In vivo tracking of superparamagnetic iron oxide nanoparticle–labeled mesenchymal stem cell tropism to malignant gliomas using magnetic resonance imaging

Laboratory investigation

Restricted access


Mesenchymal stem cells (MSCs) have been shown to migrate toward tumors, but their distribution pattern in gliomas has not been completely portrayed. The primary purpose of the study was to assay the tropism capacity of MSCs to gliomas, to delineate the pattern of MSC distribution in gliomas after systemic injection, and to track the migration and incorporation of magnetically labeled MSCs using 1.5-T magnetic resonance (MR) imaging.


The MSCs from Fischer 344 rats were colabeled with superparamagnetic iron oxide nanoparticles (SPIO) and enhanced green fluorescent protein (EGFP). The tropism capacity of MSCs was quantitatively assayed in vitro using the Transwell system. To track the migration of MSCs in vivo, MR imaging was performed both 7 and 14 days after systemic administration of labeled MSCs. After MR imaging, the distribution patterns of MSCs in rats with gliomas were examined using Prussian blue and fluorescence staining.


The in vitro study showed that MSCs possessed significantly greater migratory capacity than fibroblast cells (p < 0.001) and that lysis of F98 glioma cells and cultured F98 cells showed a greater capacity to induce migration of cells than other stimuli (p < 0.05). Seven days after MSC transplantation, the SPIO–EGFP colabeled cells were distributed throughout the tumor, where a well-defined dark hypointense region was represented on gradient echo sequences. After 14 days, most of the colabeled MSCs were found at the border between the tumor and normal parenchyma, which was represented on gradient echo sequences as diluted amorphous dark areas at the edge of the tumors.


This study demonstrated that systemically transplanted MSCs migrate toward gliomas with high specificity in a temporal–spatial pattern, which can be tracked using MR imaging.

Abbreviations used in this paper: DMEM = Dulbecco modified Eagle medium; EDTA = ethylenediaminetetraacetic acid; EGFP = enhanced green fluorescent protein; FBS = fetal bovine serum; MR = magnetic resonance; MSC = mesenchymal stem cell; NSC = neural stem cell; PBS = phosphate-buffered saline; PFA = paraformaldehyde; SDF-1 = stromal cell-derived factor-1; SPIO = super-paramagnetic iron oxide nanoparticles.

Article Information

Address correspondence to: Xing Wu, M.D., Department of Neurosurgery, Huashan Hospital, Shanghai Medical School, Fudan University, 12 Wulumuqizhong Road, Shanghai, 200040, China. email:

© AANS, except where prohibited by US copyright law.



  • View in gallery

    Photomicrographs (A and B) and electromicrographs (C and D) of MSCs in culture after labeling with iron oxide nanoparticles (Prussian blue staining). A cluster of iron nanoparticles is shown surrounded by a cell membrane in the close vicinity of the Golgi apparatus (C and D, arrows), confirming the presence of iron inside the cell. The boxed area in C is shown at higher magnification in D. Original magnification × 200 (A), × 400 (B), × 4000 (C), and × 21,000 (D).

  • View in gallery

    Photomicrographs of the EGFP-labeled MSCs cultured with F98 cells for 48 hours. The MSCs migrated toward F98 clones and clustered around the clones, whereas no green MSCs could be seen in any other place (A). The process of MSC tropism to an F98 clone was captured by sequential photographs at 12-hour intervals (B–D). Only 1 green MSC was found in the F98 clone 12 hours after coculturing MSCs and F98 cells (B), whereas 24 hours later at least 5 green MSCs clustered around the F98 clone (C), and 36 hours later the green MSCs occupied almost half of the F98 clones (D). Original magnification × 100 (A) and × 200 (B–D).

