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Differentiation of human embryonic stem cells into dopaminergic neurons for transplantation in Parkinson's disease

Correia, Sofia LU (2008) In Lund University, Faculty of Medicine Doctoral Dissertation Series 2008:34.
Abstract
Parkinson’s disease (PD) is a progressive neurodegenerative disorder, characterized by tremor, muscle rigidity and bradykinesia, which is primarily due to the degeneration of nigrostriatal dopaminergic neurons. Currently, patients are primarily treated with L-dopa and other drugs that enhance dopaminergic function. Those are effective during the first years, but eventually the majority of patients develop side effects and experience loss of efficacy. Replacement of the lost dopaminergic neurons by transplantation has been tested, with varying degrees of success, as a therapy for PD. These trials have used ventral mesencephalic donor tissue obtained from routine abortions. This source of donor tissue is, however, associated with several... (More)
Parkinson’s disease (PD) is a progressive neurodegenerative disorder, characterized by tremor, muscle rigidity and bradykinesia, which is primarily due to the degeneration of nigrostriatal dopaminergic neurons. Currently, patients are primarily treated with L-dopa and other drugs that enhance dopaminergic function. Those are effective during the first years, but eventually the majority of patients develop side effects and experience loss of efficacy. Replacement of the lost dopaminergic neurons by transplantation has been tested, with varying degrees of success, as a therapy for PD. These trials have used ventral mesencephalic donor tissue obtained from routine abortions. This source of donor tissue is, however, associated with several problems. For example, it is not readily obtainable, it is heterogeneous in cell composition and it is inherently connected to ethical issues related to the abortion. An ideal source of cell material should be available in large quantities in a predictable manner, highly enriched in midbrain dopaminergic neurons, and safe to transplant with no risks of infection or tumor growth. Human embryonic stem cells (hESCs), derived from the inner cell mass of human blastocysts, are such a potential source of cells. They are pluripotent, can self-renew and have already demonstrated the ability to generate dopaminergic neurons in culture. Presently, the application of hESCs as a therapy in PD has serious limitations, including low survival of derived dopaminergic neurons after transplantation, unclear functional integration of grafted dopaminergic neurons in the host brain and risk of teratoma/tumor formation.



The general aim of this thesis is to find possible solutions for some of the limitations in the use of hESCs in a cell replacement therapy for PD. We found that prolonged in vitro differentiation of hESCs (for at least 3 weeks) was essential to prevent formation of teratomas/tumor. We observed that addition of FGF-20 increased the yield of neurons expressing tyrosine hydroxylase (TH, the key enzyme in the biosynthesis of dopamine) derived from hESCs. This growth factor probably both promoted cell differentiation towards the dopaminergic phenotype and reduced cell death. Thus, FGF-20 can increase the chance of success for hESC-based therapy against PD by enhancing the number of dopaminergic neurons in hESC cultures. We have differentiated hESCs into dopaminergic neurons in sphere cultures. When we grafted hESC-derived spheres we found higher numbers of TH-expressing cells in the implants, compared to transplantation of a single cell suspension. The sphere culture step eliminated pluripotent cells, which can generate teratomas, but there were still proliferating neural stem cells present in the spheres that would continue to expand after transplantation. Further studies are essential to control the growth of the grafted cells in order to avoid tumor formations in the recipient brain. In conclusion, this thesis shows that prolonged differentiation of hESCs in the presence of FGF-20 in sphere cultures may provide the foundations of a protocol that can be used for a successful cell therapy against PD. (Less)
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author
supervisor
opponent
  • Dr. Minger, Stephen, King's College London
organization
publishing date
type
Thesis
publication status
published
subject
in
Lund University, Faculty of Medicine Doctoral Dissertation Series
volume
2008:34
pages
127 pages
publisher
Division of Neuroscience, Department of Experimental Medical Sciences, Lund University
defense location
Segerfalksalen
defense date
2008-03-19 09:15
ISSN
1652-8220
ISBN
978-91-85897-87-2
language
English
LU publication?
yes
id
0d78b2f4-50ae-48e2-8d65-5ea24e5c6b2d (old id 1043582)
date added to LUP
2008-03-05 08:51:16
date last changed
2016-09-19 08:44:49
@phdthesis{0d78b2f4-50ae-48e2-8d65-5ea24e5c6b2d,
  abstract     = {Parkinson’s disease (PD) is a progressive neurodegenerative disorder, characterized by tremor, muscle rigidity and bradykinesia, which is primarily due to the degeneration of nigrostriatal dopaminergic neurons. Currently, patients are primarily treated with L-dopa and other drugs that enhance dopaminergic function. Those are effective during the first years, but eventually the majority of patients develop side effects and experience loss of efficacy. Replacement of the lost dopaminergic neurons by transplantation has been tested, with varying degrees of success, as a therapy for PD. These trials have used ventral mesencephalic donor tissue obtained from routine abortions. This source of donor tissue is, however, associated with several problems. For example, it is not readily obtainable, it is heterogeneous in cell composition and it is inherently connected to ethical issues related to the abortion. An ideal source of cell material should be available in large quantities in a predictable manner, highly enriched in midbrain dopaminergic neurons, and safe to transplant with no risks of infection or tumor growth. Human embryonic stem cells (hESCs), derived from the inner cell mass of human blastocysts, are such a potential source of cells. They are pluripotent, can self-renew and have already demonstrated the ability to generate dopaminergic neurons in culture. Presently, the application of hESCs as a therapy in PD has serious limitations, including low survival of derived dopaminergic neurons after transplantation, unclear functional integration of grafted dopaminergic neurons in the host brain and risk of teratoma/tumor formation.<br/><br>
<br/><br>
The general aim of this thesis is to find possible solutions for some of the limitations in the use of hESCs in a cell replacement therapy for PD. We found that prolonged in vitro differentiation of hESCs (for at least 3 weeks) was essential to prevent formation of teratomas/tumor. We observed that addition of FGF-20 increased the yield of neurons expressing tyrosine hydroxylase (TH, the key enzyme in the biosynthesis of dopamine) derived from hESCs. This growth factor probably both promoted cell differentiation towards the dopaminergic phenotype and reduced cell death. Thus, FGF-20 can increase the chance of success for hESC-based therapy against PD by enhancing the number of dopaminergic neurons in hESC cultures. We have differentiated hESCs into dopaminergic neurons in sphere cultures. When we grafted hESC-derived spheres we found higher numbers of TH-expressing cells in the implants, compared to transplantation of a single cell suspension. The sphere culture step eliminated pluripotent cells, which can generate teratomas, but there were still proliferating neural stem cells present in the spheres that would continue to expand after transplantation. Further studies are essential to control the growth of the grafted cells in order to avoid tumor formations in the recipient brain. In conclusion, this thesis shows that prolonged differentiation of hESCs in the presence of FGF-20 in sphere cultures may provide the foundations of a protocol that can be used for a successful cell therapy against PD.},
  author       = {Correia, Sofia},
  isbn         = {978-91-85897-87-2},
  issn         = {1652-8220},
  language     = {eng},
  pages        = {127},
  publisher    = {Division of Neuroscience, Department of Experimental Medical Sciences, Lund University},
  school       = {Lund University},
  series       = {Lund University, Faculty of Medicine Doctoral Dissertation Series},
  title        = {Differentiation of human embryonic stem cells into dopaminergic neurons for transplantation in Parkinson's disease},
  volume       = {2008:34},
  year         = {2008},
}