Development of stem (neural stem-, embryonic stem-, medullary stem) cells to dopaminergic neurons and their application to reconstruction medicine.

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Neural stem cells (NSCs) are self-renewing and multi-potent, thus are reasonable candidates of donor cells for neural transplantation once they steadily directed to neurons. In the present study, differentiation from NSCs to DAergic neurons and their application to reconstruction medicine were investigated. NSCs developed mostly to astrocytes in an ordinary culture system with serum. However, majority (over 80%) of ES cells-derived NSCs could be developed to neurons using 5 steps culture protocol with cytokines and trophic factors. Micro-array analysis revealed many genes were up-regulated or down-regulated in association with differentiation to neurons from NSCs. Among them, pleiotrophin highly expressed in NSCs showed a strong trophic and survival effects on DAergic neurons and progenitors. In embryonic brains, a homeotic factor ATBF1 was expressed in cells located in differentiation zone coexpressing with β- tubulin and MAP2. In expression study of neuroepithelial cells, Neuro 2A cells and P19 cells, it was found that when ATBF1 was expressed in cytosole cells proliferated continuously, and when ATBF1 was expressed inside nucleus cells left cell cycle and differentiated to neurons. Intra-nuclear transport of ATBF1 was dependent on the activity of PI(3) K-kinase and extranuclear (cytosol) localization was CRM1dependent. Thus, a homeotic factor ATBF1 was found be involved in neuronal differentiation, brain development, formation of brain architecture and so on. Results of the present study would be applied to various fields such as neural differentiation, reconstruction medicine, brain development and education.