Reduced transition from DN to DP in E16.5 Gli2^−/− mice. (A) Thymocyte development on E16.5 (WT, n = 7; Gli2^+/−, n = 15; Gli2^−/−, n = 7). Left bar chart shows relative number of thymocytes on E16.5. Error bars show SE. The difference between WT and Gli2^−/− was not statistically significant. Dot plots show anti-CD4 and anti-CD8 staining on E16.5 thymocytes. Right bar chart shows the relative percentage of DP cells in E16.5 thymi. The difference between WT and Gli2^−/− was statistically significant (P = .007). Histograms show CD2 expression in E16.5 thymi. Left panels: CD2 expression on all thymocytes. Right panels: CD2 expression on DN cells. The difference in percentage of CD2⁺ cells overall and on DN cells between Gli2^+/+ and Gli2^−/− thymi were statistically significant (P = .01 and P = .04, respectively). (B) Apoptosis in thymocyte populations on E16.5, measured by annexin V staining. Scatterplots show the percentage of annexin V⁺ cells in DN1, CD25⁺DN, DN4, CD8ISP, and DP thymocyte populations in WT (n = 4), +/− (n = 8), and −/− (n = 4) embryos. Mean percentage positive was: DN1 WT, 1.85; +/−, 2.29; −/−, 2.62. CD25⁺DN WT, 1.55; +/−, 1.79; −/−, 2.0. DN4 WT, 0.57; +/−, 0.78; −/−, 0.72. CD8ISP WT, 2.10; +/−, 2.19; −/−, 2.75. DP WT, 1.01; +/−, 1.21; −/−, 1.32. There were no significant differences in the mean number of annexin V⁺ cells between genotype of mice in any thymocyte population. (C) Thymocyte development on E17.5 (WT, n = 7; Gli2^+/−, n = 4; Gli2^−/−, n = 7). Relative thymocyte number on E17.5. The difference between WT and Gli2^−/− was statistically significant (P = .02). Dot plots show anti-CD4 and anti-CD8 staining for E17.5 thymocytes. (D) Relative thymocyte number from E13.5 to E18.5, normalized to mean WT littermate value of 1.0. (For E18.5: WT, n = 5; Gli2^+/−, n = 5; Gli2^−/−, n = 5.) Error bars show SD.