Oude Engberink RD, van der Pol SM, Dopp EA, de Vries HE, Blezer ELComparison of SPIO and USPIO for in vitro labeling of human monocytes: MR detection and cell function. Radiology 243:467-474

Department of Molecular Cell Biology and Immunology, VU University Amsterdam, Amsterdamo, North Holland, Netherlands
Radiology (Impact Factor: 6.87). 05/2007; 243(2):467-74. DOI: 10.1148/radiol.2432060120
Source: PubMed


To label human monocytes with superparamagnetic iron oxide (SPIO) and compare labeling efficiency with that of ultrasmall SPIO (USPIO) and evaluate the effect of iron incorporation on cell viability, migratory capacity, and proinflammatory cytokine production.
The study was approved by the institutional ethics committee; informed consent was obtained from donors. Freshly isolated human monocytes were labeled with iron particles of two sizes, USPIOs of 30 nm and SPIOs of 150 nm, for 1.5 hours in culture medium containing 0.1, 0.5, 1.0, and 3.7 mg of iron per milliliter. Labeling efficiency was determined with relaxation time magnetic resonance (MR) imaging (4.7 T) and Prussian blue staining for presence of intracellular iron. Cell viability was monitored; migratory capacity of monocytes after labeling was evaluated by using an in vitro assay with monolayers of brain endothelial cells. Levels of proinflammatory cytokines, interleukin (IL) 1 and IL-6, were measured with enzyme-linked immunosorbent assay 24 hours after labeling. Data were analyzed with Student t test or two-way analysis of variance followed by a multiple-comparison procedure.
R2 relaxation rates increased for cell samples incubated with SPIOs, whereas rates were not affected for samples incubated with highest concentration of USPIOs. Labeling monocytes with SPIOs (1.0 mg Fe/mL) resulted in an R2 of 13.1 sec(-1) +/- 0.8 (standard error of the mean) (7 sec(-1) +/- 0.2 for vehicle-treated cells, P < .05) and had no effect on cell viability. On the basis of T2 relaxation times, the in vitro MR detection limit of 58 labeled monocytes per 0.05 microL was calculated. Migration of labeled monocytes was not different from that of vehicle-treated cells. Intracellular iron had no effect on production of IL-1 and IL-6 24 hours after labeling.
In vitro labeling of human monocytes is effective by using SPIOs, not USPIOs. Incubation with SPIOs (1.0 mg Fe/mL) results in efficient labeling detectable on MR images and does not affect cellular viability and activation markers such as cell migration and cytokine production.