  • View in gallery

    Bar graph showing the migratory capacity of MSCs and fibroblast cells. The F98 cells and lysis of F98 glioma cells significantly stimulated the directional migration of the MSCs compared with saline or normal brain tissue extract (*p < 0.01 for lysis of F98 glioma cells and *p < 0.05 for F98 cells, t-test). As expected, glioma cell lines (F98 cells) significantly stimulated MSC migration (up to 3-fold compared with the control) but interestingly, lysis of F98 glioma cells induced the highest chemotactic response (up to 5-fold). Normal brain tissue extract or saline did not change the basal migration rate. In addition, the response of fibroblast cells in migration capacity was not significantly different under the stimulus. Compared with fibroblast cells, F98 cells and lysis of glioma cells all promoted migration of MSCs significantly (#p < 0.001, t-test). Values shown are means ± standard error from 3 independent experiments.

  • View in gallery

    Photomicrographs of SPIO–EGFP colabeled MSCs after transplantation. A–C: Images obtained 7 days after transplantation showing that the SPIO–EGFP colabeled MSCs infiltrated and were distributed throughout the tumor. Prussian blue staining (A), EGFP staining (B), and combination of A and B (C). D–F: After 14 days, most colabeled MSCs were found at the border between the tumor and normal parenchyma, and only a few of the MSCs had infiltrated the tumor bed. Prussian blue staining (D), EGFP staining (E), and combination of D and E (F). G and H: The blue MSCs appeared to “follow” the invading tumor cell into surrounding tissue (G). This “trailing” of individual glioblastoma cells migrating away from the main tumor bed is examined in greater detail in H. The blue-stained MSCs are in direct juxtaposition to a single migrating and invading neutral red spindle-shaped tumor cell; the MSC “rides” the glioma cell in a “piggyback” fashion. The sections were costained with Prussian blue (allowing the SPIO-labeled MSCs to stain blue) and neutral red (allowing the elongated glioblastoma cells to stain dark red). I and J: Immunostaining showed EGFP-labeled green MSCs incorporated into the vessels at the edge of the tumor (Tu). The boxed area in I is shown at higher magnification in J. Some of the EGFP-labeled MSCs penetrated the vessels and then migrated in a chain pattern toward the glioma bed after 14 days (I). Original magnification × 200 (A–G and I) and × 400 (H and J).

  • View in gallery

    Bar graph showing the distribution pattern of systemically injected MSCs in different organs. At 14 days after MSC transplantation, Prussian blue–stained cells were found to scarcely distribute in the organs examined, except for within the brain tumor mass (glioma), where blue MSCs were highly concentrated. The number of blue MSCs was significantly higher in the brain tumors compared with the contralateral side of the nontumor-bearing part of the brain and other organs. *p < 0.001.

  • View in gallery

    Magnetic resonance images of SPIO–EGFP colabeled MSCs. These MSCs were examined at 7-day intervals for 2 weeks after MSC transplantation with T2-weighted spin echo and T2*-weighted gradient echo pulse sequence MR imaging. At 7 days the glioma showed a slightly high-signal intensity on the spin echo sequence (A, white arrow), whereas a well-defined dark hypointense region was shown on gradient echo sequence (B). Fourteen days after MSC transplantation, the tumor grew larger as demonstrated by a hyperintense edema region surrounding a significantly hyperintense necrosis core on the spin echo sequence (D), whereas the hypointense areas developed as 2 diluted amorphous curves at the tumor's edge on the gradient echo sequence (E). The hypointense signals on MR images in panels B and E correspond to multiple Prussian blue–stained cells in the histological sections in panels A and D of Fig. 4. As the 3D reconstructions (C and F) show, hypointense regions—which represent SPIO-labeled MSCs—in the MR slices are indicated by the yellow structures with the yellow arrows. The 3D images of SPIO-labeled MSCs in C and F are fitted with the histological sections (insets) and MR images in B and E, respectively. The tumors cannot be reconstructed in 3D because they cannot be shown in gradient echo sequences.


