Quantitative Magnetic Particle Imaging Monitors the Transplantation, Biodistribution, and Clearance of Stem Cells In Vivo

Abstract

Stem cell therapies have enormous potential for treating many debilitating diseases, including heart failure, stroke and traumatic brain injury. For maximal efficacy, these therapies require targeted cell delivery to specific tissues followed by successful cell engraftment. However, targeted delivery remains an open challenge. As one example, it is common for intravenous deliveries of mesenchymal stem cells (MSCs) to become entrapped in lung microvasculature instead of the target tissue. Hence, a robust, quantitative imaging method would be essential for developing efficacious cell therapies. Here we show that Magnetic Particle Imaging (MPI), a novel technique that directly images iron-oxide nanoparticle-tagged cells, can longitudinally monitor and quantify MSC administration in vivo. MPI offers near-ideal image contrast, depth penetration, and robustness; these properties make MPI both ultra-sensitive and linearly quantitative. Here, we imaged, for the first time, the dynamic trafficking of intravenous MSC administrations using MPI. Our results indicate that labeled MSC injections are immediately entrapped in lung tissue and then clear to the liver within one day, whereas standard iron oxide particle (Resovist) injections are immediately taken up by liver and spleen. Longitudinal MPI-CT imaging also indicated a clearance half-life of MSC iron oxide labels in the liver at 4.6 days. Finally, our ex vivo MPI biodistribution measurements of iron in liver, spleen, heart, and lungs after injection showed excellent agreement (R(2) = 0.943) with measurements from induction coupled plasma spectrometry. These results demonstrate that MPI offers strong utility for noninvasively imaging and quantifying the systemic distribution of cell therapies and other therapeutic agents.

Keywords: Magnetic particle imaging; cell therapy tracking; mesenchymal stem cells; quantitative imaging.

Figure 1

Comparison of MPI/CT, fluorescent imaging, and MRI in mouse. Two probes filled with a mixture of Nanomag-MIP SPIOs and Angiosense 680 EX fluorescent probe were implanted 1.0 mm and 2.8 mm below the dorsal skin surface of a mouse. (A) A representative MPI image visualizes SPIO tracer with high image contrast and no signal modulation by biological tissue. MPI: 5 × 3.75 × 10 cm FOV, 6.8 min scan. CT: 15 min scan, 184 μm isotropic resolution. (B) Fluorescent imaging of implanted probes shows a decrease in signal with tissue depth. Fluorescent imaging: 5 sec scan. (C) In MR images, the presence of SPIO tracer corresponds to a signal dropout similar to those generated at air-tissue interfaces. MRI: 4 × 8 cm FOV, 313 μm in-plane resolution, 17 min scan.

Figure 2

MPI scanning process. (A) Opposing magnets form a gradient field with a central field-free region, labeled here as a field-free point (FFP). The FFP can be shifted using a series of electromagnets in a scan trajectory to cover the imaging volume. (B) The magnetization of SPIOs changes nonlinearly due to an externally applied magnetic field, which can be modeled using a Langevin function. The MPI point spread function (PSF) corresponds to the derivative of this Langevin function. (C) The traversal of the FFP across a spatially-varying SPIO distribution (shown in black) causes SPIO magnetization to change according to the Langevin curve. This change in SPIO magnetization induces a voltage signal in a detector coil, which can be assigned to the instantaneous FFP location for image reconstruction. (D-E) The prototype FFP MPI scanner used in this study has a 7 T/m magnetic gradient along the X direction and 3.5 T/m gradient along the Y and Z directions, which are generated using neodymium permanent magnets. The FFP can be shifted via electromagnets in the X and Y directions and a drive coil (not shown) in the Z direction.

Figure 3

Quantitative comparison of MPI and fluorescent signal with tissue depth. Six pairs of equivalent SPIO and fluorescent probes in sealed capillary tubes were placed at various depths in a sectioned porcine muscle phantom. (A-B) Maximum intensity projection MPI/CT images show constant SPIO signal at various depths in tissue, but equivalently placed fluorescent probes show significant signal reduction with tissue depth. (C) Log plot of image signal with tissue depth for MPI and fluorescent imaging shows no significant dependence of MPI signal on tissue depth, but exponential decrease of optical signal with increasing tissue depth. MPI imaging (n = 3): 4 cm × 3.75 cm × 8 cm FOV, 5.3 min scan. Fluorescent imaging (n = 5): 2 second exposure, F/2, 675 and 720 nm excitation/emission filters. CT imaging: 25 min acquisition, 184 μm isotropic resolution.

