MR tracking of transplanted cells with "positive contrast" using manganese oxide nanoparticles

Abstract

Rat glioma cells were labeled using electroporation with either manganese oxide (MnO) or superparamagnetic iron oxide (SPIO) nanoparticles. The viability and proliferation of SPIO-labeled cells (1.9 mg Fe/ml) or cells electroporated with a low dose of MnO (100 microg Mn/ml) was not significantly different from unlabeled cells; a higher MnO dose (785 microg Mn/ml) was found to be toxic. The cellular ion content was 0.1-0.3 pg Mn/cell and 4.4 pg Fe/cell, respectively, with cellular relaxivities of 2.5-4.8 s(-1) (R(1)) and 45-84 s(-1) (R(2)) for MnO-labeled cells. Labeled cells (SPIO and low-dose MnO) were each transplanted in contralateral brain hemispheres of rats and imaged in vivo at 9.4T. While SPIO-labeled cells produced a strong "negative contrast" due to the increase in R(2), MnO-labeled cells produced "positive contrast" with an increased R(1). Simultaneous imaging of both transplants with opposite contrast offers a method for MR "double labeling" of different cell populations.

FIG. 1

In vivo MRI of labeled 9L cells 24 h after transplantation in the striata of rat brain. a–c: Spin echo image (TR = 1000 ms, TE = 14.1 ms). d–f: R₁ maps. g–i: R₂ maps. j–l: R₁/R₂ merged maps. Shown are representative images of three out of seven rats, two injected with MnO- and FeO-labeled cells (a,b), and one with FeO-labeled cells and unlabeled cells (c; control). Note the simultaneous double contrast in (j–l).

FIG. 2

R₁ and R₂ histograms. a: The R₁ of each pixel is presented for two representative rats. A region of interest (ROI) was selected to cover both the transplanted cells and brain tissue. Pixels of unlabeled transplanted cells (green, N = 71 pixels) have more uniform and lower R₁ values, while MnO-labeled transplanted cells (red, N = 58 pixels) exhibit a broader distribution, shifting toward higher R₁ values. The overlapping pixels are from brain tissue surrounding the labeled transplanted cells; the “tail” can be attributed to pixels with a higher R₁ that represent MnO-labeled cells. The average R₁ for unlabeled and MnO-labeled cells was 0.52 s⁻¹ and 0.63 s⁻¹, respectively. b: These findings are consistent with the average distribution of R₁ from MnO-labeled transplanted cells (red, N = 5) and unlabeled transplanted cells (green, N = 2). c: The R₂ distribution is similar for MnO-labeled transplanted cells (red, N = 5) and unlabeled transplanted cells (green, N = 2), implying that MnO can be used as dominant T₁ positive contrast agent.

FIG. 3

Spin echo images (TR = 1000 ms, TE = 14.1 ms) (a,b) and R₁ maps (c,d) of the same rat brain at 1 (a,c) and 3 (b,d) days after cell grafting. The contrast decreases for both MnO- and SPIO-labeled cells over time. Although it is difficult to see MnO-labeled cells on the spin echo image (b), the cells are still visible on the R₁ map (d).

FIG. 4

In vitro characterization of relaxation enhancement. The relative change in signal I/I₀ intensity (mean of region of interest [ROI]) is plotted for a series of TRs and TEs for (a,b) PBS samples (◇), 4% gelatin only samples (△), cells suspended in 4% gelatin labeled with MnO (•), and unlabeled cells (□); (c,d) MnO-labeled cell pellets (•) and unlabeled cell pellets (□).

FIG. 5

Assessment of cell viability and toxicity at 24 h (a) and 48 h (b) after electroporation with MnO (low: 100 μg Mn/ml) or (high: 785 μg Mn/ml) and SPIO (1.9 mg Fe/ml). Cell viability was tested using three different assays: MTS (black bars); Cell Titer Blue (gray bars); and Calcein-AM (empty bars). All bars represent the average ± SD of the percentage relative to control (electroporation without contrast agent).