Reduced transition from DN to DP thymocytes in E16.5 Gli3^+/– and Gli3^–/– mice. (A) Thymocytes from Gli3^+/+, Gli3^+/–, and Gli3^–/– mice were isolated and stained for expression of the cell surface markers, CD4 and CD8. Percentage of thymocytes in each quadrant is shown. (B) Data from multiple litters represented in panel A are shown graphically. The mean percentage of DN, CD8⁺ ISP, and DP thymocytes relative to the corresponding mean percentage in wild-type littermates for each litter are shown in the left, middle, and right panels, respectively. P values for the comparisons between the mean values of Gli3^–/– and Gli3^+/–, between Gli3^+/– and Gli3^+/+, and between Gli3^–/– and Gli3^+/+ were all less than .05. (C) Expression of CD2 on thymocytes isolated from E16.5 Gli3^+/+, Gli3^+/–, and Gli3^–/– mice. A reduction in CD2 expression accompanies the reduction in CD8 ISP and DP thymocyte populations seen in Gli3^+/– and Gli3^–/– mice. Histogram overlays show the expression of CD2 on total E16.5 thymocytes (shaded curves), DN cells (thin lines), and CD8 ISP cells (bold lines). Percentages of total thymocytes positive for CD2 are shown. (D) Data from multiple experiments as shown in panel C. The results are expressed as relative mean percentage of CD2 expression in thymocyte DN, CD8 ISP, and DP populations relative to the wild-type control. Error bars in panels B and D represent the standard error of the mean.