Figure 4.

The inhibitory effect of CD4⁺CD25^high T cells requires cell contact and is not affected by the addition of TNFα in vitro. (A) CD4⁺CD25^high and responder T cells derived from patients responding to anti-TNFα therapy were stimulated with 5 mg/ml of soluble anti-CD3/anti-CD28 either with or without the use of a transwell membrane to prevent cell contact. (top) 2 μM monensin was added for the last 5 h of culture, and cells were stained for detection of intracellular cytokine production. (bottom) Cytokines from the supernatants collected from the same experiment were measured by CBA. (B) CD4⁺CD25^high and CD4⁺CD25⁻ T cells isolated from healthy as well as responding patients were cultured 1:1 ratio, upon anti-CD3/anti-CD28 stimulation, in the presence of different concentrations of TNFα. The percentage of suppression of cytokine production was calculated in each individual. One out of two experiments is shown.