Stromal factors <tex>$SDF1\alpha$</tex>, sFRP1, and VEGFD induce dopaminergic neuron differentiation of human pluripotent stem cells
Faculty of Pharmaceutical, Biomedical and Veterinary Sciences . Biomedical Sciences
Journal of neuroscience research. - New York
, p. 1367-1381
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 a (SDF1a), 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 SDF1a, 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 beta III-tubulin (a marker for immature neurons) in both the NTera2 and I6 cell lines. These results indicate that SDF1a, 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. (c) 2012 Wiley Periodicals, Inc.