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Journal Article
Research Support, U.S. Gov't, P.H.S.
Neurotrophin channeling of neural progenitor cell differentiation.
Journal of Neurobiology 2001 March
The act of defining neuropoietic progenitor/stem cells is still in its early phases. Epidermal growth factor (EGF) stimulates extended proliferation of aggregates of subventricular striatal cells, taken from E15 mouse striatum, termed neurospheres in liquid culture. We have shown here and in previous work, using either immunohistochemistry or RT-PCR, that neurosphere cells express 13 cytokines (32 tested) and 20 cytokine receptors (28 tested), with 11 potential paracrine and nine potential autocrine loops. The neurotrophin receptors, Trk A, B, and C, were all expressed. Using a newly developed FACS single cell deposition technique, we evaluated the capacity of single EGF stimulated neurosphere cells to respond to the ligands for Trk A and B, nerve growth factor (NGF), and brain-derived neurotrophin factor (BDNF). Addition of NGF or BDNF to EGF for 14 days had no effect, but removal of EGF at day 14 with subsequent addition of BDNF or NGF resulted in an increase in neuronal and astroglial, but not oligodendrocyte, colony cells at 21 and 28 days of culture for BDNF, and of both cell types at 28 days for NGF. Tri-lineage colonies increased at day 21 with BDNF and at day 28 for both NGF and BDNF. Gross colony morphology also showed changes with neurotrophin addition, forming multiple individual cell balls or filamentous spreads. When EGF was withdrawn, a threshold effect was observed, with small, but not large, colonies ceasing growth. BDNF and NGF showed no effects on cell proliferation when compared to EGF controls, as determined by 5'-bromo-2-deoxyuridine (BrdU) incorporation and thus, they appear to affect differentiation of progenitor cells. These data indicate a sequential action of cytokines with EGF maintaining viability and proliferation and blocking differentiation. Removal of EGF is then permissive for the differentiating effects of BDNF and NGF. These data further indicate that the majority of EGF neurosphere clones have neurotrophin dependent tri-lineage potential.
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