Molecular and Magnetic Resonance Imaging of Human Embryonic Stem Cell-Derived Neural Stem Cell Grafts in Ischemic Rat Brain

Department of Neurosurgery, Stanford Stroke Center, Stanford University School of Medicine, California 94305-5487, USA.
Molecular Therapy (Impact Factor: 6.23). 06/2009; 17(7):1282-91. DOI: 10.1038/mt.2009.104
Source: PubMed


Real-time imaging of transplanted stem cells is essential for understanding their interactions in vivo with host environments, for tracking cell fate and function and for successful delivery and safety monitoring in the clinical setting. In this study, we used bioluminescence (BLI) and magnetic resonance imaging (MRI) to visualize the fate of grafted human embryonic stem cell (hESC)-derived human neural stem cells (hNSCs) in stroke-damaged rat brain. The hNSCs were genetically engineered with a lentiviral vector carrying a double fusion (DF) reporter gene that stably expressed enhanced green fluorescence protein (eGFP) and firefly luciferase (fLuc) reporter genes. The hNSCs were self-renewable, multipotent, and expressed markers for neural stem cells. Cell survival was tracked noninvasively by MRI and BLI for 2 months after transplantation and confirmed histologically. Electrophysiological recording from grafted GFP(+) cells and immuno-electronmicroscopy demonstrated connectivity. Grafted hNSCs differentiated into neurons, into oligodendrocytes in stroke regions undergoing remyelination and into astrocytes extending processes toward stroke-damaged vasculatures. Our data suggest that the combination of BLI and MRI modalities provides reliable real-time monitoring of cell fate.

