FIGURE 2.

Dominant negative Tcf inhibits Wnt11-induced gene expression and osteoblast maturation. MC3T3 control (GFP) and MC3T3 Wnt11⁺ cells were transduced with a retrovirus encoding dnTcf. The cells were cultured for 2 days and placed in osteo-inductive conditions with and without BMP2. The cells were harvested for RNA at day 9 of induction. cDNA was synthesized for each sample, and gene expression was analyzed by Q-RT-PCR. A, Rspo2; B, Dmp1; C, Phex; D, Bsp. Values represent fold-change in gene expression relative to non-BMP-treated GFP controls, as described previously. Statistical analysis was performed using analysis of variance with subsequent Tukey-Kramer multiple comparison testing. n >3, p < 0.01. # ⊥ ¥ indicates statistical significantly increased expression in non-BMP (no GF)-treated Wnt11 cells relative to the other non-BMP treated groups (#, control; ⊥, control dnTcf; ¥, Wnt11 dnTcf). *†‡ indicates statistically significant differences between BMP-treated Wnt11⁺ cells and all other BMP-treated groups (*, control; †, control dnTcf; ‡, Wnt11⁺ dnTcf). E, MC3T3 control (GFP), Wnt11⁺, and Rspo2⁺ cells were placed in osteogenic media with or without BMP (at the indicated concentration). The cells were harvested after the onset of mineralization (days 12–16) and stained with Alizarin Red S. F, control/dnTcf, Wnt11⁺/dnTcf, and Rspo2⁺/dnTcf were cultured in parallel with the MC3T3 cells above. Results repeated three independent times. No GF, no growth factor.