 마우스의 각 조직을 분석한 결과, 골수뿐 아니라 림파절, 비장, 간장 세포 등에서 유전자가 안정적으로 발현하는 것을 관찰하였다. 또한, $6.4{\pm}2.7%$의 골수조직 존재 조혈줄기/전구세포에서 도입된 유전자가 안정적으로 발현하고 있는 사실을 확인하였다. 본 연구의 결과를 바탕으로 IBM 이식 방법을 이용한 생체 조직 내 레트로바이러스의 유전자 도입은 조혈 줄기 세포를 이용한 유전자 치료에 매우 효과적인 방법이라는 사실을 시사해주고 있다.

Keywords

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