Fig. 2.

TCR/CD3-ligation-induced apoptosis of semimature CD4⁺8^-HSA⁺ thymocytes is independent of Fas- and FADD-transduced death receptor signaling. Semimature CD4⁺8^-HSA⁺ thymocytes from WT, Fas-deficient lpr, and FADD-DN transgenic mice were purified by immunofluorescent staining with surface-marker-specific mAbs and cell sorting. Cells were cultured for 20 h in plates coated with graded concentrations of anti-CD3ε mAb in the absence (A) or presence (B) of optimal doses of anti-CD28 mAb, left untreated (C), or treated with FLAG-FasL (100 ng/ml) crosslinked with anti-FLAG mAb (1 μg/ml) (C). Apoptosis was assessed after 20 h as described in Fig. 1. Data represent arithmetic means ± SE of three to four independent experiments for animals of each genotype and for each treatment regime. No significant differences were observed in A, comparing WT vs. lpr (P > 0.52) and WT vs. FADD-DN (P > 0.163), or B, comparing WT vs. lpr (P > 0.171) and WT vs. FADD-DN (P > 0.218).