Magnetic resonance tracking of human CD34+ progenitor cells separated by means of immunomagnetic selection and transplanted into injured rat brain.

Magnetic resonance imaging (MRI) provides a noninvasive method for studying the fate of transplanted cells in vivo. We studied whether superparamagnetic nanoparticles (CD34 microbeads), used clinically for specific magnetic sorting, can be used as a magnetic cell label for in vivo cell visualization. Human cells from peripheral blood were selected by CliniMACS CD34 Selection Technology (Miltenyi). Purified CD34+ cells were implanted into rats with a cortical photochemical lesion, contralaterally to the lesion. Twenty-four hours after grafting, the implanted cells were detected in the contralateral hemisphere as a hypointense spot on T2 weighted images; the hypointensity of the implant decreased during the first week. At the lesion site we observed a hypointensive signal 10 days after grafting that persisted for the next 3 weeks, until the end of the experiment. Prussian blue and anti-human nuclei staining confirmed the presence of magnetically labeled human cells in the corpus callosum and in the lesion 4 weeks after grafting. CD34+ cells were also found in the subventricular zone (SVZ). Human DNA (a human-specific 850 base pair fragment of alpha-satellite DNA from human chromosome 17) was detected in brain tissue sections from the lesion using PCR, confirming the presence of human cells. Our results show that CD34 microbeads superparamagnetic nanoparticles can be used as a magnetic cell label for in vivo cell visualization. The fact that microbeads coated with different commercially available antibodies can bind to specific cell types opens extensive possibilities for cell tracking in vivo.

MRI of transplanted pancreatic islets.

A promising treatment method for type 1 diabetes mellitus is transplantation of pancreatic islets containing beta-cells. The aim of this study was to develop an MR technique to monitor the distribution and fate of transplanted pancreatic islets in an animal model. Twenty-five hundred purified and magnetically labeled islets were transplanted through the portal vein into the liver of experimental rats. The animals were scanned using a MR 4.7-T scanner. The labeled pancreatic islets were clearly visualized in the liver in both diabetic and healthy rats as hypointense areas on T2*-weighted MR images during the entire measurement period. Transmission electron microscopy confirmed the presence of iron-oxide nanoparticles inside the cells of the pancreatic islets. A significant decrease in blood glucose levels in diabetic rats was observed; normal glycemia was reached 1 week after transplantation. This study, therefore, represents a promising step toward possible clinical application in human medicine.

Temporal profile of ultrastructural changes in cortical neurons after a photochemical lesion.

A photochemical lesion was induced in the right sensory motor cortex of rat brains. We examined at various time points the occurrence of different types of neuronal death with respect to a potential therapeutic window. The lesion appearance was documented by magnetic resonance imaging, and functional recovery was evaluated by behavioral tests showing recovery at 48 hr after lesioning. At 0.5, 1, 3, 6, 12, 24, 48, and 72 hr postlesion, cortical layers IV and V were examined by light and electron microscopy. Ultrastructural changes, which corresponded well to light microscopy findings, were found in both hemispheres. In the lesioned area, the neuropil appeared disorganized at 0.5 hr, and apoptotic and necrotic cell death was found at 0.5-3 hr. After 3 hr, the tissue was disintegrated. On the contralateral side, chromatin clumping appeared at 0.5-3 hr. At 3 hr, ruptured membranes were found, a sign of irreversible cell death. At 6-72 hr, the membranes were intact, and the chromatin was not clumped but heterogeneously distributed. The nuclei contained dispersed nucleoli at 48-72 hr. The morphology correlated well with magnetic resonance images and functional behavior. Our study demonstrates that a photochemical lesion is a useful model for studying morphological changes in injured cells. It results in a permanent infarction within 3 hr. In that the morphology on the contralateral side drastically changed between 3 and 6 hr, the cellular alterations at these time points might represent a break point at which cells either progress toward cell death or recover.

Magnetic resonance tracking of transplanted bone marrow and embryonic stem cells labeled by iron oxide nanoparticles in rat brain and spinal cord.

Nuclear magnetic resonance (MR) imaging provides a noninvasive method for studying the fate of transplanted cells in vivo. We studied, in animals with a cortical photochemical lesion or with a balloon-induced spinal cord compression lesion, the fate of implanted rat bone marrow stromal cells (MSCs) and mouse embryonic stem cells (ESCs) labeled with superparamagnetic iron oxide nanoparticles (Endorem). MSCs were colabeled with bromodeoxyuridine (BrdU), and ESCs were transfected with pEGFP-C1 (eGFP ESCs). Cells were either grafted intracerebrally into the contralateral hemisphere of the adult rat brain or injected intravenously. In vivo MR imaging was used to track their fate; Prussian blue staining and electron microscopy confirmed the presence of iron oxide nanoparticles inside the cells. During the first week postimplantation, grafted cells migrated to the lesion site and populated the border zone of the lesion. Less than 3% of MSCs differentiated into neurons and none into astrocytes; 5% of eGFP ESCs differentiated into neurons, whereas 70% of eGFP ESCs became astrocytes. The implanted cells were visible on MR images as a hypointense area at the injection site, in the corpus callosum and in the lesion. The hypointense signal persisted for more than 50 days. The presence of GFP-positive or BrdU-positive and nanoparticle-labeled cells was confirmed by histological staining. Our study demonstrates that both grafted MSCs and eGFP ESCs labeled with a contrast agent based on iron oxide nanoparticles migrate into the injured CNS. Iron oxide nanoparticles can therefore be used as a marker for the long-term noninvasive MR tracking of implanted stem cells.

Imaging the fate of implanted bone marrow stromal cells labeled with superparamagnetic nanoparticles.

Bone marrow stromal cells (MSCs) are pluripotent progenitor cells that have the capacity to migrate toward lesions and induce or facilitate site-dependent differentiation in response to environmental signals. In animals with a cortical photochemical lesion, the fate of rat MSCs colabeled with magnetic iron-oxide nanoparticles (Endorem) and bromodeoxyuridine (BrdU) was studied. MSCs were either grafted intracerebrally into the contralateral hemisphere of adult rat brain or injected intravenously. In vivo MRI was used to track their fate; Prussian blue staining and transmission electron microscopy (TEM) confirmed the presence of iron-oxide nanoparticles inside the cells. During the first week posttransplantation, the transplanted cells migrated to the lesion site and populated the border zone of the damaged cortical tissue. The implanted cells were visible on MR images as a hypointense area at the injection site and in the lesion. The hypointense signal persisted for more than 50 days. The presence of BrdU-positive and iron-containing cells was confirmed by subsequent histological staining. Three to 4 weeks after injection, <3% of MSCs around the lesion expressed the neuronal marker NeuN. Our study demonstrates that a commercially available contrast agent can be used as a marker for the long-term noninvasive MR tracking of implanted cells.