Figure 6
Pulmonary cells infected with RSV inhibit STAT3 phosphorylation and cytotoxic activity in IL-21-treated human CD8 T cells. Human pulmonary cells (MRC5) were infected with RSV (1 × 104 PFU/ml) for 1 h. Afterwards, the medium was replenished, and human CD8 T cells isolated from PBMCs were placed in coculture with the infected MRC5 cells and then stimulated with IL-21 (25 ng/ml) for 30 min. (A) Cropped blot bands of total phosphorylated STAT3 protein (on serine 727) in memory CD8 T cells analyzed by Western blot. β-Actin was used to normalize protein quantification. (B) pSTAT3 protein quantification from Western blot analysis. Data are shown as percentages over untreated/uninfected controls. Full-length blots are shown in Supplementary Fig. 4. (C) Human pulmonary MRC-5 cells (1 × 105 cells) were infected with RSV and cocultured in 96-well plates with human memory CD8 T cells (8 × 104 cells) stimulated with IL-21 (25 ng/ml) in RPMI with 2% FBS. Human CD8 T cell cytotoxicity against MRC-5 cells was assessed by lactate dehydrogenase detection in the coculture supernatant. Data are expressed as the mean ± SEM. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparison test. *p < 0.05, **p < 0.01, ***p < 0.001.
