J Korean Neurosurg Soc.  2013 Dec;54(6):467-476. 10.3340/jkns.2013.54.6.467.

Engraftment of Human Mesenchymal Stem Cells in a Rat Photothrombotic Cerebral Infarction Model : Comparison of Intra-Arterial and Intravenous Infusion Using MRI and Histological Analysis

Affiliations
  • 1Department of Radiology, Chung-Ang University College of Medicine, Seoul, Korea. kwakbk@cau.ac.kr

Abstract


OBJECTIVE
This study aimed to evaluate the hypotheses that administration routes [intra-arterial (IA) vs. intravenous (IV)] affect the early stage migration of transplanted human bone marrow-derived mesenchymal stem cells (hBM-MSCs) in acute brain infarction.
METHODS
Male Sprague-Dawley rats (n=40) were subjected to photothrombotic infarction. Three days after photothrombotic infarction, rats were randomly allocated to one of four experimental groups [IA group : n=12, IV group : n=12, superparamagnetic iron oxide (SPIO) group : n=8, control group : n=8]. All groups were subdivided into 1, 6, 24, and 48 hours groups according to time point of sacrifice. Magnetic resonance imaging (MRI) consisting of T2 weighted image (T2WI), T2* weighted image (T2*WI), susceptibility weighted image (SWI), and diffusion weighted image of rat brain were obtained prior to and at 1, 6, 24, and 48 hours post-implantation. After final MRI, rats were sacrificed and grafted cells were analyzed in brain and lung specimen using Prussian blue and immunohistochemical staining.
RESULTS
Grafted cells appeared as dark signal intensity regions at the peri-lesional zone. In IA group, dark signals in peri-lesional zone were more prominent compared with IV group. SWI showed largest dark signal followed by T2*WI and T2WI in both IA and IV groups. On Prussian blue staining, IA administration showed substantially increased migration and a large number of transplanted hBM-MSCs in the target brain than IV administration. The Prussian blue-positive cells were not detected in SPIO and control groups.
CONCLUSION
In a rat photothrombotic model of ischemic stroke, selective IA administration of human mesenchymal stem cells is more effective than IV administration. MRI and histological analyses revealed the time course of cell migration, and the numbers and distribution of hBM-MSCs delivered into the brain.

Keyword

MRI; Mesenchymal stem cell; Photothrombotic cerebral infarction; Superparamagnetic iron oxide; Intravascular

MeSH Terms

Animals
Brain
Brain Infarction
Cell Movement
Cerebral Infarction*
Diffusion
Humans*
Infarction
Infusions, Intravenous*
Iron
Lung
Magnetic Resonance Imaging*
Male
Mesenchymal Stromal Cells*
Rats*
Rats, Sprague-Dawley
Stroke
Transplants
Iron

Figure

  • Fig. 1 In vitro cell staining. A : Non-labeled human mesenchymal stem cell, Prussian blue staining, original magnification ×400. B : Labeled hMSC, Prussian blue staining, original magnification ×400. C : Combined staining with antimitochondrial antibody-Prussian, original magnification ×400.

  • Fig. 2 Rat brain 5 days after photothrombotic infarction in right frontoparietal lobes. Whole cerebrum of rat brain (A) and coronal cross section (B).

  • Fig. 3 In vitro cellular MRI of PBS phantoms containing suspensions of different concentrations of SPIO labeled hBM-MSCs. A : T2 weighted image. B : T2* weighted image (GRE). C : Susceptibility weighted image. (a) 6.3×104 cells/mL, (b) 1.3×105 cells/mL, (c) 2.5×105 cells/mL, (d) 5.0×105 cells/mL, (e) unlabeled hMSC 5.0×105 cells/mL, (f) no cells (normal saline). PBS : phosphate-buffered saline, hBM-MSCs : human bone marrow-derived mesenchymal stem cells, SPIO : superparamagnetic iron oxide, hMSC : human mesenchymal stem cell.

  • Fig. 4 Serial brain MRI after regions IA (A) and IV (B) administration after photothrombotic infarction of rat model. Acute infarction are located on the right side of the brain (the left side of the images). Dark signal intensity regions increased over time especially in peripheral portion of infarction. SWI shows largest dark signal intensity regions following T2*WI and T2WI (arrows). Linear parenchymal low-signal intensities suggestive of veins are noted in all groups on SWI (double arrows). In the IA group, dark signal intensity in peripheral portion of infarction are more prominent than incorresponding images from a the IV group. PTCI : photothrombotic cerebral infarction, IA : intra-arterial, IV : intravenous, T2*WI : T2* weighted image, T2WI : T2 weighted image, SWI : susceptibility weighted image, DWI : diffusion weighted image.

  • Fig. 5 Quantitative analysis of MRI. Graph depicting mean (±SE) number of pixel count of dark signal intensity on SWI after either selective intra-arterial (IA) or intravenous (IV) transplantation in a rat model of photothrombotic infarction (*p<0.05). SWI : susceptibility weighted image.

  • Fig. 6 MRI of rat brain (IA 48 hr), SPIO-labeled hBM-MSCs directly grafted into a rat brain, and Prussian blue-stained cells and antimitochondrial-Prussian blue stained cells in a rat brain. A : Implanted cells are visualized as low signal intensity on SWI of MRI. B : Hypointense areas [square in (A)] represent massive invasions by Prussian blue stained cells (original magnification ×200, ×400). C : Graph showing the mean (±SE) number of engrafted cells in brain after either selective IA or IV transplantation (*p<0.05). D : Among these cells antimitochondrial-Prussian blue stained cells are noted (arrows, original magnification ×400). IA : intra-arterial, IV : intravenous, SPIO : superparamagnetic iron oxide, SWI : susceptibility weighted image, hBM-MSCs : human bone marrow-derived mesenchymal stem cells.

  • Fig. 7 Prussian blue-stained cells in lung specimen and graph showing the mean (±SE) number of engrafted cells after either selective intra-arterial (IA) or intravenous (IV) transplantation. A : Several Prussian blue-stained cells are noted in alveolar wall (arrows, original magnification ×400). B : hBM-MSC entrapment were significantly larger in the IV group than in the IA group at 1 hour time point (*p<0.05). hBM-MSCs : human bone marrow-derived mesenchymal stem cells.

  • Fig. 8 Aggregated MSCs at arterial lumen in IA rat model on combined Anti-mitochondrial antibody-Prussian blue staining. Note aggregated cells (arrows) which are positive on Anti-mitochondrial antibody (brown color) and Prussian blue staining (blue color).


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