Figure 3.

STING agonist cGAMP enhanced NK cell cytotoxicity to pancreatic cancer cells. (a) AsPC-1 cells were stimulated with 4 μg/mL cGAMP for 20 h and co-cultured with NK-92 cells for 4 h. Cell lysis was detected by the LDH assay. (b) Capan-2 cells were stimulated with 4 μg/mL cGAMP for 20 h and co-cultured with NK-92 cells. Cell lysis was detected by the LDH assay. (c) Flow cytometry analysis of the fusion protein expression of ULBP2/5/6 in AsPC-1 cells after stimulation with 4 μg/mL cGAMP. (d) Flow cytometry analysis of the fusion protein expression of ULBP2/5/6 in Capan-2 cells after stimulation with 4 μg/mL cGAMP. (e) The mRNA and protein levels of CCL5 were determined respectively using RT-PCR and ELISA assays in AsPC-1 cells after stimulation with 4 μg/mL cGAMP. (f) The mRNA and protein levels of CCL5 were determined respectively using RT-PCR and ELISA assays in Capan-2 cells after stimulation with 4 μg/mL cGAMP. (g) Flow cytometry analysis of CCR5 expression in NK-92 cells. (h) A schematic diagram of the Transwell migration assay of NK-92 cells. (i) Migration of NK-92 cells into the lower chamber of the Transwell containing the conditioned media from the AsPC-1 cell cultures with or without the stimulation with 4 μg/mL cGAMP, or anti-CCL5 antibody. (j) Both AsPC-1 cells and NK-92 cells were pre-stimulated with 4 μg/mL cGAMP for 20 h and then co-cultured for 4 h. Cell lysis was detected by LDH assay. (k) Both Capan-2 cells and NK-92 cells were pre-stimulated with 4 μg/mL cGAMP for 20 h and then co-cultured for 4 h. Cell lysis was detected by LDH assay. A control group of the cGAMP-treated tumor cells alone (no NK cells) were used as a baseline group. Data are shown as means ± SEM, and statistical significance was determined as ns: not significant, ***p < .001, **p < .01 and *p < .05.