Fig. 1
25KbPEI enhances the production of IFN-γ by NK cells without transcriptional activation. A Chem_NK cells were generated by treating 1 × 106 NK-92MI cells for 12 h with 5 μg/ml of 25KbPEI. B IFN-γ levels present in the co-culture medium of NK cells and target cancer cells was measured by ELISA. C IFN-γ levels in culture medium collected from C_NK and Chem_NK cells were measured by ELISA. Statistical analysis was performed using Student’s t-test (vs. C_NK and Chem_NK). D IFN-γ transcripts in Chem_NK and C_NK cells were measured by qRT-PCR. NK cells treated with PMA/ionomycin were used as a positive control for the analysis of IFN-γ transcripts [1]. Statistical analysis was performed using one-way ANOVA with Tukey’s multiple comparisons test (vs. C_NK). E Intracellular IFN-γ levels were quantified by flow cytometry after treatment of NK cells with 5 μg/ml 25KbPEI for the indicated times. Statistical analysis was performed using one-way ANOVA with Dunnett’s multiple comparisons test (vs. 0 h). F Intracellular IFN-γ levels were evaluated by flow cytometry following treatment for 12 h with 25KbPEI and incubation with cycloheximide (CHX) (100 μg/ml) for the indicated times. The IFN-γ level at each time point is represented relative to the level at CHX 0 h. Statistical analysis was conducted using two-way ANOVA with Sidak’s multiple comparisons test (vs. C_NK). All experiments were conducted at least three times. *P < 0.05; **P < 0.01; ***P < 0.001, and ****P < 0.0001