  • 1

    Aboody KSBrown ARainov NGBower KALiu SYang W: Neural stem cells display extensive tropism for pathology in adult brain: evidence from intracranial gliomas. Proc Natl Acad Sci U S A 97:12846128512000

  • 2

    Anderson SAGlod JArbab ASNoel MAshari PFine HA: Noninvasive MR imaging of magnetically labeled stem cells to directly identify neovasculature in a glioma model. Blood 105:4204252005

  • 3

    Arbab ASBashaw LAMiller BRJordan EKLewis BKKalish H: Characterization of biophysical and metabolic properties of cells labeled with superparamagnetic iron oxide nanoparticles and transfection agent for cellular MR imaging. Radiology 229:8388462003

  • 4

    Arbab ASPandit SDAnderson SAYocum GTBur MFrenkel V: Magnetic resonance imaging and confocal microscopy studies of magnetically labeled endothelial progenitor cells trafficking to sites of tumor angiogenesis. Stem Cells 24:6716782006

  • 5

    Arbab ASYocum GTKalish HJordan EKAnderson SAKhakoo AY: Efficient magnetic cell labeling with protamine sulfate complexed to ferumoxides for cellular MRI. Blood 104:121712232004

  • 6

    Arbab ASYocum GTRad AMKhakoo AYFellowes VRead EJ: Labeling of cells with ferumoxides-protamine sulfate complexes does not inhibit function or differentiation capacity of hematopoietic or mesenchymal stem cells. NMR Biomed 18:5535592005

  • 7

    Arbab ASYocum GTWilson LBParwana AJordan EKKalish H: Comparison of transfection agents in forming complexes with ferumoxides, cell labeling efficiency, and cellular viability. Mol Imaging 3:24322004

  • 8

    Askari ATUnzek SPopovic ZBGoldman CKForudi FKiedrowski M: Effect of stromal-cell-derived factor 1 on stem-cell homing and tissue regeneration in ischaemic cardiomyopathy. Lancet 362:6977032003

  • 9

    Barbero SBonavia RBajetto APorcile CPirani PRavetti JL: Stromal cell-derived factor 1alpha stimulates human glioblastoma cell growth through the activation of both extracellular signal-regulated kinases 1/2 and Akt. Cancer Res 63:196919742003

  • 10

    Benedetti SPirola BPollo BMagrassi LBruzzone MGRigamonti D: Gene therapy of experimental brain tumors using neural progenitor cells. Nat Med 6:4474502000

  • 11

    Bos CDelmas YDesmouliere ASolanilla AHauger OGrosset C: In vivo MR imaging of intravascularly injected magnetically labeled mesenchymal stem cells in rat kidney and liver. Radiology 233:7817892004

  • 12

    Brown ABYang WSchmidt NOCarroll RLeishear KKRainov NG: Intravascular delivery of neural stem cell lines to target intracranial and extracranial tumors of neural and non-neural origin. Hum Gene Ther 14:177717852003

  • 13

    Bulte JWKraitchman DL: Iron oxide MR contrast agents for molecular and cellular imaging. NMR Biomed 17:4844992004

  • 14

    Ceradini DJKulkarni ARCallaghan MJTepper OMBastidas NKleinman ME: Progenitor cell trafficking is regulated by hypoxic gradients through HIF-1 induction of SDF-1. Nat Med 10:8588642004

  • 15

    De Palma MVenneri MAGalli RSergi LSPoliti LSSampaolesi M: Tie2 identifies a hematopoietic lineage of proangiogenic monocytes required for tumor vessel formation and a mesenchymal population of pericyte progenitors. Cancer Cell 8:2112262005

  • 16

    Dunn IFBlack PM: The neurosurgeon as local oncologist: cellular and molecular neurosurgery in malignant glioma therapy. Neurosurgery 52:141114242003

  • 17

    Dwenger ARosenthal FMachein MWaller CSpyridonidis A: Transplanted bone marrow cells preferentially home to the vessels of in situ generated murine tumors rather than of normal organs. Stem Cells 22:86922004

  • 18

    Ehtesham MKabos PGutierrez MAChung NHGriffith TSBlack KL: Induction of glioblastoma apoptosis using neural stem cell-mediated delivery of tumor necrosis factor-related apoptosis-inducing ligand. Cancer Res 62:717071742002

