title: Dopaminergic differentiation of human mesenchymal stem cells--utilization of bioassay for tyrosine hydroxylase expression.
abstract: Parkinson's disease (PD) is a neurodegenerative disorder, caused by a selective loss of dopaminergic neurons in the substantia nigra. In PD, the best therapeutic modalities cannot halt the degeneration. The selective hallmark pathology and the lack of effective treatment make PD an appropriate candidate for cell replacement therapy. Adult autologous bone-marrow-derived mesenchymal stem cells (MSCs) have been investigated as candidates for cell replacement strategies. Several laboratories, including ours, have induced MSCs into neuron-like cells demonstrating a variety of neuronal markers including dopaminergic characteristics, such as the expression of tyrosine hydroxylase (TH). This project aimed to induce MSCs into mature dopamine secreting cells and to generate a bioassay to evaluate the induction. For that purpose, we created a reporter vector containing a promoter of TH, the rate-limiting enzyme in the dopamine synthesis and red fluorescent protein DsRed2. Transfection of human neuroblastoma, dopamine synthesizing, SH-SY5Y cells confirmed the reliability of the constructed reporter plasmid. Following dopaminergic differentiation of the transfected human MSCs cells, TH expressing cells were identified and quantified using flow cytometry. Further study revealed that not only did the differentiated cells activate TH promoter but they also expressed TH protein and secreted dopamine. The reported results indicate that MSCs may be primed in vitro towards a dopaminergic fate offering the promise of innovative therapy for currently incurable human disorders, including PD.
authors.affiliation: Laboratory of Neurosciences, Felsenstein Medical Research Center, Rabin Medical Center, Beilinson Campus, Tel Aviv University, Petah Tiqwa 49100, Israel. innakan@post.tau.ac.il
authors.forename: Inna
authors.initials: I
authors.lastname: Kan
authors.forename: Tali
authors.initials: T
authors.lastname: Ben-Zur
authors.forename: Yael
authors.initials: Y
authors.lastname: Barhum
authors.forename: Yossef S
authors.initials: YS
authors.lastname: Levy
authors.forename: Alex
authors.initials: A
authors.lastname: Burstein
authors.forename: Tirza
authors.initials: T
authors.lastname: Charlow
authors.forename: Shlomo
authors.initials: S
authors.lastname: Bulvik
authors.forename: Eldad
authors.initials: E
authors.lastname: Melamed
authors.forename: Daniel
authors.initials: D
authors.lastname: Offen
mesh_terms: D001681:Biological Assay / Q000379:methods; D002454:Cell Differentiation / Q000502:physiology*; D045744:Cell Line, Tumor; D004298:Dopamine / Q000378:metabolism*; D005434:Flow Cytometry / Q000379:methods; D015870:Gene Expression; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000502:physiology*; D009447:Neuroblastoma; D014162:Transfection / Q000379:methods; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism*
chemical_list: D014446:Tyrosine 3-Monooxygenase; D004298:Dopamine