Figure 2.

The threshold of stimulation resulting in TNF-α production does not correlate with the threshold required for apoptotic death. CTLs were assayed concurrently for production of TNF-α and for proliferation in response to stimulation with splenocytes pulsed with various concentrations of I10 peptide. Production of TNF-α was quantitated by ELISA of culture supernatants. CTLs were stimulated by APCs pulsed with the indicated concentration of peptide antigen. After 24 h of culture, supernatant was collected and assayed for TNF-α. Proliferation was assessed as the incorporation of [³H]thymidine between 24 and 48 h of culture. As peptide concentration increased, TNF-α production rose concurrently with the rate of proliferation until both reached maximums at similar concentrations of peptide. With further increases in peptide concentration, proliferation decreased, characteristic of the high antigen induction of apoptosis, whereas TNF-α production remained elevated. Because TNF-α production was maximal at antigen concentrations too low to observe apoptosis or inhibition of proliferation, the requirement for high dose antigen to trigger apoptotic death is not the result of the stimulation threshold for TNF-α production.