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    • "Recently, brain organoids derived from induced PSCs have also been shown to provide neural disease models for pathological study [22]. For these applications, in vitro and in vivo tracking of PSC-derived NPCs using MRI can provide valuable information toward the clinical translations [3]. NPCs are usually generated from PSCs through the formation of aggregate structure known as embryoid bodies (EBs) [23]. "
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    ABSTRACT: Background aims: Pluripotent stem cell (PSC)-derived neural progenitor cells (NPCs) represent an unlimited source for the treatment of various neurological disorders. NPCs are usually derived from PSCs through the formation of embryoid body (EB), an aggregate structure mimicking embryonic development. This study investigated the effect of labeling multicellular EB-NPC aggregates with micron-sized particles of iron oxide (MPIO) for cell tracking using magnetic resonance imaging (MRI). Methods: Intact and dissociated EB-NPC aggregates were labeled with various concentrations of MPIOs (0, 2.5, 5 and 10 μg Fe/mL). The labeled cells were analyzed by fluorescent imaging, flow cytometry and in vitro MRI for labeling efficiency and detectability. Moreover, the biological effects of intracellular MPIO on cell viability, cytotoxicity, proliferation and neural differentiation were evaluated. Results: Intact EB-NPC aggregates showed higher cell proliferation and viability compared with the dissociated cells. Despite diffusion limitation at low MPIO concentration, higher concentration of MPIO (i.e., 10 μg Fe/mL) was able to label EB-NPC aggregates at similar efficiency to the single cells. In vitro MRI showed concentration-dependent MPIO detection in EB-NPCs over 2.0-2.6 population doublings. More important, MPIO incorporation did not affect the proliferation and neural differentiation of EB-NPCs. Conclusions: Multicellular EB-NPC aggregates can be efficiently labeled and tracked with MPIO while maintaining cell proliferation, phenotype and neural differentiation potential. This study demonstrated the feasibility of labeling EB-NPC aggregates with MPIO for cellular monitoring of in vitro cultures and in vivo transplantation.
    Cytotherapy 01/2015; 17(1). DOI:10.1016/j.jcyt.2014.09.008 · 3.29 Impact Factor
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    • "Another multimodal approach is the combination of MRI with BLI to characterize transplanted NSCs in the stroke pathology. Firefly bioluminescence was used as a viability marker while transplant location was visualized with iron oxide labeling by MRI for 8 weeks (Daadi et al., 2009). Sequential MRI/BLI was successfully employed to investigate the amount and distribution of intra-arterially (i.a.) versus intra-venously (i.v.) injected stem cells in an ischemia model, showing brain accumulation after i.a. but not after i.v. "
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    ABSTRACT: Transplanted stem cells can induce and enhance functional recovery in experimental stroke. Invasive analysis has been extensively used to provide detailed cellular and molecular characterization of the stroke pathology and engrafted stem cells. But post mortem analysis is not appropriate to reveal the time scale of the dynamic interplay between the cell graft, the ischemic lesion and the endogenous repair mechanisms. This review describes non-invasive imaging techniques which have been developed to provide complementary in vivo information. Recent advances were made in analyzing simultaneously different aspects of the cell graft (e.g., number of cells, viability state, and cell fate), the ischemic lesion (e.g., blood-brain-barrier consistency, hypoxic, and necrotic areas) and the neuronal and vascular network. We focus on optical methods, which permit simple animal preparation, repetitive experimental conditions, relatively medium-cost instrumentation and are performed under mild anesthesia, thus nearly under physiological conditions. A selection of recent examples of optical intrinsic imaging, fluorescence imaging and bioluminescence imaging to characterize the stroke pathology and engrafted stem cells are discussed. Special attention is paid to novel optimal reporter genes/probes for genetic labeling and tracking of stem cells and appropriate transgenic animal models. Requirements, advantages and limitations of these imaging platforms are critically discussed and placed into the context of other non-invasive techniques, e.g., magnetic resonance imaging and positron emission tomography, which can be joined with optical imaging in multimodal approaches.
    Frontiers in Cellular Neuroscience 08/2014; 8:226. DOI:10.3389/fncel.2014.00226 · 4.29 Impact Factor
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    • "Stem cells of various origins can provide behavioral improvements in animal models of stroke (see, e.g., (Lindvall and Kokaia, 2011)) through several mechanisms , such as neuronal replacement (Daadi et al., 2009, 2008), modulation of inflammation (Bacigaluppi et al., 2009; Horie et al., 2011; Lee et al., 2008), neuroprotection (Bacigaluppi et al., 2009), or stimulation of angiogenesis (Chen et al., 2003; Horie et al., 2011; Lee et al., 2008) and brain plasticity (Andres et al., 2011; Daadi et al., 2010; Liu et al., 2008). Ischemic stroke occurs more often in elderly humans (Marini et al., 2001), but most transplantation studies have been performed in adult animals. "
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    ABSTRACT: Purpose: Induced pluripotent stem cells (iPSCs) improve behavior and form neurons after implantation into the stroke-injured adult rodent brain. How the aged brain responds to grafted iPSCs is unknown. We determined survival and differentiation of grafted human fibroblast-derived iPSCs and their ability to improve recovery in aged rats after stroke. Methods: Twenty-four months old rats were subjected to 30 min distal middle cerebral artery occlusion causing neocortical damage. After 48 h, animals were transplanted intracortically with human iPSC-derived long-term neuroepithelial-like stem (hiPSC-lt-NES) cells. Controls were subjected to stroke and were vehicle-injected. Results: Cell-grafted animals performed better than vehicle-injected recipients in cylinder test at 4 and 7 weeks. At 8 weeks, cell proliferation was low (0.7 %) and number of hiPSC-lt-NES cells corresponded to 49.2% of that of implanted cells. Transplanted cells expressed markers of neuroblasts and mature and GABAergic neurons. Cell-grafted rats exhibited less activated microglia/macrophages in injured cortex and neuronal loss was mitigated. Conclusions: Our study provides the first evidence that grafted human iPSCs survive, differentiate to neurons and ameliorate functional deficits in stroke-injured aged brain.
    Restorative neurology and neuroscience 06/2014; 32(4). DOI:10.3233/RNN-140404 · 2.49 Impact Factor
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