Lptn impairs the induction of Th1 but not Th2 type genes.

(A) CD4⁺ T cells were collected after activation for 24 or 48 hours, followed by RNA extraction, reverse transcription, and PCR using primer pairs specific for both Huβ_2m and IL-2 at 24 hours, and for both Huβ_2m and IFN-γ or IL-13 (not shown for IL-13) at 48 hours, as described in “Materials and methods.” One experiment of 4 is shown for either IL-2 or INF-γ. (B) Quantification of the RT-PCR experiments shown in panel A. Integrated intensity of the respective bands was determined as described in “Materials and methods.” Data shown represent the mean value ± SD (error bar) of the lymphokine/Huβ_2m signal ratio, calculated from 4 different experiments. The expression of IL-13 mRNA gave similar intensities with or without Lptn (not shown). (C) JA16 cells were electroporated with pIL-2, pIFN-γ, or pIL-4-Fluc constructs, together with pβ-actin-Rluc, as described in “Materials and methods.” After transfection, cells were either left unstimulated (not shown) or stimulated overnight as indicated. For each lymphokine promoter construct and each activation condition, the relative luciferase activity (RLU) corresponds to the ratio of Fluc expression to that of Rluc, and each RLU value is representative of at least 3 independent experiments. RLU obtained for PMA+ionomycin were 123.2 (2.44), 1.77 (0.024), 1.07 (0.81) for pIL-2, pIFN-γ, pIL-4, respectively, with SD in parentheses. Whatever the promoter used, RLU of unstimulated cells was 5- to 15-fold lower than in CD3-stimulated cells (not shown).