FIG. 1.

Micrococci (Micro) and sPGN induce transcription of IL-8 in HEK293 cells expressing TLR2, and this induction is NF-κB dependent. (A) HEK293/TLR2 cells were stimulated with micrococci at 40 μg/ml or sPGN at 10 μg/ml, total RNA was isolated, IL-8 and GAPDH cDNAs were synthesized and amplified using primers specific for IL-8 and GAPDH, and the amplified products were separated on a 3% agarose gel. The results shown are from one of two similar experiments. (B) 293 cells were cotransfected with the following plasmids: TLR2, TLR1, or a control vector at 0.25 μg/ml, CD14 at 0.025 μg/ml, and IL-8 reporter pIL8(wt)CAT at 0.125 μg/ml. At 24 h after transfection, cells were stimulated with micrococci at 40 μg/ml, sPGN at 10 μg/ml, or TNF-α at 100 ng/ml for 16 h and cell lysates were assayed for CAT activity. (C) 293 cells were cotransfected with plasmids TLR2, CD14, and pIL8(wt)CAT (as described for panel B) or with an IL-8 reporter plasmid with a nonfunctional NF-κB site, pIL8(−κB)CAT, at 0.125 μg/ml. Cells were stimulated and assayed as described for panel B. The results are means of duplicate samples from one of three similar experiments. CV, control vector.