Mesenchymal stem cells:: Isolation, characterisation and in vivo fluorescent dye tracking
Mesenchymal stem cells:: Isolation, characterisation and in vivo fluorescent dye tracking
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DOI：
10.1016/j.hlc.2008.01.006
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发表时间：
2008-10-01
影响因子：
2.6
通讯作者：
Ward, Christopher
中科院分区：
文献类型：
--
作者：
Weir, Christopher;Morel-Kopp, Marie-Christine;Ward, Christopher
细胞疗法已被用于通过直接心肌输送、冠状动脉输注和表面附着支架来再生心脏。具有向心肌细胞和其他细胞系分化能力的多能间充质干细胞（MSC）已在啮齿动物中进行了大量试验。然而，为了将基础研究转化为新的、安全的再生疗法，越来越需要大型动物模型。了解细胞治疗在哺乳动物心脏中的作用模式一直受到细胞追踪能力的限制。本研究考察了利用不同的荧光染料追踪绵羊同种异体间充质干细胞命运的能力。从羊骨髓中分离的间充质干细胞经提取和流式细胞术鉴定后培养。在用荧光跟踪染料（例如CFSE和DiI）标记后，对细胞进行长达六周的体外和体内信号测试。研究了标记对细胞分裂和分化的影响。一些染料失去了荧光，减慢了细胞分裂。然而，巯基活性染料CM-DiI在注射到羊骨骼肌后6周和植入MSC涂层生物材料支架后2周，在标记的MSC中显示出可检测的体内荧光。CM-DiI标记的MSC体外分化在四周内显示标签保留。荧光膜染料CM-DiI跟踪植入的绵羊间充质干细胞，并提供传统细胞标记物（如基因修饰的GFP）的替代选择。
Cell therapies have been used to regenerate the heart by direct myocardial delivery, by coronary infusion and by surface attached scaffolds. Multipotent mesenchymal stem cells (MSC) with capacity to differentiate into cardiomyocytes and other cell lines have been predominantly trialled in rodents. However, large animal models are increasingly needed to translate basic research into new, safe regenerative therapies. Understanding the mode of action of cell therapies in the mammalian heart has been limited by cell tracking capability. This study examined the ability to track the fate of allogeneic MSC in sheep using various fluorescent dyes.MSC isolated from sheep bone marrow were grown in culture following extraction and flow cytometric characterisation. After labelling with fluorescent tracking dyes (e.g. CFSE and DiI) cells were tested for in vitro and in vivo signal up to six weeks. Labelling effect on cell division and differentiation was studied.Several dyes lost fluorescence and slowed cell division. However, the thiol reactive dye CM-DiI showed detectable in vivo fluorescence in labelled MSC six weeks after injection into sheep skeletal muscle and two weeks after implantation of an MSC coated biomaterial scaffold. CM-DiI labelled MSC differentiated in vitro showed label retention over four weeks.The fluorescent membrane dye CM-DiI tracks implanted sheep MSC and provides an alternative to traditional cell markers such as gene modified GFP.