ATP inhibits the capacity of mature DCs to attract type 1 but not type 2 polarized T-cell lines.

Polarized type 1 and type 2 T-cell lines were obtained from PBMCs stimulated with PHA in the presence of rhu IL-12 and anti–IL-4 mAb or rhu IL-4 and anti–IL-12 mAb, respectively. Cells were cultured for 10 to 14 days in the presence of 30 U/mL IL-2 and then examined for intracellular IFN-γ and IL-4 by flow cytometry after 6 hours of incubation with plate-bound anti-CD3 and soluble anti-CD28 mAbs to assess polarization (A,B). Expression of the chemokine receptors CCR4, CCR5, and CXCR3 was studied by flow cytometry on resting polarized type 1 (C) and type 2 (D) cells. Type 1 (E,G) and type 2 (F,H) T lymphocytes were tested for their migratory capacity to DC-derived supernatants diluted in complete medium and 0.5% BSA. Shown are results with supernatants from immature DCs (E,F) that were either untreated (○) or stimulated with 250 μM ATP (■) or 250 μM UTP (▵) and results with supernatants from DCs treated with 10 μg/mL LPS alone (●), LPS and ATP (▪), or LPS and UTP (▴) (G,H). Data from the migration assays are mean ± SD net migration results from 4 independent experiments using T-cell lines from 2 different donors. In panel G, differences in T-cell migration induced by supernatants from DCs stimulated with LPS alone and from DCs stimulated with LPS and ATP were significant (P < .03) at supernatants dilutions of 1:500 or less.