  • 19

    Ehtesham MKabos PKabosova ANeuman TBlack KLYu JS: The use of interleukin 12-secreting neural stem cells for the treatment of intracranial glioma. Cancer Res 62:565756632002

  • 20

    Ehtesham MYuan XKabos PChung NHLiu GAkasaki Y: Glioma tropic neural stem cells consist of astrocytic precursors and their migratory capacity is mediated by CXCR4. Neoplasia 6:2872932004

  • 21

    Ferrari NGlod JLee JKobiler DFine HA: Bone marrow-derived, endothelial progenitor-like cells as angiogenesis-selective gene-targeting vectors. Gene Ther 10:6476562003

  • 22

    Frank JAMiller BRArbab ASZywicke HAJordan EKLewis BK: Clinically applicable labeling of mammalian and stem cells by combining superparamagnetic iron oxides and transfection agents. Radiology 228:4804872003

  • 23

    Grossman SABatara JF: Current management of glioblastoma multiforme. Semin Oncol 31:6356442004

  • 24

    Hamada HKobune MNakamura KKawano YKato KHonmou O: Mesenchymal stem cells (MSC) as therapeutic cytoreagents for gene therapy. Cancer Sci 96:1491562005

  • 25

    Heeschen CLehmann RHonold JAssmus BAicher AWalter DH: Profoundly reduced neovascularization capacity of bone marrow mononuclear cells derived from patients with chronic ischemic heart disease. Circulation 109:161516222004

  • 26

    Hilbe WDirnhofer SOberwasserlechner FSchmid TGunsilius EHilbe G: CD133 positive endothelial progenitor cells contribute to the tumour vasculature in non-small cell lung cancer. J Clin Pathol 57:9659692004

  • 27

    Imitola JRaddassi KPark KIMueller FJNieto MTeng YD: Directed migration of neural stem cells to sites of CNS injury by the stromal cell-derived factor 1α/CXC chemokine receptor 4 pathway. Proc Natl Acad Sci U S A 101:18117181222004

  • 28

    Keles GEBerger MS: Advances in neurosurgical technique in the current management of brain tumors. Semin Oncol 31:6596652004

  • 29

    Kim SKCargioli TGMachluf MYang WSun YAl-Hashem R: PEX-producing human neural stem cells inhibit tumor growth in a mouse glioma model. Clin Cancer Res 11:596559702005

  • 30

    Lee JElkahloun AGMessina SAFerrari NXi DSmith CL: Cellular and genetic characterization of human adult bone marrow-derived neural stem-like cells: a potential antiglioma cellular vector. Cancer Res 63:887788892003

  • 31

    Lefranc FBrotchi JKiss R: Possible future issues in the treatment of glioblastomas: special emphasis on cell migration and the resistance of migrating glioblastoma cells to apoptosis. J Clin Oncol 23:241124222005

  • 32

    Machein MRRenninger Sde Lima-Hahn EPlate KH: Minor contribution of bone marrow-derived endothelial progenitors to the vascularization of murine gliomas. Brain Pathol 13:5825972003

  • 33

    Magnitsky SWatson DJWalton RMPickup SBulte JWWolfe JH: In vivo and ex vivo MRI detection of localized and disseminated neural stem cell grafts in the mouse brain. Neuro-image 26:7447542005

  • 34

    Matuszewski LPersigehl TWall ASchwindt WTombach BFobker M: Cell tagging with clinically approved iron oxides: feasibility and effect of lipofection, particle size, and surface coating on labeling efficiency. Radiology 235:1551612005

  • 35

    Moore XLLu JSun LZhu CJTan PWong MC: Endothelial progenitor cells' “homing” specificity to brain tumors. Gene Ther 11:8118182004

  • 36

    Nakamizo AMarini FAmano TKhan AStudeny MGumin J: Human bone marrow-derived mesenchymal stem cells in the treatment of gliomas. Cancer Res 65:330733182005

