Figure 2.

IL-5 expression requires de novo transcription and translation. (a) Cells were incubated with PMA (50 nm) and PHA (5 μg/ml), as indicated, for 24 hr. Supernatants were harvested and IL-5 release was measured. Data from at least five independent experiments are expressed as pg/ml means ± SEM. (b) Cells were incubated with PMA + PHA and RNA was harvested at 6 hr for semi-quantitative RT-PCR analysis of IL-5 and GAPDH. Agarose gels representative of six such experiments are shown. Data from six independent experiments are expressed as means ± SEM of percentage of the PMA + PHA stimulation. (c) PBMC were incubated with PMA + PHA as above in the presence or absence of actinomycin D (10 μg/ml) or cycloheximide (10 μg/ml). Supernatants were harvested at 24 hr and IL-5 release measured by ELISA. (d) Cells were treated with PMA + PHA and cycloheximide (10 μg/ml) as indicated and RNA was harvested after 6 hr for semi-quantitative RT-PCR analysis of IL-5 and GAPDH. Data (n = 3) were normalized to GAPDH. In (c) and (d) data from at least three independent experiments are expressed as percentage of stimulated as means ± SEM. ***P < 0·001, **P < 0·01.