AKT and β-catenin are required for activation of β-catenin–induced genes and inhibition of anti-inflammatory genes. wtNKG2D T cells were stimulated with anti-CD3/CD28–, anti-CD3–, anti-NKG2D–, or anti-CD3– and anti-NKG2D–coated beads in the presence of (A) β-catenin inhibitor XAV939 (10nM) or DMSO control (0nM) or (C,D) AKT inhibitor API-2 (2.5μM) or DMSO vehicle control (0μM). (A,C) After 8 hours, gene expression was determined by RT-PCR. Data are shown as the fold change in gene expression compared with T cells cultured in media at each concentration of inhibitor. Stimulation of T cells through NKG2D significantly changed gene expression compared with CD3 stimulation (**P < .05), and the presence of the inhibitor significantly changed gene expression compared with NKG2D stimulation in the presence of vehicle control (***P < .05). (B) Activated, nontransduced T cells were stimulated with media, anti-CD3–, anti-NKG2D–, or anti-CD3– and anti-NKG2D–coated beads in the presence of β-catenin inhibitor XAV939 (10nM) or DMSO vehicle control (0nM). After 24 hours, secretion of IFNγ was determined by ELISA, or after 72 hours production of IL-9, IL-10, IL-13, and VEGFα was determined by Luminex analysis. Cytokine secretion data are presented as mean ± SD and are from 2 donors (as indicated by donors 1 and 2). In the presence of the β-catenin inhibitor, T cells stimulated through CD3 and NKG2D secreted more anti-inflammatory cytokines and VEGFα than in the presence of the vehicle control. (D) chNKG2D (closed symbols) and wtNKG2D (open symbols) T cells were stimulated with media (squares), anti-CD3–coated (triangles), anti-NKG2D–coated (diamonds), or anti-CD3 and anti-NKG2D–coated (wtNKG2D T cells only; gray diamonds) beads in the presence of AKT inhibitor or vehicle DMSO. β-Catenin was measured by staining cells with anti–β-catenin Abs after 5 hours. Data shown were obtained from 1 donor and are representative of data from 3 donors.