title: Stromal factors SDF1α, sFRP1, and VEGFD induce dopaminergic neuron differentiation of human pluripotent stem cells.
abstract: Human embryonic stem cell (hESC)-derived dopaminergic (DA) neurons hold potential for treating Parkinson's disease (PD) through cell replacement therapy. Generation of DA neurons from hESCs has been achieved by coculture with the stromal cell line PA6, a source of stromal cell-derived inducing activity (SDIA). However, the factors produced by stromal cells that result in SDIA are largely undefined. We previously reported that medium conditioned by PA6 cells can generate functional DA neurons from NTera2 human embryonal carcinoma stem cells. Here we show that PA6-conditioned medium can induce DA neuronal differentiation in both NTera2 cells and the hESC I6 cell line. To identify the factor(s) responsible for SDIA, we used large-scale microarray analysis of gene expression combined with mass spectrometric analysis of PA6-conditioned medium (CM). The candidate factors, hepatocyte growth factor (HGF), stromal cell-derived factor-1 α (SDF1α), secreted frizzled-related protein 1 (sFRP1), and vascular endothelial growth factor D (VEGFD) were identified, and their concentrations in PA6 CM were established by immunoaffinity capillary electrophoresis. Upon addition of SDF1α, sFRP1, and VEGFD to the culture medium, we observed an increase in the number of cells expressing tyrosine hydroxylase (a marker for DA neurons) and βIII-tubulin (a marker for immature neurons) in both the NTera2 and I6 cell lines. These results indicate that SDF1α, sFRP1, and VEGFD are major components of SDIA and suggest the potential use of these defined factors to elicit DA differentiation of pluripotent human stem cells for therapeutic intervention in PD.
authors.affiliation: National Institute on Aging Intramural Research Program, National Institutes of Health, Laboratory of Neurosciences, Baltimore, MD 21224, USA.
authors.forename: Catherine M
authors.initials: CM
authors.lastname: Schwartz
authors.forename: Tahereh
authors.initials: T
authors.lastname: Tavakoli
authors.forename: Charmaine
authors.initials: C
authors.lastname: Jamias
authors.forename: Sung-Soo
authors.initials: SS
authors.lastname: Park
authors.forename: Stuart
authors.initials: S
authors.lastname: Maudsley
authors.forename: Bronwen
authors.initials: B
authors.lastname: Martin
authors.forename: Terry M
authors.initials: TM
authors.lastname: Phillips
authors.forename: Pamela J
authors.initials: PJ
authors.lastname: Yao
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Itoh
authors.forename: Wu
authors.initials: W
authors.lastname: Ma
authors.forename: Mahendra S
authors.initials: MS
authors.lastname: Rao
authors.forename: Ernest
authors.initials: E
authors.lastname: Arenas
authors.forename: Mark P
authors.initials: MP
authors.lastname: Mattson
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D018929:Cell Culture Techniques; D002460:Cell Line; D045744:Cell Line, Tumor; D054377:Chemokine CXCL12 / Q000096:biosynthesis / Q000502:physiology; D059290:Dopaminergic Neurons / Q000166:cytology* / Q000378:metabolism; D054278:Embryonal Carcinoma Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology / Q000502:physiology; D008565:Membrane Proteins / Q000494:pharmacology / Q000502:physiology; D051379:Mice; D009414:Nerve Growth Factors / Q000096:biosynthesis*; D058953:Neural Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D055495:Neurogenesis / Q000502:physiology*; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D017154:Stromal Cells / Q000378:metabolism; D014404:Tubulin / Q000096:biosynthesis; D014446:Tyrosine 3-Monooxygenase / Q000096:biosynthesis; D042643:Vascular Endothelial Growth Factor D / Q000096:biosynthesis / Q000502:physiology; D000094742:Secreted Frizzled-Related Proteins
chemical_list: D015415:Biomarkers; D054377:Chemokine CXCL12; D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009414:Nerve Growth Factors; D014404:Tubulin; D014446:Tyrosine 3-Monooxygenase; D042643:Vascular Endothelial Growth Factor D; C117636:SFRP1 protein, human; D000094742:Secreted Frizzled-Related Proteins