Factors influencing the differentiation of dopaminergic traits in transplanted neural stem cells.

1. Our previous studies demonstrated that when neural stem cells (NSCs) of the C17.2 clonal line are transplanted into the intact or 6-hydroxydopamine (6-OHDA) lesioned rat striatum, in most, but not all grafts, cells spontaneously express the dopamine (DA) biosynthetic enzymes, tyrosine hydroxylase (TH), and aromatic L-amino acid decarboxylase (Yang, M., Stull, N. D., Snyder. E. Y., Berk, M. A., and Iacovitti, L. (2002). Exp. Neurol.). 2. These results suggested that there were certain conditions which were more conducive to the development of DA traits in NSCs and possibly other neurotransmitter phenotypes. 3. In the present study, we modified a number of variables in vitro (i.e. passage number, confluence) and/or in vivo (degree, type, and site of injury) before assessing the survival, migration. and differentiation of engrafted NSCs. 4. We found that low confluence cultures were comprised exclusively of flattened polygonal cells, which when transplanted, migrated widely in the brain but did not express TH. 5. In contrast, high confluence cultures contained both polygonal cells and an overlying bed of fusiform cells. 6. When these NSCs were maintained for 12-20 passages and then transplanted, virtually all engrafted cells in 65% of the grafts expressed TH but not markers of other neurotransmitter systems. 7. Importantly, all TH+ grafts were accompanied by significant physical damage to the brain while TH- grafts were not, suggesting that local injury-related factors were also important. 8. Of no apparent influence on TH expression, regardless of how cells were grown prior to implantation, was the site of transplantation (cortex or striatum) or the degree of chemical lesion (intact, partial or full). 9. We conclude that transplanted NSCs can express traits specifically associated with DA neurons but only when cells are grown under certain conditions in vitro and then transplanted in proximity to injury-induced factors present in vivo.