Korean J Radiol.  2009 Dec;10(6):613-622. 10.3348/kjr.2009.10.6.613.

Dual-Modal Nanoprobes for Imaging of Mesenchymal Stem Cell Transplant by MRI and Fluorescence Imaging

• Sung CK
• Hong KA
• Lin S
• Lee Y
• Cha J
• Lee JK
• Hong CP
• Han BS
• Jung SI
• Kim SH
• Yoon KS

• KMID: 1102565
• DOI: http://doi.org/10.3348/kjr.2009.10.6.613

Abstract

OBJECTIVE
To determine the feasibility of labeling human mesenchymal stem cells (hMSCs) with bifunctional nanoparticles and assessing their potential as imaging probes in the monitoring of hMSC transplantation. MATERIALS AND METHODS: The T1 and T2 relaxivities of the nanoparticles (MNP@SiO2[RITC]-PEG) were measured at 1.5T and 3T magnetic resonance scanner. Using hMSCs and the nanoparticles, labeling efficiency, toxicity, and proliferation were assessed. Confocal laser scanning microscopy and transmission electron microscopy were used to specify the intracellular localization of the endocytosed iron nanoparticles. We also observed in vitro and in vivo visualization of the labeled hMSCs with a 3T MR scanner and optical imaging. RESULTS: MNP@SiO2(RITC)-PEG showed both superparamagnetic and fluorescent properties. The r1 and r2 relaxivity values of the MNP@SiO2(RITC)-PEG were 0.33 and 398 mM-1 s-1 at 1.5T, respectively, and 0.29 and 453 mM-1 s-1 at 3T, respectively. The effective internalization of MNP@SiO2(RITC)-PEG into hMSCs was observed by confocal laser scanning fluorescence microscopy. The transmission electron microscopy images showed that MNP@SiO2(RITC)-PEG was internalized into the cells and mainly resided in the cytoplasm. The viability and proliferation of MNP@SiO2(RITC)-PEG-labeled hMSCs were not significantly different from the control cells. MNP@SiO2(RITC)-PEG-labeled hMSCs were observed in vitro and in vivo with optical and MR imaging. CONCLUSION: MNP@SiO2(RITC)-PEG can be a useful contrast agent for stem cell imaging, which is suitable for a bimodal detection by MRI and optical imaging.

Keyword

Stem cells; Nanoparticles; Magnetic resonance (MR); Optical imaging, contrast agent

MeSH Terms

Animals
Biocompatible Materials
Cells, Cultured
Cobalt
Feasibility Studies
Ferric Compounds
Humans
Magnetic Resonance Imaging/*methods
*Mesenchymal Stem Cells
Mice
Mice, Nude
Microscopy, Confocal
Microscopy, Electron
Nanoparticles/*chemistry
Phantoms, Imaging
Polyethylene Glycols
Rats
Rhodamines
Silicon Dioxide
Staining and Labeling/methods

Figure

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  Fig. 1 Transverse relaxation rate (R2) versus MNP@SiO2(RITC)-PEG concentration at 1.5T and 3T.

• 

  Fig. 2 Cellular distribution of MNP@SiO2(RITC)-PEG in human mesenchymal stem cells. Confocal fluorescence images of human mesenchymal stem cells cultured with different concentrations of MNP@SiO2(RITC)-PEG (0, 0.5, 1, 2 and 4 mg/ml) for 2, 4, and 6 hours.

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  Fig. 3 Transmission electron microscopy images of human mesenchymal stem cells embedded in MNP@SiO2(RITC)-PEG. Human mesenchymal stem cells were treated with 4 mg of MNP@SiO2 (RITC)-PEG/ml medium for four hours, which was then processed for transmission electron microscopy study. MNP@SiO2 (RITC)-PEG (arrows) exhibited after internalization into cells. MNP@SiO2 (RITC)-PEG-treated cell (×10,000, 20,000). N = nucleus, M = mitochondria

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  Fig. 4 Cell viability. Human mesenchymal stem cells were incubated for 2 (⋄), 4, (■) and 6 hours (▵) with different magnetic nanoparticle concentrations (0, 0.5, 1, 2, 4 mg/ml) after 0 (A), 2 (B), and 4 days (C).

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  Fig. 5 Proliferation index of human mesenchymal stem cells, which were incubated for four hours with 4 mg/ml MNP@SiO2(RITC)-PEG is shown (■). Control (⋄).

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  Fig. 6 Experimental MRI of phantom of labeled human mesenchymal stem cells placed in 4% phantom gelatin in plastic wells. Coronal FIESTA (GE, b-SSFP) sequence image of phantom shows marked decrease in signal intensity with increasing magnetic nanoparticle concentration. A. 5×105 cells in 4% gelatin, B. 1×105 cells in 4% gelatin. 1 = gelatin, 2 = unlabeled hMSCs in 4% gelatin, 3-6 = labeled hMSCs with 2-hour incubation; 7-10 = 4-hour incubation, 11-14 = 6-hour incubation. 3, 7, 11 = labeled human mesenchymal stem cells at magnetic nanoparticle concentration of 0.5 mg/ml, 4, 8, 12, at 1 mg/ml; 5, 9, 13, at 2 mg/ml; 6, 10 ,14, at 4 mg/ml.

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  Fig. 7 Sequential in vivo MR imaging (3T, 3.0 mm slices) and fluorescence imaging of SD rat after injection with MNP@SiO2(RITC)-PEG-labeled human mesenchymal stem cells. A-D. MR images with fast spin echo (TR/TE: 3,500/89.8) (A), 3D fast imaging employing steady state acquisition (TR/TE/flip angle: 8.4/4.1/35) (B), multiple gradient-recalled echo (TR/TE/flip angle: 800/13.4/20) (C), and fluorescent signal coming from labeled human mesenchymal stem cells in kidney of SD rat (arrows) can only be detected in excised state (D).

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  Fig. 8 In vivo MR imaging (A) and fluorescence imaging (B) of nude mouse after subcutaneous injection of labeled and unlabeled human mesenchymal stem cells with MNP@SiO2(RITC)-PEG (1 = 1×105 unlabeled human mesenchymal stem cells for control, 2 = 1×106 labeled human mesenchymal stem cells, 3 = 1×105 labeled human mesenchymal stem cells). Labeled human mesenchymal stem cells are clearly seen as dark dot (arrow) in subcutaneous layer of nude mouse on axial scan of fast spin echo sequence. Fluorescent signal of subcutaneously injected human mesenchymal stem cells is detected at injection sites of labeled cells. Injection sites of unlabeled cells shows autofluorescence-induced artifact.

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