Figure 3.

Wnt7a and active β-catenin stimulate neural stem cell (NSC) proliferation and self-renewal. (a) Western blot analysis of control IgG and Wnt7a–IgG expression in stable HEK 293 cells with the use of human IgG antibody. (b) Proliferation of low-density adult mouse NSCs during co-culture with control IgG (C) and Wnt7a–IgG (Wnt)-expressing cells was monitored by cell numbers in each input well. Data are means ± s.d.; asterisk, P < 0.01 by Student’s t-test, n = 6. (c) The self-renewal capacity of adult mouse NSCs co-cultured with cells expressing control IgG and Wnt7a–IgG was determined by secondary clonal rate. Data are means ± s.d.; asterisk, P < 0.05 by Student’s t-test, n = 5. (d) Multipotency of the Wnt7a-treated NSCs. Cells were immunostained for a neuronal marker (Tuj1), an astrocyte marker (GFAP) and an oligodendrocyte marker (O4). Scale bar, 100 μm. (e) Expression of β-catenin ΔN90 in transduced adult rat NSCs analysed by western blot analysis with anti-β-catenin antibody. WT, wild-type β-catenin; ΔN90, β-catenin ΔN90. (f) Cell-cycle analysis of control and β-catenin ΔN90-transduced adult rat NSCs. The percentages of cells at G1 phase and at G2/S/M phases are indicated. (g) The self-renewal capacity of adult rat NSCs transduced with control vector (C) or β-catenin ΔN90-expressing virus (ΔN90) was determined by secondary clonal rate. Data are represented as means ± s.d.; asterisk, P < 0.05 by Student’s t-test, n = 5. (h) Representative clonal derivatives of adult rat NSCs transduced with control vector or β-catenin ΔN90-expressing virus. d1, d10: days 1 and 10 of clonal analysis, respectively. Scale bar, 100 μm. (i) Multipotency of β-catenin ΔN90-transduced NSCs. Cells were differentiated and immunostained for Tuj1, GFAP, and O4. 4,6-Diamidino-2-phenylindole (DAPI) staining is shown in blue. The right-hand panel is the merged image. Representative O4-positive cells are shown in the inset. Scale bar, 50 μm. Uncropped images of blots in (a) and (e) are shown in Supplementary Information, Fig. S5.