Figure 5

Treatment with rIL-17A led to down-regulation of T regulatory cells in Tyk-2 deficient mice.

(a). Mice were treated with OVA alone and OVA + rIL-17A as shown in the experimental design. (b).–(d). Lung CD4⁺T cells from OVA alone and OVA + rIL-17A treated mice were isolated and incubated for 24 h with α-CD3 and α-CD28. The supernatants were analyzed by ELISA for IL-4, IL-5 and IL-13 ((b):IL-4: n = 3–4; p = 0.004; p = 0.003; (c): IL-5: n = 3–4; p = 0.0002; p = 0.040; (d): IL-13: n = 3–4; p = 0.002; p = 0.018; p = 0.017). (e). BALF was retrieved and BAL cells were incubated with antibodies against CD3, CD45R, CCR3 and Gr-1 and subsequently analyzed by flow cytometry. CD3⁻CD45R⁻CCR3⁻Gr-1⁺ cells were counted as neutrophils ((b). n = 3–4; p = 0.039; p = 0.029; p = 0.036; p = 0.015). (f). Total lung cells from OVA and OVA + rIL-17A treated mice were incubated with antibodies against CD4, CD25 and Foxp3 and analyzed by flow cytometry (n = 3–4; p = 0.001; p = 0.002). (g). Lung CD4⁺T cells from OVA alone and OVA + rIL-17A treated mice were isolated and incubated for 24 h with α-CD3 and α-CD28. The supernatants were analyzed by ELISA for IL-9 (n = 2–3; p = 0.0004; p = 0.016).