Journal
EXPERIMENTAL NEUROLOGY
Volume 211, Issue 1, Pages 301-310Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.expneurol.2008.02.007
Keywords
stem cells; stem cell niche; neural tissue-spheres; mitogen; EGF; FGF2
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Tissue blocks containing neural precursor cells were isolated from the rat forebrain subventricular zone (SVZ) and ventral mesencephalon (VM) and propagated as neural tissue-spheres (NTS). In the presence of fibroblast growth factor-2 (FGF2) and epidermal growth factor (EGF), SVZ-derived NTS were propagated and maintained for more than 6 months with a cell population doubling time of 21.5 days. The replacement of EGF by leukemia inhibitory factor (LIF) resulted in a cell population doubling time of 19.8 days, Corresponding to a 10-fold increase in estimated cell numbers over a period of 70 days, at which point these NTS ceased to grow. In the presence of FGF2 and LIF, VM-derived NTS displayed a cell population doubling time of 24.6 days, which was maintained over a period of more than 200 days. However, when LIF was replaced by EGF, the cell numbers only increased 1.2 fold over 50 days. Using different immunohistochemical markers, we observed a distinct compartmentalization of cells within the spheres. In SVZ-derived NTS an outer compartment of proliferating (nestin(+)/Ki67(+)), preferentially neurogenic (beta-tubulin III+/MAP2(+)) cells, surrounded by an inner compartment of glial (GFAP(+)/CNPase(+)) cells. The inner compartment of long-term propagated VM-derived NTS contained GFAP(+) cells as well as cells immunoreactive for the precursor cell Market nestin, even where minimal cell proliferation was observed. Our results demonstrate that tissues from rat SVZ and VM can be propagated as NTS. However, the cellular organization of the NTS and the need for mitogens to maintain long-term proliferative capacity differ with the origin of the tissue. (C) 2008 Elsevier Inc. All rights reserved.
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