Figure 4

In vivo MPI experimental protocol. 4 groups of 3 rats each were used in this study. Groups 1 and 2 received tail vein injections of 5×10⁶ to 8×10⁶ Resovist-labeled mesenchymal stem cells. Group 3 received an equivalent tail vein injection of Resovist alone, and Group 4 received a tail vein injection of isotonic saline. Groups 1, 3, and 4 were sacrificed within one hour of injection and imaged using MPI and CT, while Group 2 was imaged in vivo using MPI over 12 days after injection, followed by sacrifice and MPI-CT imaging. Post-sacrifice, all animals were dissected and the liver, lungs, heart, and spleen were excised for MPI imaging and iron distribution analysis.

Figure 5

hMSC labeling using Resovist SPIO tracer. (A) Prussian blue staining of hMSCs labeled with four Resovist labeling concentrations ranging from 0 to 40 μg Fe / mL. Cells labeled with 40 μg Fe / mL Resovist show around 90-95% labeling efficiency with no differences in cell morphology. Scale bars: 100 μm. (B) An MTT-based cell viability assay shows no significant differences in cell viability between unlabeled and labeled hMSC populations.

Figure 6

MPI-CT imaging of intravenously injected hMSCs, Resovist, and saline control, with representative coronal, sagittal, and axial slices shown from full 3D MPI datasets. (A) MPI imaging of hMSC tail vein injections less than one hour post-injection shows substantial hMSC localization to lung. (B) At 12 days, hMSC tail vein injections show significant total clearance and liver migration. (C) MPI imaging of Resovist-only tail vein injections less than one hour post-injection shows immediate SPIO uptake in liver and spleen. (D) Control injections of isotonic saline show no detectable MPI signal. MPI imaging (n = 4 for each animal): 4 × 3.75 × 10 cm FOV, 9 minute acquisition. CT imaging: 25 minute acquisition, 184 μm isotropic resolution.

Figure 7

MPI quantification of in vivo SPIO clearance from hMSC injections. (A) In vivo MPI monitoring of intravenous hMSC injections (shown here as coronal summed intensity projections) shows gradual clearance from liver tissue from 1 day post-injection to 12 days post-injection. In vivo MPI imaging also shows benign respiratory motion artifacts (seen most prominently at 3 and 9 days post-injections), which do not affect MPI quantification and can be removed via the use of respiratory gating. MPI imaging (N = 3 animals, 2 MPI scans per time-point per animal): 4 × 3.75 × 10 cm FOV, 9 minute acquisition. (B) MPI measurements of in vivo iron clearance from labeled hMSC injections indicate in vivo clearance half-life at 4.6 days, with 95% confidence intervals at 3.7 and 6.0 days.

Figure 8

MPI analysis of postmortem SPIO biodistribution. MPI analysis of SPIO biodistribution for all groups was performed on liver, spleen, heart, and lung organs harvested postmortem. (A) MPI imaging of intravenous hMSC injections indicate localization of cells to lung tissue within one hour of injection. (B) 12 days after hMSC injection, MPI imaging shows substantial SPIO clearance and signal migration to liver and spleen. (C-D) Resovist SPIO intravenous injections localize immediately to liver and spleen, while no detectable MPI signal is found for control saline injections. The abbreviations li, s, h, and lu refer respectively to liver, spleen, heart, and lung. MPI imaging (n = 4 scans per animal): 4 × 3.75 × 10 cm FOV, 9 minute acquisition. All units are in µg Fe / mm².

Figure 9

Comparison of MPI-measured SPIO biodistribution to induction-coupled plasma spectrometry. (A) MPI-measured SPIO biodistribution for each experimental group. (B) A scatterplot of MPI and ICP-measured iron content in each organ for each experimental group shows high correlation (R² = 0.943, 95% linear confidence intervals shown). Data points are mean ± s.d. for both MPI and ICP measurements for each organ for each experimental group (n = 3 animals each).