Mechanism of inhibition of IL-2 transcription by cAMP in primary human T cells.
(A) T cells were stimulated with either anti-CD3 plus anti-CD28 or PMA plus anti-CD28 in the presence of either media alone or with forskolin and IBMX. At the indicated time intervals (3 and 6 hours) of culture whole cell extracts were prepared and expression of transcription factors was determined by SDS-PAGE and Western blot, using antibodies specific for c-Jun, phospho–c-Jun, c-Fos, and NFATp. Results are representative of 2 experiments. (B) Jurkat T cells were transiently transfected with a reporter construct driven by the 2-kb IL-2 promoter/enhancer or with reporter constructs of luciferase driven by either AP-1, NF-κB, or NFAT. Forty hours after transfection 5 × 105 cells were aliquoted and cultured with either media, a combination of anti-CD3 plus anti-CD28 mAb, or with phorbol ester and anti-CD28 mAb either alone or with forskolin and IBMX. After 6 hours of culture luciferase activity was measured. Results are representative of 3 experiments.
