Figure 4.

Deletion of Cbl-b, Grail, or PD-1 in W131AOTII mice did not rescue anergic phenotypes in W131AOTII mice. (A) Relative expression of Cbl-b mRNA in naive and memory Vα2⁺ CD4 T cells from indicated mice (n = 3 mice/group). (B and C) Immunoblot analysis of Cbl-b (B) and Grail (C) on peripheral CD25⁻ naive OTII and W131AOTII T cells (n = 4–8 mice/group). (D) Flow cytometry of peripheral CD4⁺Vα2⁺ T cells to identify T reg cells (Foxp3⁺). (E) Bar graph depicts the frequencies of peripheral T reg cells of Vα2⁺CD4⁺ T cells. (F) Flow cytometry of peripheral CD4⁺Vα2⁺Foxp3⁻ T cells to identify anergic T cells (Foxp3⁻FR4⁺CD73⁺). (G) Bar graph shows the frequencies of peripheral anergic T cells of CD4⁺Vα2⁺ T cells. (H) Flow cytometry of peripheral CD4⁺Vα2⁺ T cells stained for CD44 and CD62L expression. (I) Bar chart shows the frequencies of peripheral memory T cells (CD44⁺CD62L⁻) of Va2⁺CD4⁺ T cells. Data are representative of three independent experiments (A–C) and combined from five independent experiments (n = 3–17 mice/group). Two-tailed Student’s t test was performed. *, P < 0.05; **, P < 0.005; ***, P < 0.0005.