Figure 4.
Treatment of normal human leukocytes, activated T cells, and HUVECs with scFvCD7:sFasL. (A) Resting PBLs were subjected to treatment with scFvCD7:sFasL (325 ng/mL), or cross-linked sFasL (100 ng/mL) for up to 7 days, after which experimental apoptosis was assessed by annexin V/PI staining. Indicated values are representatives of 3 independent experiments. (B) Isolated PBLs were mixed at a ratio of 1:10 with DiI-labeled Jurkat cells. Mixed cultures were treated for 24 hours with scFvCD7:sFasL (100 ng/mL) or secondarily cross-linked sFasL (100 ng/mL). Differential fluorescent labeling of Jurkat target cells and PBLs was used to separately evaluate apoptosis induction by annexinV staining. Indicated values are representatives of 3 independent experiments. (C) Activated T cells were subjected to treatment with scFvCD7:sFasL (325 ng/mL) for up to 7 days, after which apoptosis was assessed by annexin V/PI staining. Indicated values are representatives of 3 independent experiments. (D) Resting HUVECs were treated for 24 hours with scFvCD7:sFasL (100 ng/mL), secondarily cross-linked sFasL (100 ng/mL), or actinomycin D (2 μg/mL). Apoptosis was assessed by ΔΨ. (E) Resting HUVECs were mixed with fluorescently labeled Jurkat cells (ratio 1:1) and treated with scFvCD7:sFasL (100 ng/mL) or actinomycin D (2 μg/ml) for 24 hours. Differential fluorescent labeling of Jurkat target cells and HUVEC bystander cells was used to separately evaluate apoptosis by ΔΨ.