Figure 4.

DGKαζDKO-enhanced airway Th17 responses. Thy1.1⁺Thy1.2⁺ congenic mice injected with 1.5 × 10⁶ Thy1.1⁻Thy1.2⁺Vα2⁺CD4⁺ WT or DKO naïve OT2 T cells on day −1 were intranasally injected with OVA_323−339 peptide on days 0, 1, and 2. Draining mediastinal lymph nodes and spleens were harvested on the seventh day. (A) Representative dot plots of dLN cells and splenocytes. Top panels: CD4 vs. TCRVα2 staining. Bottom panels: Thy1.1 vs. Thy1.2 staining of the gated TCRVα2⁺CD4⁺ population. (B,C) Percentages (B) and number (C) of donor-derived OT2 T cells in dLNs and spleens. (D–H) Splenocytes and dLN cells from recipients were stimulated with PMA and ionomycin for 4–5 h, followed by cell surface and intracellular staining. (D) Representative dot plots of indicated cytokines in donor-derived OT2 T cells. (E,F) Percentages (E) and number (F) of donor-derived IFN-γ-producing OT2 T cells. (G,H) Percentages (G) and number (H) of donor-derived IL-17A- and IL-17F-producing OT2 T cells. (I–M) Splenocytes and dLN cells were stimulated with OVA_323−339 for 2 days with GolgiPlug added in the last 5 h, followed by cell surface and intracellular staining. (I) Representative dot plots of indicated cytokine staining in gated donor-derived OT2 T cells. (J,K) Percentages of IFN-γ- (J) and IL-17-producing cells (K) in donor OT2 T cells. (L,M) IFN-γ (L) and IL-17A (M) concentrations in culture supernatants. Data shown are representative of or calculated from two independent experiments (n = 8). *P < 0.05; **P < 0.01; ***P < 0.001 as determined by the Student t-test.