A fraction of CD8
+ T cells becomes DN upon stimulation.
A, TCR-
αβ+ CD25
− T cells were sorted into CD4
+ and CD8
+ populations and cultured in the absence or presence of plate-bound anti-CD3 and anti-CD28 (5
μg/ml). Cells were harvested and stained with CD4 and CD8 at days 1, 3, and 5 and the quantity of DN cells was quantified. A representative experiment is shown.
B, Cumulative data (mean ± SEM) from one experiment is shown (
n = 4).
C, TCR-
αβ+ CD25
− CD8
+ T cells were sorted and stimulated in plates coated with anti-CD3 and anti-CD28 (5
μg/ml). After 5 days, cells were restained and sorted into CD8
+ and CD8
− subsets. Total RNA from nonstimulated CD8
+ T cells (
), as well as from poststimulation CD8
+ (□) and CD8
− (■) cells was isolated and cDNA was prepared. CD8
α transcript was quantified by real time PCR. GADPH was used as control.
D, Purified T cells were labeled with CFSE and incubated during 5 days without stimulation (■); with soluble anti-CD3 (5
μg/ml), anti-CD28 (2.5
μg/ml), and goat anti-mouse cross-linker Ab (2.5
μg/ml) (dark gray bar); in a transwell, separated from autologous monocytes by a permeable membrane (0.4
μm pores); in the same well with autologous monocytes. *,
p < 0.05.