Figure 5.
p21 loss does not directly affect neural stem cell potential to generate neurons or glia both in vivo or in vitro, but exhaustion is associated with accelerated decline in production of new cells in the olfactory bulb. (A) Neurosphere differentiation during passaging induced by exposure to serum for 7 d. The proportions of β-III-tubulin+ (neurons) or GFAP+ (astrocytes) cells in neurospheres derived from p21-/- and p21+/+ mice at 60 d of age. Data are presented as mean ± SEM. No significant effect of genotype was observed, indicating maintenance of multipotentiality and lack of terminal differentiation in neurospheres undergoing exhaustion in vitro; photomicrographs depict examples of staining of differentiated p21-/- primary neurospheres. Bars, 50 μm. (B) The numbers of Ki67-labeled cells in the subependyma of p21+/+ and p21-/- mice at 60 and 480 d of age; data are presented as mean ± SEM per section of lateral ventricle subependyma. There was a significant interaction between genotype and age [F (1, 11) = 7.33, p < 0.05] with significantly fewer Ki67+ cells in p21-/- than p21+/+ at 480 d of age (p < 0.05), but no difference at 60 d of age. (C) The numbers of BrdU and NeuN double-labeled cells (left) and single-labeled BrdU+ cells (middle) in sections of the granule cell layer of the olfactory bulb of p21+/+ and p21-/- mice at 60 d and 480 d of age; data are presented as mean ± SEM per section of olfactory bulb. (Right) The confocal image depicts examples of BrdU-labeled cells that are single-labeled or double-labeled for NeuN (double arrowhead) from the p21-/- olfactory bulb granule cell layer at 60 d of age. Bars, 50 μm. There were significant interactions between genotype and age for both the numbers of BrdU+/NeuN+ double-labeled cells [F (1, 15) = 5.78, p < 0.05] and the numbers of BrdU+/NeuN- single-labeled cells [F (1, 15) = 4.92, p < 0.05] with significantly fewer doubled-labeled and single-labeled cells in p21-/- than p21+/+ at 480 d of age (ps < 0.05), but no difference at 60 d of age.