Costimulation through LIGHT enhances MAPK activity.
(A) MAPK activity is enhanced after LIGHT costimulation. BALB/c T cells were precultured in medium for 2 hours in the presence of 0.1% DMSO, or 0.1% DMSO plus 15 μM cytochalasin D. After washing, the cells were cultured in medium (MED), or stimulated with solid phase TR6-Fc, a suboptimal concentration of anti-CD3, or both, for 90 minutes at 37°C. The levels of phosphorylated p44/42 MAPK and total p44/42 MAPK were assessed by immunoblotting with the same membrane. The 42-kDa band representing phosphorylated MAPK and the 44-kDa and 42-kDa bands representing total p44/42 MAPK protein are indicated by arrows. (B) P44/42 MAPK activity is essential for LIGHT-enhanced T-cell proliferation. BALB/c T cells were cultured in wells coated with TR6-Fc (10 μg/mL), a suboptimal concentration of anti-CD3 (0.2 μg/mL), or both. The cells were cultured in medium, or in the presence of a p44/42 MAPK inhibitor PD98059 (15 μM) or its noninhibitory structural homologue SB202474 (15 μM). The 3H-thymidine uptake of these cells was measured between 48 and 64 hours. The samples were tested in triplicate, and means ± SD of counts per minute are shown. The experiments were performed twice with similar results, and the data of a representative experiment are presented.
