FK778 allows the activation but inhibits the proliferation of allogeneic stimulated PBMCs. (A) A primary MLC was performed with 10⁵ responder PBMCs and 10⁵ allogeneic stimulator cells. Increasing concentrations of FK778 were added at day 0. Proliferation was measured at day 6, by means of ³H uptake, and percentage of inhibition of the primary MLC is depicted for increasing FK778 concentrations. Error bars represent SD counts. (B) The kinetics of proliferation during primary MLC (y-axis) was investigated during treatment with 0 μg/mL (•), 10 μg/mL (▵), or 50 μg/mL FK778 (○). At the peak of proliferation (day 5), **P < .002 for differences between untreated and FK778 treated cells. (C) Annexin-V expression was analyzed at day 6 of the primary MLC on CD3⁺CD4⁺ gated cells using flow cytometry. The number of annexin V⁺ cells is indicated in each plot. (D) At day 3, blast formation of untreated control cells (left panel), cells treated with 50 μg/mL FK778 (middle panel), and naive responder PBMC (right panel) was analyzed on CD3⁺CD4⁺ cells using flow cytometry. The number of blasts is indicated in each plot. (E) Cell-surface marker expression at day 6 of the primary MLC (left column shows CD25; right column, HLA-DR) of untreated control cells (top row), FK778-treated cells (middle row), and unstimulated responder PBMCs (bottom row) was analyzed on CD3⁺CD4⁺ gated cells. Percentage of positive cells is depicted in each plot, and the mean fluorescent intensity (MFI) of CD25 expression in the positive fractions is given. One representative experiment of 8 is presented.