Figure 5.

Constitutively active β-catenin and Wnt7a rescued TLX siRNA-mediated proliferation deficiency in neural stem cells (NSCs). (a) Active β-catenin rescued TLX siRNA-induced NSC proliferation deficiency. Control vector (C), TLX siRNA (siTLX), active β-catenin, or siTLX and β-catenin double-transduced adult mouse NSCs were cultured in N2-supplemented medium containing EGF and FGF for 46 h and pulse-labelled with BrdU for 2 h. Cell proliferation was determined by BrdU labelling (red). Merged images of BrdU labelling and phase contrast images (grey) are shown in the lower panels. Scale bar, 100 μm. (b) Quantification of the percentage of BrdU-positive (BrdU⁺) cells in total living cells of control vector (C), active β-catenin (β-cat.), TLX siRNA (siTLX), or both TLX siRNA and active β-catenin-transduced NSCs. Data are represented as means ± s.d.; asterisk, P < 0.01 by Student’s t test, (n = 3). (c) Recovery of TLX siRNA-mediated NSC proliferation deficiency by Wnt7a. Untreated adult mouse NSCs were plated at low cell density at the base of transwells and co-cultured with control vector or TLX siRNA-transduced NSCs, or co-cultured with TLX siRNA-transduced NSCs plus Wnt7a–IgG (Wnt7a)-expressing cells that were seeded in the upper chambers of transwells. Cells were cultured in N2-supplemented medium containing EGF and FGF. NSC proliferation was monitored by cell numbers in each input well at the base of transwells on day 5 of co-culture. Data are represented as means ± s.d.; asterisk, P < 0.05 by Student’s t test, (n = 3).