Inhibition of maturation of DCs by TPT. (A) Human monocytes were cultured for 7 days with 50 ng/mL GM-CSF and 50 ng/mL IL-4 in the presence of TPT (D2-7, 2.5 nM; TPT-DC) or medium alone (Control-DC). Cells were stained with the designated mAb and analyzed by FACS. A typical experiment from at least 5 independent experiments with similar result is shown. Open profiles show staining with an isotype control, and black profiles show staining pattern with mAbs of the indicated specificity. (B) Human monocytes were cultured for 7 days with 50 ng/mL GM-CSF and 50 ng/mL IL-4 in the presence of TPT (TPT[D2-7], 2.5 nM) or medium alone (control). After thorough washing, the medium was restored in the presence of 10 ng/mL of LPS without TPT (columns 1 and 2). In another setting, control DCs were suspended in medium containing 10 ng/mL LPS supplemented with 2.5 nM of TPT (TPT (D8-9); column 3). After 48 hours of incubation, cells were labeled with the designated mAb and analyzed by FACS. Data shown are representative of at least 5 experiments with similar results. Open profiles show staining of LPS-stimulated DCs with an isotype control, black profiles show staining patterns with mAbs of the indicated specificity. For comparison, the staining pattern for day-7 cultured DCs was shown by dashed profiles. (C) Human monocytes were cultured with GM-CSF and IL-4 for 7 days. Then, the cells were simulated with sCD40L (300 ng/mL) or TNF (10 ng/mL), or poly (I:C) (12.5 μg/mL), in the presence of medium alone or TPT (2.5 nM). After 48 hours of incubation, cells were labeled with the designated mAb and analyzed by FACS. Data shown are representative of 3 experiments with similar results. Gray profiles show staining pattern of untreated DCs with mAbs of the indicated specificity, open profiles show staining pattern of TPT(D8-9)–treated DCs with indicated mAbs, and dashed profiles show isotype control.