Fig. 4

NK cell mediated killing of HD-Ad gene transduced epithelial cells is blocked by Ruxolitinib and CAPE. NK-92 cells were deprived from IL-2 for 48 h and THP-1 cells were activated by 5 ng/ml PMA to prepare mature macrophages. Mature macrophages were harvested and co-cultured with NK-92 cells in presence of ruxolitinib or CAPE, and in presence and absence of both the inhibitors for 24 h. NK-92 cells alone were also cultured in presence of ruxolitinib or CAPE, and in presence and absence of both the inhibitors for 24 h. Both the cohorts of NK-92 and macrophages were transduced with C4HSU vector. Simultaneously, GFP-C4HSU transduced BEAS-2b cells were also cultured in presence of ruxolitinib or CAPE, and in presence and absence of both the inhibitors for 24 h. After culturing them for 24 h, NK-92 and macrophages were transferred to respective BEAS-2b cultures. After incubating those co-cultures for 5 h, BEAS-2b cells were harvested and stained for APC-Annexin V and 7AAD. Samples were run on flowcytometer and analysed by gating on GFP positive, gene transduced BEAS-2b cells. Percentages of 7AAD positive dead gene transduced BEAS-2b cells are depicted on upper right quadrant of panel (a). Percent dead cells from each row are depicted as separate bar diagram in panel (b) to better depict significant differences. Results from one experiment is depicted in the figure. The significance was calculated by one-way ANOVA with Tukey’s multiple comparison. *p < 0.05, **p < 0.01, ***p < 0.001