Investigative Magnetic Resonance Imaging

Korean Society of Magnetic Resonance in Medicine (대한자기공명의과학회)
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$In$ $vitro$ MRI and Characterization of Rat Mesenchymal Stem Cells Transduced with Ferritin as MR Reporter Gene

페리틴 리포터 유전자를 발현하는 백서 중간엽 줄기세포의 특성과 자기공명영상 연구

  • Shin, Cheong-Il (Department of Radiology, Seoul National University Hospital) ;
  • Lee, Whal (Department of Radiology, Seoul National University Hospital) ;
  • Woo, Ji-Su (Department of Radiology, Seoul National University Hospital) ;
  • Park, Eun-Ah (Department of Radiology, Seoul National University Hospital) ;
  • Kim, Pan-Ki (SNU-Duke Cardiovascular MR Research Center, Seoul National University) ;
  • Song, Hyun-Bok (SNU-Duke Cardiovascular MR Research Center, Seoul National University) ;
  • Kim, Hoe-Suk (Department of Radiology, Seoul National University Hospital)
  • 신청일 (서울대학교병원 영상의학과) ;
  • 이활 (서울대학교병원 영상의학과) ;
  • 우지수 (서울대학교병원 영상의학과) ;
  • 박은아 (서울대학교병원 영상의학과) ;
  • 김판기 (서울대학교 서울대-듀크 심혈관 자기공명영상 연구센터) ;
  • 송현복 (서울대학교 서울대-듀크 심혈관 자기공명영상 연구센터) ;
  • 김회숙 (서울대학교병원 영상의학과)
  • Received : 2012.04.01
  • Accepted : 2012.04.25
  • Published : 2012.04.30
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Abstract

Purpose : This study was performed to evaluate the characteristics of rat mesenchymal stem cells (RMSCs) transduced with human ferritin gene and investigate $in$ $vitro$ MRI detectability of ferritin-transduced RMSCs. Materials and Methods: The RMSCs expressing both myc-tagged human ferritin heavy chain subunit (myc-FTH) and green fluorescence protein (GFP) were transduced with lentiviurs. Transduced cells were sorted by GFP expression using a fluorescence-activated cell sorter. Myc-FTH and GFP expression in transduced cells were detected by immunofluorescence staining. The cell proliferative ability and viability were assessed by MTT assay. The RMSC surface markers (CD29+/CD45-) were analyzed by flow cytometry. The intracellular iron amount was measured spectrophotometically and the presence of ferritin-iron accumulation was detected by Prussian blue staining. $In$ $vitro$ magnetic resonance imaging (MRI) study of cell phantoms was done on 9.4 T MR scanner to evaluate the feasibility of imaging the ferritin-transduced RMSCs. Results: The myc-FTH and GFP genes were stably transduced into RMSCs. No significant differences were observed in terms of biologic properties in transduced RMSCs compared with non-transduced RMSCs. Ferritin-transduced RMSCs exhibited increased iron accumulation ability and showed significantly lower $T_2$ relaxation time than non-transduced RMSCs. Conclusion: Ferritin gene as MR reporter gene could be used for non-invasive tracking and visualization of therapeutic mesenchymal stem cells by MRI.

목적: 백서 중간엽 줄기세포에 페리틴 유전자를 형질 도입시켜 생물학적 특성의 변화 유무를 평가하고, 자기공명영상에서 신호강도의 차이를 확인해보고자 하였다. 대상과 방법: 백서 중간엽 줄기세포에 렌티바이러스를 이용하여 사람유래 재조합 페리틴과 녹색형광단백질 유전자의 과발현을 유도하였다. 페리틴 유전자가 발현된 백서 중간엽 줄기세포의 증식성과 생존능을 분석하기 위해 MTT 어세이를 수행하였으며, 유세포 분석을 수행하여 중간엽 줄기세포의 표면 마커 발현을 평가하고, 세포 내 철 함량을 측정하고 프러시안 블루 염색을 시행하여 철 축적능력을 분석하였다. 세포 팬텀을 이용하여 9.4 T 자기공영영상 기기를 이용하여 검출가능성을 평가하였다. 결과: 페리틴과 녹색형광 유전자는 백서 중간엽 줄기세포에서 안정적으로 발현되었다. 페리틴 유전자의 과발현으로 인해 백서 중간엽 줄기세포의 생물학적 특성 (증식능력, 생존능, 표면마커)은 영향을 받지 않았다. 페리틴을 발현하는 중간엽 줄기세포에서 철의 축적능력이 증가된 것이 확인되었고, T2 이완 시간은 유의하게 감소하였다. 결론: 줄기세포 치료 연구에서 자기공명 리포터 유전자 페리틴은 자기공명영상법을 이용하여 중간엽 줄기세포를 비침습적으로 가시화 할 수 있고 이를 이용하여 생체추적이 가능할 것으로 기대된다.

Keywords

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