Neurotransplantation of magnetically labeled oligodendrocyte progenitors: magnetic resonance tracking of cell migration and myelination

Abstract

Demyelination is a common pathological finding in human neurological diseases and frequently persists as a result of failure of endogenous repair. Transplanted oligodendrocytes and their precursor cells can (re)myelinate axons, raising the possibility of therapeutic intervention. The migratory capacity of transplanted cells is of key importance in determining the extent of (re)myelination and can, at present, be evaluated only by using invasive and irreversible procedures. We have exploited the transferrin receptor as an efficient intracellular delivery device for magnetic nanoparticles, and transplanted tagged oligodendrocyte progenitor cells into the spinal cord of myelin-deficient rats. Cell migration could be easily detected by using three-dimensional magnetic resonance microscopy, with a close correlation between the areas of contrast enhancement and the achieved extent of myelination. The present results demonstrate that magnetic resonance tracking of transplanted oligodendrocyte progenitors is feasible; this technique has the potential to be easily extended to other neurotransplantation studies involving different precursor cell types.

Figure 1

(a) CG-4 cells labeled for 48 h with 0.5 mg Fe/ml unconjugated MION-46L. No particle uptake can be detected by using the Prussian blue stain for ferric iron. (b–d) CG-4 cells labeled for 48 h with 0.05 mg Fe/ml MION-46L-OX-26. (b) numerous iron-containing vesicles are visible. These vesicles also contain the covalently linked, endocytosed OX-26 mAb, as visualized in c by using immunoperoxidase staining. Under these conditions the total Tfr binding capacity for OX-26 appears nearly saturated, given the similar staining pattern in d by using additional OX-26 mAb added afterward. (e) Transmission electron microscopy of MION-46L-OX-26-tagged CG-4 cells reveals the presence of numerous vesicles (arrows), which measure approximately 0.6–1.0 μm in diameter, and which are filled with the electron-dense magnetic nanoparticles. One of the vesicles (double arrows) is shown at a higher magnification in f to demonstrate the association of particles with a (reversed) endocytosed membrane. All cells in a–f were trypsinized before staining. (Bars represent 1 μm in e and 200 nm in f.)

Figure 2

1/T1 (A) and 1/T2 (B) as function of Larmor frequency for MION-46L-labeled (0.5 mg Fe/ml) and MION-46L-OX-26-labeled (0.05 mg Fe/ml) cells. The dependence of 1/T2 on TE is indicative of (intra)cellular clustering of the magnetic nanoparticles.

Figure 3

Md rat 10 days after transplantation of magnetically labeled CG-4 cells. (a) Shown are the three MRI planes of view for TE = 6 msec at 78 μm isotropic resolution. In the sagittal plane (top images, consecutive slices), cellular migration can be appreciated over a distance of 8.4 mm. The contrast in the transverse images (enlarged in bottom row, shown is each third interleaved slice) corresponds to the area of Prussian blue staining (b Upper) and antiproteolipid protein immunolabeling (c Upper) and shows a blooming effect caused by an extended-range susceptibility effect of the magnetic particles. (b and c Lower) Cell migration from the injection site toward the dorsal column, where the majority of the newly formed myelin was found. Note the differential orientation of the individual myelin fibers, which is along the direction of CG-4 cell migration. (Bars represent 100 μm in b and c Lower.)

Figure 4

Md rat 14 days after transplantation. Shown is the sagittal MRI plane for TE = 6 msec (a, consecutives slices) at 78-μm isotropic resolution, with cellular migration over a distance of 4.5 mm. The antiproteolipid protein immunolabeling (b and c) show that the observed MRI contrast corresponds closely to the myelination. In b, note the injection track with cell migration toward the dorsal column (the spherical appearance of the injection track in the MR images is the result of its direction relative to the orientation of the external magnetic field gradient). Prussian blue-positive cells (d) in the area of new myelination resembled the cellular morphology of oligodendrocytes. (Bars represent 1 mm in b and c and 10 μm in d.)

Figure 5

Md rat 14 days after transplantation of magnetically labeled and paraformaldehyde-fixed (dead) CG-4 cells. Shown are the three MRI planes of view for TE = 6 msec, with consecutive slices for the sagittal plane (top images) and interleaved slices for the transverse plane (bottom row, enlarged). In this control experiment, contrast is visible at the injection site only; no cellular migration can be observed. Magnifications: ×4, Top and Middle.