Figure 4

The S20-3 peptide–induced cell death involves TNFRI. (A) Immunoblot analysis of total cellular levels of TNF receptors I and II in BJAB, Jurkat, and Daudi cells. Numbers represent expression levels relative to GAPDH (loading control). (B) Daudi cells were pre-incubated for 1 hour with 5 μg/mL of TNFRI- or TNFRII-blocking antibodies, followed by 1 hour of treatment with 5 ng/mL of TNF-α or 100 μM peptide S20-3, and immediately analyzed for necrosis by LDH release assay. (C) BJABK1 cells (left panel) and BJAB cells (right panel) were pre-incubated for 1 hour with 5 μg/mL of TNFRI-blocking antibody, subsequently treated with 100 μM peptide S20-3 or 5 ng/mL of TNF-α for 1 hour, and analyzed as in (B). The relative cytotoxicity values in (B) and (C) were calculated as LDH release in [(treated-control)/(high control-control)]x100 and are shown as means ± SD of triplicate wells; *P < 0.005, **P < 0.02.