Tracking stem cells with superparamagnetic iron oxide nanoparticles: perspectives and considerations

Abstract

Superparamagnetic iron oxide nanoparticles (SPIONs) have been used for diagnoses in biomedical applications, due to their unique properties and their apparent safety for humans. In general, SPIONs do not seem to produce cell damage, although their long-term in vivo effects continue to be investigated. The possibility of efficiently labeling cells with these magnetic nanoparticles has stimulated their use to noninvasively track cells by magnetic resonance imaging after transplantation. SPIONs are attracting increasing attention and are one of the preferred methods for cell labeling and tracking in preclinical and clinical studies. For clinical protocol approval of magnetic-labeled cell tracking, it is essential to expand our knowledge of the time course of SPIONs after cell incorporation and transplantation. This review focuses on the recent advances in tracking SPION-labeled stem cells, analyzing the possibilities and limitations of their use, not only focusing on myocardial infarction but also discussing other models.

Keywords: cell tracking; in vivo imaging; myocardial infarction; nanoparticles; stem cells; superparamagnetic iron oxide nanoparticles.

Figure 1

Mesenchymal stem cells incubated with superparamagnetic iron oxide nanoparticles. Notes: Mesenchymal cells were harvested from bone marrow of Wistar rats and cultured for three passages. They were then incubated with Feridex (50 μg/mL) and protamine (5.0 μg/mL) for 4 hours and trypsinized for subsequent transplantation. Immunostaining using the primary antibody anti-dextran (1:1000; Stem Cell Technologies, Vancouver, BC, Canada) was performed in trypsinized cells to detect the superparamagnetic iron oxide nanoparticles. (A) Graph showing the high proportion of dextran-positive cells. (B–B‴) Fluorescent microscopy images, showing (B) nuclei stained with 4′,6-diamidino-2-phenylindole (blue), (B′) dextran-positive cells, and (B″) merged images. (B‴) Higher-magnification image of the area indicated by the white box. Scale bar =50 μm.

Figure 2

Differentiation potential of mesenchymal stem cells labeled with SPIONs. Notes: Mesenchymal cells were harvested from bone marrow of Wistar rats, cultured, and induced to differentiate in chondrocytes and adipocytes after the exposure to Feridex (50 μg/mL) and protamine (5.0 μg/mL) for 4 hours. (A) Alcian blue staining showing chondrogenesis in Feridex-labeled cells. The nuclei were counterstained with nuclear fast red. (A′) Higher magnification of the black box, showing brown deposits indicating the presence of SPIONs. (B) Oil Red O staining indicating adipogenesis in labeled cells. (C–C″) Representative images showing higher magnification of dextran-positive cells after adipocyte differentiation. These images were acquired in a black/white charge-coupled device camera, and thus red color from Oil Red staining is not visible in the figure. (C) Bright-field microscopy image of an adipocyte revealed by Oil Red staining, (C′) nuclei counterstained with 4′,6-diamidino-2-phenylindole (blue) and dextran-positive cell (green), and (C″) merged images. Scale bar =50 μm. Abbreviation: SPION, superparamagnetic iron oxide nanoparticle.

Figure 3

Tracking-labeled mesenchymal stem cells 24 hours after transplantation in infarcted hearts. Notes: Positive cells for dextran (green) were found both at the injection site and distant (myocardial septum) from the injection site. The nuclei were counterstained with 4′,6-diamidino-2-phenylindole (blue). (A) Photomontage showing cells in the injected area in the left ventricle wall (the arrowheads point for dextran-positive cells into the heart tissue). (B) Representative image of dextran-positive cells found in areas close to the injection site. (C) Representative image of labeled cells in a region far from the injection area, the myocardial septum. Scale bar =200 μm in (A) and 50 μm in (B) and (C).

Figure 4

Tracking of transplanted labeled mesenchymal stem cells 12 days after transplantation in the infarcted heart. Notes: Confocal microscopy of the tissue, showing rare positive cells for dextran (green) and nuclei counterstained with DAPI (blue). (A) Cells stained with dextran, showing different morphologies. (B–B″) Higher magnification of the box in (A), showing (B) nuclei counterstained with DAPI, (B′) dextran-positive cell, and (B″) merged image. (C–C‴) Higher magnification of the box in (A) showing (C) dextran-positive cell, (C′) troponin I staining (red), (C″) nuclei counterstained with DAPI, and (C‴) merged image. Scale bar =50 μm. Abbreviation: DAPI, 4′,6-diamidino-2-phenylindole.

Figure 5

Qualitative analysis of proliferation in infarcted myocardium 48 hours after injury and 24 hours after Feridex-labeled mesenchymal cell transplantation. Notes: Fluorescence microscopy images showing Ki-67-positive cells (red) and dextran-positive cells (green) in an area close to the injection site. The nuclei were counterstained with DAPI (blue). (A, B) DAPI staining, (A′, B′) Ki-67 expression, (A″, B″) dextran staining, and (A‴, B‴) merged images. (B″″) Higher magnification image of the area is indicated by the white box showing a cell positive for both Ki-67 and dextran. Scale bar =50 μm. Abbreviation: DAPI, 4′,6-diamidino-2-phenylindole.

Figure 6

Qualitative analysis of apoptosis in infarcted heart region 48 hours after injury and 24 hours after cell transplantation. Notes: Fluorescence microscopy images showing activated caspase-3-positive cells (red), dextran-positive cells (green), and nuclei counterstained with DAPI (blue). (A) Representative image close to the region of cell injection. (B–C″) Images in higher magnification are also close to the injection site. (B, C) DAPI staining, (B′, C′) activated caspase-3 expression, (B″, C″) dextran staining, and (B‴, C‴) merged images. Scale bar =50 μm. Abbreviation: DAPI, 4′,6-diamidino-2-phenylindole.