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    • "One key attribute of SPIO particles is that they are biologically inert and believed to safely degrade via normal iron recycling pathways when released from dying cells [16]. Another beneficial feature is that cells can be heavily-labeled with SPIOs and remain viable without affecting their proliferative capacity [17]. On the other hand, the intracellular concentration of SPIO particles can be diluted by cell division resulting in eventual loss of MRI signal [18]. "
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    ABSTRACT: The purpose of this study was to determine the ability of superparamagnetic iron oxide (SPIO) nanoparticles to function as a long-term tracking label for multi-modal imaging of implanted engineered tissues containing muscle-derived progenitor cells using magnetic resonance imaging (MRI) and X-ray micro-computed tomography (μCT). SPIO-labeled primary myoblasts were embedded in fibrin sealant and imaged to obtain intensity data by MRI or radio-opacity information by μCT. Each imaging modality displayed a detection gradient that matched increasing SPIO concentrations. Labeled cells were then incorporated in fibrin sealant, injected into the atrioventricular groove of rat hearts, and imaged in vivo and ex vivo for up to 1 year. Transplanted cells were identified in intact animals and isolated hearts using both imaging modalities. MRI was better able to detect minuscule amounts of SPIO nanoparticles, while μCT more precisely identified the location of heavily-labeled cells. Histological analyses confirmed that iron oxide particles were confined to viable, skeletal muscle-derived cells in the implant at the expected location based on MRI and μCT. These analyses showed no evidence of phagocytosis of labeled cells by macrophages or release of nanoparticles from transplanted cells. In conclusion, we established that SPIO nanoparticles function as a sensitive and specific long-term label for MRI and μCT, respectively. Our findings will enable investigators interested in regenerative therapies to non-invasively and serially acquire complementary, high-resolution images of transplanted cells for one year using a single label.
    Full-text · Article · Sep 2014 · PLoS ONE
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    • "Iron oxide MNP with total size of 100 nm were preferred in this study as MNP in this range have been shown to be more effective for in vitro monocyte / macrophage labeling compared to smaller nanoparti - cles size ( Oude Engberink et al . 2007 ) . Zeta potential analysis has revealed that SPIO , SPIO – PEG , and SPIO – PEG – COOH have a surface charge of - 1 . 8 , - 3 . 3 , and - 5 . 8 mV , respectively . Ferrozine - based spectro - photometry ( Fig . 1 ) revealed higher uptake of iron by M2 macrophages compared to M1 subsets for the different SPIO nanoparticles ( i . e . , a"
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    ABSTRACT: Noninvasive imaging of macrophages activity has raised increasing interest for diagnosis of different diseases, which make them attractive vehicles to deliver contrast agents or drugs for diagnostic or therapeutic purposes. In this study, the effect of polyethylene glycol functionalization of magnetic iron oxide nanoparticles and their further surface modification with carboxylic groups on bone marrow-derived M1 and M2 macrophages phenotype, labeling efficiency, uptake mechanism, biocompatibility, and their in vivo MR detection was assessed. An enhanced labeling efficiency was observed for carboxylic surface-modified superparamagnetic iron oxide (SPIO) compared to PEGylated SPIO and to a higher extent to plain SPIO along with a higher uptake by M2 subsets. Magnetic nanoparticles were found located in the periphery of the vesicles dispersed in the cytoplasm in TEM. Investigation of the labeling mechanism by inhibiting different uptake pathways revealed that endocytosis via scavenger receptor A, a process known to be clathrin mediated, plays a central role in the cellular uptake kinetics of both macrophages subpopulations. Biocompatibility evaluation showed no variation in cell viability and mitochondrial membrane potential with a low release of ROS. Flow cytometry and measurement of iNOS and Arginase 1 activity as marker of M1 and M2 macrophages polarization confirmed that magnetic labeling of macrophages subsets did not affect their polarization. In addition, no variation was observed in the biodistribution of magnetic iron oxide-labeled M1 and M2 macrophages subsets when monitored using noninvasive magnetic resonance imaging with a better detection for the enhanced SPIO–PEG–COOH-labeled cells.
    Full-text · Article · Jul 2013 · Journal of Nanoparticle Research
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    • "As demonstrated in the present study, a longer incubation time is needed for optimal labeling with USPIO particles. For labeling of phagocytic cells though, SPIO particles might be more suitable, as SPIO particles are easily recognized and internalized into monocytes and macrophages [37]. Another advantage of USPIO particles is that they have longer plasmatic half-life (>36 hours) and this allows for longer cell tracking observation periods [25]. "
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    ABSTRACT: Background. Regenerative therapy is an emerging treatment modality. To determine migration and retention of implanted cells, it is crucial to develop noninvasive tracking methods. The aim was to determine ex vivo magnetic resonance imaging (MRI) detection limits of ultrasmall superparamagnetic iron-oxide (USPIO) labeled mesenchymal stromal cells (MSCs). Materials and Methods. 248 gel-phantoms were constructed and scanned on a 1.5T MRI-scanner. Phantoms contained human MSCs preincubated with USPIO nanoparticles for 2, 6, or 21 hours using 5 or 10 μ g USPIO/10(5) MSCs. In addition, porcine hearts were scanned after injection of USPIO labeled MSCs. Results. Using 21 h incubation time and 10 μ g USPIO/10(5) MSCs, labeled cells were clearly separated from unlabeled cells on MRI using 250.000 (P < 0.001), 500.000 (P = 0.007), and 1.000.000 MSCs (P = 0.008). At lower incubation times and doses, neither labeled nor unlabeled cells could be separated. In porcine hearts labeled, but not unlabeled, MSCs were identified on MRI. Conclusions. As few as 250.000 MSCs can be detected on MRI using 21 h incubation time and 10 μ g USPIO/10(5) MSCs. At lower incubation times and doses, several million cells are needed for MRI detection. USPIO labeled cells can be visualized by MRI in porcine myocardial tissue.
    Full-text · Article · Mar 2013 · Stem cell International
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