Proliferative response and cytokine profile of B6.pfp^−/− and B6.gld T cells are intact.

(A) Proliferative response of splenic T cells from recipients of TCD-BM with B6, B6.gld, or B6.pfp^−/− T cells with (▪) or without (▩) irradiated (20 Gy) C3FeB6F1/J splenic stimulator cells (“Materials and methods”). (B) Splenic T cells from B6 (■), B6.gld (▪), and B6.pfp^−/− (░) mice were incubated for 4 hours with PMA (10 ng/mL) and ionomycin (2 μM). Brefeldin A (10 μg/mL) was added after the first hour of incubation. Intracellular cytokine expression in CD4 memory cells (CD4⁺, CD62L⁻, CD44⁺) and CD8 memory cells (CD8⁺, CD122⁺, CD44⁺) was measured by flow cytometric analysis. Cytokine expression in unstimulated controls was less than 7% (data not shown). (C) Splenic T cells from B6 (■), B6.gld (▪), and B6.pfp^−/− (░) mice were incubated with equal amounts of T-cell–depleted, irradiated (20 Gy) C3FeB6F1/J splenic stimulator cells in 24-well plates for 16 hours. Brefeldin A (10 μg/mL) was added after the first hour of incubation. Intracellular cytokine expression in CD4 memory cells (CD4⁺, CD62L⁻, CD44⁺) and CD8 memory cells (CD8⁺, CD122⁺, CD 44⁺) was measured by flow cytometric analysis. (D) Splenic T cells from B6 (■), B6.gld (▪), and B6.pfp^−/− (░) mice were incubated with equal amounts of irradiated (20 Gy) C3FeB6F1/J splenic stimulator cells in 24-well plates for 5 days. Cells were harvested and restimulated with T-cell–depleted, irradiated (20 Gy) C3FeB6F1/J splenic stimulator cells for 16 hours. Brefeldin A (10 μg/mL) was added after the first hour of incubation. Intracellular cytokine expression in CD4 memory cells (CD4⁺, CD62L⁻, CD44⁺) and CD8 memory cells (CD8⁺, CD122⁺, CD44⁺) was measured by flow cytometric analysis.