Labeling Stem Cells with Ferumoxytol, an FDA-Approved Iron Oxide Nanoparticle
Labeling Stem Cells with Ferumoxytol, an FDA-Approved Iron Oxide Nanoparticle
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DOI：
10.3791/3482
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发表时间：
2011-11-01
影响因子：
1.2
通讯作者：
Daldrup-Link, Heike E.
中科院分区：
文献类型：
--
作者：
Castaneda, Rosalinda T.;Khurana, Aman;Daldrup-Link, Heike E.
基于干细胞的疗法为再生医学领域提供了巨大的潜力。然而，关于移植细胞的体内动力学仍有许多待了解。一种在体内重复监测移植干细胞的非侵入性方法将使研究人员能够直接监测干细胞移植并确定成功或不成功的植入结果。人们继续研究各种干细胞的无数应用。该协议重点关注 3 种不同的干细胞群：人胚胎肾 293 (HEK293) 细胞、人间充质干细胞 (hMSC) 和诱导多能干 (iPS) 细胞。 HEK 293 细胞源自在含有剪切腺病毒 5 DNA 的培养物中生长的人胚胎肾细胞。这些细胞因其易于培养、生长迅速且易于转染而被广泛用于研究。 hMSCs 存在于成人骨髓中。这些细胞可以作为未分化细胞进行复制，同时保持多能性或分化为有限数量的细胞命运的潜力。 hMSC 可以分化为间充质组织谱系，包括成骨细胞、脂肪细胞、软骨细胞、肌腱、肌肉和骨髓基质。 iPS 细胞是经过基因重编程的成体细胞，经过修饰可表达类似于胚胎干细胞特性的基因和因子。这些细胞具有多能性，这意味着它们具有分化成所有细胞谱系的能力 (1)。 hMSC 和 iPS 细胞均已表现出体内组织再生能力。事实证明，磁共振 (MR) 成像与超顺磁性氧化铁 (SPIO) 纳米粒子细胞标记的使用对于干细胞的体内跟踪非常有效，因为 SPIO 纳米粒子的 MR 成像具有接近微观的解剖分辨率、较长的血液半衰期（允许纵向成像）以及细胞检测的高灵敏度 (2-4)。此外，使用 SPIO 的 MR 成像具有临床可转化性。 SPIO 由氧化铁核心和葡聚糖、羧基葡聚糖或淀粉表面涂层组成，用于包含血浆成分中的生物反应性铁核心。这些试剂会产生局部磁场不均匀性，导致 T2 加权 MR 图像上的信号减弱 (5)。不幸的是，SPIO 已不再生产。然而，第二代超小型 SPIO (USPIO) 提供了一种可行的替代方案。 Ferumoxytol (FerahemeTM) 是一种 USPIO，由非化学计量的磁铁矿核心和聚葡萄糖山梨醇羧甲基醚涂层组成。通过光散射测定，ferumoxytol 的胶体颗粒尺寸为 17-30 nm。分子量为750 kDa，2T MRI 场的弛豫常数为58.609 mM-1 sec-1 强度(4)。 Ferumoxytol 最近被 FDA 批准作为铁补充剂，用于治疗肾衰竭患者的铁缺乏症 (6)。我们的小组已将该试剂应用于细胞标记应用的“标签外”用途。我们的技术证明了用 ferumoxytol 可以有效标记干细胞，从而导致标记细胞在 MR 图像上产生显着的 MR 信号效应。该技术可应用于临床前和临床环境中干细胞治疗的非侵入性监测。
Stem cell based therapies offer significant potential for the field of regenerative medicine. However, much remains to be understood regarding the in vivo kinetics of transplanted cells. A non-invasive method to repetitively monitor transplanted stem cells in vivo would allow investigators to directly monitor stem cell transplants and identify successful or unsuccessful engraftment outcomes.A wide range of stem cells continues to be investigated for countless applications. This protocol focuses on 3 different stem cell populations: human embryonic kidney 293 (HEK293) cells, human mesenchymal stem cells (hMSC) and induced pluripotent stem (iPS) cells. HEK 293 cells are derived from human embryonic kidney cells grown in culture with sheared adenovirus 5 DNA. These cells are widely used in research because they are easily cultured, grow quickly and are easily transfected. hMSCs are found in adult marrow. These cells can be replicated as undifferentiated cells while maintaining multipotency or the potential to differentiate into a limited number of cell fates. hMSCs can differentiate to lineages of mesenchymal tissues, including osteoblasts, adipocytes, chondrocytes, tendon, muscle, and marrow stroma. iPS cells are genetically reprogrammed adult cells that have been modified to express genes and factors similar to defining properties of embryonic stem cells. These cells are pluripotent meaning they have the capacity to differentiate into all cell lineages (1). Both hMSCs and iPS cells have demonstrated tissue regenerative capacity in-vivo.Magnetic resonance (MR) imaging together with the use of superparamagnetic iron oxide (SPIO) nanoparticle cell labels have proven effective for in vivo tracking of stem cells due to the near microscopic anatomical resolution, a longer blood half-life that permits longitudinal imaging and the high sensitivity for cell detection provided by MR imaging of SPIO nanoparticles (2-4). In addition, MR imaging with the use of SPIOs is clinically translatable. SPIOs are composed of an iron oxide core with a dextran, carboxydextran or starch surface coat that serves to contain the bioreactive iron core from plasma components. These agents create local magnetic field inhomogeneities that lead to a decreased signal on T2-weighted MR images (5). Unfortunately, SPIOs are no longer being manufactured. Second generation, ultrasmall SPIOs (USPIO), however, offer a viable alternative. Ferumoxytol (FerahemeTM) is one USPIO composed of a non-stoichiometric magnetite core surrounded by a polyglucose sorbitol carboxymethylether coat. The colloidal, particle size of ferumoxytol is 17-30 nm as determined by light scattering. The molecular weight is 750 kDa, and the relaxivity constant at 2T MRI field is 58.609 mM-1 sec-1 strength(4). Ferumoxytol was recently FDA-approved as an iron supplement for treatment of iron deficiency in patients with renal failure (6). Our group has applied this agent in an "off label" use for cell labeling applications. Our technique demonstrates efficient labeling of stem cells with ferumoxytol that leads to significant MR signal effects of labeled cells on MR images. This technique may be applied for non-invasive monitoring of stem cell therapies in pre-clinical and clinical settings.