Fig. 4.

IL-10 mRNA is increased in cells exposed first to DEP or benzo[a]pyrene and then to LPS. (a, left) PBMCs (n = 3) were incubated with 0, 1 and 10 ng/ml of DEP, and 10 ng/ml LPS was added after 24 h. RNA was extracted and hybridized to a panel of ³²P-labelled probes, and the hybridized IL-10 probe was quantified using a PhosphorImager SI, and normalized to the housekeeping probe G3PDH. Results are shown for a representative subject, and show a dose-dependent increase in IL-10 mRNA with increasing doses of DEP. (a, right) PBMCs were also incubated for 24 h with 1 or 10 nm benzo[a]pyrene as a surrogate for the PAH component of the DEP, then 10 ng/ml of LPS was added and RNA extracted after a further 16 h. Parallel cultures also received 1 nm of the competitive antagonist [α]naphthoflavone with each dose of benzo[a]pyrene. IL-10 mRNA was quantified and normalized to G3PDH. Results from a representative subject show increased IL-10 mRNA with benzo[a]pyrene preincubation, which is diminished with the coaddition of [α]naphthoflavone. Pyr = benzo[a]pyrene; Naph = [α]naphthoflavone. (b) Gel of a representative S1 nuclease protection assay showing increased IL-10 mRNA in cells exposed first to DEP (left) or benzo[a]pyrene (right), followed by LPS.