J Korean Neurosurg Soc.  2014 Nov;56(5):383-389. 10.3340/jkns.2014.56.5.383.

Enhanced Efficacy of Human Brain-Derived Neural Stem Cells by Transplantation of Cell Aggregates in a Rat Model of Parkinson's Disease

Affiliations
  • 1Department of Neurological Surgery, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Korea. srjeon@amc.seoul.kr
  • 2Department of Biochemistry and Molecular Biology, University of Ulsan College of Medicine, Seoul, Korea.
  • 3Department of Maxillofacial Biomedical Engineering, Institute of Oral Biology, School of Dentistry, Kyung Hee University, Seoul, Korea.
  • 4Center for Bionics of Korea Institute of Science and Technology, Seoul, Korea.
  • 5Department of Neurosurgery, Konkuk University School of Medicine, Seoul, Korea.

Abstract


OBJECTIVE
Neural tissue transplantation has been a promising strategy for the treatment of Parkinson's disease (PD). However, transplantation has the disadvantages of low-cell survival and/or development of dyskinesia. Transplantation of cell aggregates has the potential to overcome these problems, because the cells can extend their axons into the host brain and establish synaptic connections with host neurons. In this present study, aggregates of human brain-derived neural stem cells (HB-NSC) were transplanted into a PD animal model and compared to previous report on transplantation of single-cell suspensions.
METHODS
Rats received an injection of 6-OHDA into the right medial forebrain bundle to generate the PD model and followed by injections of PBS only, or HB-NSC aggregates in PBS into the ipsilateral striatum. Behavioral tests, multitracer (2-deoxy-2-[18F]-fluoro-D-glucose ([18F]-FDG) and [18F]-N-(3-fluoropropyl)-2-carbomethoxy-3-(4-iodophenyl)nortropane ([18F]-FP-CIT) microPET scans, as well as immunohistochemical (IHC) and immunofluorescent (IF) staining were conducted to evaluate the results.
RESULTS
The stepping test showed significant improvement of contralateral forelimb control in the HB-NSC group from 6-10 weeks compared to the control group (p<0.05). [18F]-FP-CIT microPET at 10 weeks posttransplantation demonstrated a significant increase in uptake in the HB-NSC group compared to pretransplantation (p<0.05). In IHC and IF staining, tyrosine hydroxylase and human beta2 microglobulin (a human cell marker) positive cells were visualized at the transplant site.
CONCLUSION
These results suggest that the HB-NSC aggregates can survive in the striatum and exert therapeutic effects in a PD model by secreting dopamine.

Keyword

Parkinson's disease; Cell transplantation; Human brain-derived neural stem cells; Cell aggregates; Rat model; [18F]-FP-CIT microPET

MeSH Terms

Animals
Axons
Brain
Cell Transplantation
Dopamine
Dyskinesias
Forelimb
Humans
Medial Forebrain Bundle
Models, Animal*
Neural Stem Cells*
Neurons
Oxidopamine
Parkinson Disease*
Rats
Suspensions
Tissue Transplantation
Transplants
Tyrosine 3-Monooxygenase
Dopamine
Oxidopamine
Suspensions
Tyrosine 3-Monooxygenase

Figure

  • Fig. 1 In the step test, the number of contralateral forelimb touches in the HB-NSC group (n=9) improved significantly, compared to the control group (n=7), beginning at 6 weeks after HB-NSC transplantation. The increase was maintained through 10 week posttransplantation (*p<0.05). In the apomorphine-induced rotation test, a slight decrease was observed at week 8, compared to week 4, in the HB-NSC transplantation group, but the difference was not significant.

  • Fig. 2 Coronal images of [18F]-FDG and [18F]-FP-CIT PET scans in striatal area are shown, respectively. In the [18F]-FDG PET scan 10 week posttransplantation, the uptake did not show significant differences in bilateral striatum in control group as well as HB-NSC group, which means PD models were created successfully. [18F]-FP-CIT PET scans were performed before transplantation (PD), 5 weeks after transplantation (5 w), and 10 weeks after transplantation (10 w). The HB-NSC group showed a statistically significant recovery in the uptake of [18F]-FP-CIT in the transplanted striatum (arrow) at 10 weeks posttransplantation compared to PD (*p< 0.05) whereas there was no significant improvement of uptake at 5 weeks posttransplantation (p=0.065), suggesting that dopamine is secreted by the transplanted cells. The color bar indicates the relative uptake intensity and shows maximum standardized uptake values in the image.

  • Fig. 3 The results of immunohistochemistry and immunoflourescence for tyrosin hydroxylase (TH) at 12 weeks posttransplantation in coronal sections of the striatum in the HB-NSC group are shown in B, D, E, G, and the control group in A, C, F. The cell aggregates were TH-positive, and a round margin delineating the border between HB-NSC aggregates and host striatal tissue (red arrows) can been seen in D. The round margin of injected cells in HB-NSC group is shown prominently by IF staining (E), and colocalization of DAPI (blue), TH (red) and human β2 microglobulin (green) positive cells is observed in G (magnification ×400) which is indicated by circle in E. There was no positive staining of IHC (A and C) and IF (F) in control group.

  • Fig. 4 IHC staining of TH after 12 weeks ipsilateral cell transplantation with single-cell suspension (A, courtesy of Yoon et al. 2013) and cell aggregates (B). Different patterns of TH-positive staining in transplantation between single-cell suspensions and cell aggregates were detected. In transplantation with single-cell suspensions, abnormal parallel staining of dendrites which considered to be the characterastics of partially differentiated and poorly connected dopaminergic cells was observed (C, magnification ×200, courtesy of Yoon et al. 2013). In transplantation of cell aggregates, dendrites of the stained cells extended in various directions, suggesting the formation of a network with the neighboring cells (D, magnification ×200).


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