Investigative Magnetic Resonance Imaging

  • 제16권1호
  • /
  • Pages.47-54
  • /
  • 2012
  • /
  • 2384-1095(pISSN)
  • /
  • 2384-1109(eISSN)
대한자기공명의과학회 (Korean Society of Magnetic Resonance in Medicine)
권호 표지

페리틴 리포터 유전자를 발현하는 백서 중간엽 줄기세포의 특성과 자기공명영상 연구

$In$ $vitro$ MRI and Characterization of Rat Mesenchymal Stem Cells Transduced with Ferritin as MR Reporter Gene

  • 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)
  • 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)
  • 투고 : 2012.04.01
  • 심사 : 2012.04.25
  • 발행 : 2012.04.30
PDF 다운로드
⟨ 이전 논문 다음 논문 ⟩

초록

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.

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.

키워드

참고문헌

  1. Cohen B, Ziv K, Plaks V, et al. MRI detection of transcriptional regulation of gene expression in transgenic mice. Nature Medicine 2007;13:498-503
  2. Kim HS, Cho HR, Choi SH, Woo JS, Moon WK. In vivo imaging of tumor transduced with bimodal lentiviral vector encoding human ferritin and green fluorescent protein on a 1.5T clinical magnetic resonance scanner. Cancer Research 2010;70:7315-24
  3. Genove G, DeMarco U, Xu H, Goins WF, Ahrens ET. A new transgene reporter for in vivo magnetic resonance imaging. Nature Medicine 2005;11:450-454
  4. Aung W, Hasegawa S, Koshikawa-Yano M, et al. Visualization of in vivo electroporation-mediated transgene expression in experimental tumors by optical and magnetic resonance imaging. Gene Therapy 2009;16:830-839
  5. Cohen B, Dafni H, Meir G, Harmelin A, Neeman M. Ferritin as an endogenous MRI reporter for noninvasive imaging of gene expression in C6 glioma tumors. Neoplasia 2005;7:109-117
  6. Liu J, Cheng EC, Long RC, et al. Noninvasive monitoring of embryonic stem cells in vivo with MRI transgene reporter. Tissue engineering Part C, Methods 2009;15:739-747
  7. Kim YJ, Huh Y-M, Choe KO, et al. In vivo magnetic resonance imaging of injected mesenchymal stem cells in rat myocardial infarction; simultaneous cell tracking and left ventricular function measurement. International Journal of Cardiovascular Imaging 2009;25:99-109
  8. Agudelo CA, Tachibana Y, Noboru T, Iida H, Yamaoka T. Long-term in vivo magnetic resonance imaging tracking of endothelial progenitor cells transplanted in rat ischemic limbs and their angiogenic potential. Tissue engineering Part A 2011;17:2079-89
  9. Agudelo CA, Tachibana Y, Hurtado AF, Ose T, Iida H, Yamaoka T. The use of magnetic resonance cell tracking to monitor endothelial progenitor cells in a rat hindlimb ischemic model. Biomaterials 2012;33:2439-48
  10. Campan M, Lionetti V, Aquaro GD, et al. Ferritin as a reporter gene for in vivo tracking of stem cells by 1.5-T cardiac MRI in a rat model of myocardial infarction. Am J Physiol Heart Circ Physiol 2011;300:H2238-2250
  11. Li Y, Zhang D, Zhang Y, He G, Zhang F. Augmentation of neovascularization in murine hindlimb ischemia by combined therapy with simvastatin and bone marrow-derived mesenchymal stem cells transplantation. J Biomed Sci 2010;17:75
  12. Laurila JP, Laatikainen L, Castellone MD, et al. Human embryonic stem cell-derived mesenchymal stromal cell transplantation in a rat hind limb injury model. Cytotherapy 2009;11:726-37
  13. Iwase T, Nagaya N, Fujii T, et al. Comparison of angiogenic potency between mesenchymal stem cells and mononuclear cells in a rat model of hindlimb ischemia. Cardiovasc Res 2005; 66:543-51