FIGURE 9.

Differential secretion of CCL20, TSLP and CCL-33. (A). SID-SRM for CCL20, TSLP and CCL-33 for phBECs vs phSAECs. Data are relative changes in expression in the secretome by RSV infection and cell type. (B). Differential expression is independent of the presence of medium supplements. SID-SRM for CCL20, TSLP, CCL-33 and IL-6 for phBECs and phSAECs grown and infected in the absence (−) or presence (+) of BSA/growth factor supplementation. Each bar is the mean ± SEM of triplicate determinations. (C). Cells were infected with or without RSV for 24h. Equal amounts of cDNA were taken for Q-RT-PCR for CCR6 and POLB expression using gene-specific primers. Each bar represents the mean ± SEM of cycle threshold (Ct) for triplicate assays. Experiments were repeated twice. ** = p<0.004. (D) Recombinant human CCL20 induces expression of mucin (MUC5AC) by hSAE cells in vitro. Different concentrations of recombinant CCL20 (rCCL20) were added to phSAEC cultures for 6 h. Shown is the fold change in MUC5AC mRNA expression by Q-RT-PCR. (E). hSAECs were incubated with different volumes of uninfected (CM) or UV-inactivated RSV conditioned medium (RSV-CM) for 6 h. MUC5AC mRNA expression was measured by Q-RT-PCR. (F). rCCL20 (2.5 ng/ml) was neutralized with mouse mAb to hCCL20 (25 ug/mL) or the same concentration of mouse IgG and added to the cell culture for 6 h. MUC5AC mRNA expression was assessed by Q-RT-PCR. (G). RSV-CM (0.125 ml) was neutralized with anti-hCCL20 mAb (25 μg) in 1 ml of culture medium or the same amount of mouse IgG, added to the cell culture and incubated for 6 h. MUC5AC mRNA expression was assessed by Q-RT-PCR.