  • 37

    Nakamura KIto YKawano YKurozumi KKobune MTsuda H: Antitumor effect of genetically engineered mesenchymal stem cells in a rat glioma model. Gene Ther 11:115511642004

  • 38

    Peters BADiaz LAPolyak KMeszler LRomans KGuinan EC: Contribution of bone marrow-derived endothelial cells to human tumor vasculature. Nat Med 11:2612622005

  • 39

    Phinney DGIsakova I: Plasticity and therapeutic potential of mesenchymal stem cells in the nervous system. Curr Pharm Des 11:125512652005

  • 40

    Rempel SADudas SGe SGutierrez JA: Identification and localization of the cytokine SDF1 and its receptor, CXC chemokine receptor 4, to regions of necrosis and angiogenesis in human glioblastoma. Clin Cancer Res 6:1021112000

  • 41

    Rich JNBigner DD: Development of novel targeted therapies in the treatment of malignant glioma. Nat Rev Drug Discov 3:4304462004

  • 42

    Rosu-Myles MStewart ETrowbridge JIto CYZandstra PBhatia M: A unique population of bone marrow cells migrates to skeletal muscle via hepatocyte growth factor/c-met axis. J Cell Sci 118:434343522005

  • 43

    Ruzinova MBSchoer RAGerald WEgan JEPandolfi PPRafii S: Effect of angiogenesis inhibition by Id loss and the contribution of bone-marrow-derived endothelial cells in spontaneous murine tumors. Cancer Cell 4:2772892003

  • 44

    Salmaggi AGelati MPollo BFrigerio SEoli MSilvani A: CXCL12 in malignant glial tumors: a possible role in angiogenesis and cross-talk between endothelial and tumoral cells. J Neurooncol 67:3053172004

  • 45

    Salmaggi AGelati MPollo BMarras CSilvani ABalestrini MR: CXCL12 expression is predictive of a shorter time to tumor progression in low-grade glioma: a single-institution study in 50 patients. J Neurooncol 74:2872932005

  • 46

    Sordi VMalosio MLMarchesi FMercalli AMelzi RGiordano T: Bone marrow mesenchymal stem cells express a restricted set of functionally active chemokine receptors capable of promoting migration to pancreatic islets. Blood 106:4194272005

  • 47

    Studeny MMarini FCChamplin REZompetta CFidler IJAndreeff M: Bone marrow-derived mesenchymal stem cells as vehicles for interferon-beta delivery into tumors. Cancer Res 62:360336082002

  • 48

    Tille JCPepper MS: Mesenchymal cells potentiate vascular endothelial growth factor-induced angiogenesis in vitro. Exp Cell Res 280:1791912002

  • 49

    Uhl MWeiler MWick WJacobs AHWeller MHerrlinger U: Migratory neural stem cells for improved thymidine kinase-based gene therapy of malignant gliomas. Biochem Biophys Res Commun 328:1251292005

  • 50

    Wang LLi YChen XChen JGautam SCXu Y: MCP-1, MIP-1, IL-8 and ischemic cerebral tissue enhance human bone marrow stromal cell migration in interface culture. Hematology 7:1131172002

  • 51

    Wynn RFHart CACorradi-Perini CO'Neill LEvans CAWraith JE: A small proportion of mesenchymal stem cells strongly expresses functionally active CXCR4 receptor capable of promoting migration to bone marrow. Blood 104:264326452004

  • 52

    Yaşargil MGKadri PAYaşargil DC: Microsurgery for malignant gliomas. J Neurooncol 69:67812004

  • 53

    Zhang ZJiang QJiang FDing GZhang RWang L: In vivo magnetic resonance imaging tracks adult neural progenitor cell targeting of brain tumor. Neuroimage 23:2812872004


Cited By



All Time Past Year Past 30 Days
Abstract Views 131 131 18
Full Text Views 147 147 6
PDF Downloads 87 87 2
EPUB Downloads 0 0 0


Google Scholar