Fig. 5.
Pharmacological and genetic inhibition of CFTR both increased IFN-γ production in human Vδ2 T cells and Vδ2-mediated cytotoxicity to human K562 leukemia cells. a Flow cytometry analysis of in vitro-expanded human Vδ2 T cells treated with anti-CD3 (10 μg/ml) and anti-CD28 (1 μg/ml) antibody stimulation for 6 h in the presence of the CFTR inhibitor CFTRinh172 (5 μM) or vehicle DMSO was performed to detect IFN-γ production (n = 6, paired t-test, **p < 0.01). b In vitro-expanded human Vδ2 T cells were pretreated with the CFTR inhibitor CFTRinh172 (5 μM) or vehicle DMSO for 2 h and then cocultured with DIR-labeled K562 cells at the indicated ratio. Six hours later, dead K562 cells were identified by PI staining (n = 3, **p < 0.01, ****p < 0.0001). c Flow cytometry analysis of in vitro-expanded human Vδ2 T cells infected with scramble shRNA or CFTR-specific shRNA lentiviral particles was performed. After 72 h of infection, the cells were stimulated with 5 μg/ml plate-bound anti-human CD3 antibodies and 1 μg/ml soluble anti-human CD28 antibodies before IFN-γ production was detected (upper panels, n = 3, **p < 0.01, ***p < 0.001). The cells were stained with anti-CFTR and PE-anti-mouse IgM (polyclonal) antibodies and analyzed by FACS. Histograms of CFTR expression (gating on GFP+CD3+ cells) and the MFI of CFTR are shown (lower panels, black line/number: scramble shRNA; red line/number: CFTR-specific shRNA; dotted line: isotype control. n = 3, ***p < 0.001)