Lung Expansion of Th1, Th2, Th17 and Tregs Figure 5. Lung Th1, Th2, Th17 and Treg cells in CD4+CD25+ and CD4+CD252 populations proliferate de novo and in situ during allergic inflammation. Number of lung CD4+CD25+ (A, C, E and G) and CD4+CD252 cells (B, D, F, H) that are expressing transcription factors (TF); T- bet (A and B; Th1), GATA-3 (C and D; Th2), RORct (E and F; Th17) and Foxp3 (G and H; Treg) and have incorporated BrdU (filled column) or have incorporated BrdU and also express a double amount of DNA (7-AAD++; hatched columns) in mice exposed to PBS or OVA. Data are shown as mean number of cells per gram tissue +SEM. ** p,0.01. doi:10.1371/journal.pone.0019889.g005 T-bet+ cells were predominantly distributed in the alveolar lung tissue, followed by the peribronchial and perivascular tissue. GATA-3+ and Foxp3+ cells were mainly expressed in the perivascular tissue, followed by peribronchial and alveolar tissue. Although the RORct+ cells were the smallest population detected in lung tissue, most were distributed in the peribronchial tissue, followed by the alveolar and perivascular tissue (Figure 6C). Cytokine concentration in lungs during allergic inflammation To determine the general inflammatory milieu in the lung, pro and anti-inflammatory cytokines, as well as cytokines relevant to the different T helper cell subsets, were quantified in the lung supernatant. Specifically, IL-2, IL-6, TNF (tumour necrosis factor), IFN-c (Th1), IL-4 (Th2), IL-17A (Th17) and IL-10 (Treg) PLoS ONE | www.plosone.org 8 May 2011 | Volume 6 | Issue 5 | e19889