Fluorophore-conjugated iron oxide nanoparticle labeling and analysis of engrafting human hematopoietic stein cells
Fluorophore-conjugated iron oxide nanoparticle labeling and analysis of engrafting human hematopoietic stein cells
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DOI：
10.1634/stemcells.2007-0016
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发表时间：
2008-02-01
期刊：
影响因子：
5.2
通讯作者：
Nolta, Jan A.
中科院分区：
文献类型：
--
作者：
Maxwell, Dustin J.;Bonde, Jesper;Nolta, Jan A.
使用纳米级氧化铁颗粒结合分子成像技术，可以动态研究人类造血干细胞（HSC）的归家和运输。确定临床适用的将纳米颗粒装入原始HSC的策略需要严格定义培养条件，以保持活力而不诱导终末分化。在目前的研究中，荧光分子与右旋包覆的氧化铁纳米颗粒（Feridex）共价连接，表征人类HSC标记，以监测植入过程。将荧光团结合到葡聚糖包被层上进行荧光激活细胞分选纯化，消除了来自非隔离纳米颗粒污染物的虚假信号。研究人员开发了一种短期定义的培养策略，可以有效地标记静止和循环的HSC，在体外或体内没有明显的毒性。将纯化的原代人脐带血谱系缺失细胞和CD34（+）细胞移植到免疫缺陷小鼠体内，可以在受体骨骼中检测到标记的人造血干细胞。流式细胞术用于精确定量隔离纳米颗粒的细胞群，并跟踪其移植后的命运。流式细胞术终点分析证实骨髓中存在纳米颗粒标记的人干细胞。使用荧光标记的氧化铁纳米颗粒结合流式细胞术进行荧光成像，可以评估标记效率和确定的人类HSC亚群的归归能力。
The use of nanometer-sized iron oxide particles combined with molecular imaging techniques enables dynamic studies of homing and trafficking of human hematopoietic stem cells (HSC). Identifying clinically applicable strategies for loading nanoparticles into primitive HSC requires strictly defined culture conditions to maintain viability without inducing terminal differentiation. In the current study, fluorescent molecules were covalently linked to dextrancoated iron oxide nanoparticles (Feridex) to characterize human HSC labeling to monitor the engraftment process. Conjugating fluorophores to the dextran coat for fluorescence-activated cell sorting purification eliminated spurious signals from nonsequestered nanoparticle contaminants. A short-term defined incubation strategy was developed that allowed efficient labeling of both quiescent and cycling HSC, with no discernable toxicity in vitro or in vivo. Transplantation of purified primary human cord blood lineage-depleted and CD34(+) cells into immunodeficient mice allowed detection of labeled human HSC in the recipient bones. Flow cytometry was used to precisely quantitate the cell populations that had sequestered the nanoparticles and to follow their fate post-transplantation. Flow cytometry endpoint analysis confirmed the presence of nanoparticle-labeled human stem cells in the marrow. The use of fluorophorelabeled iron oxide nanoparticles for fluorescence imaging in combination with flow cytometry allows evaluation of labeling efficiencies and homing capabilities of defined human HSC subsets.