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. 2022 Mar 9;41(16):2372–2389. doi: 10.1038/s41388-022-02253-6

Fig. 4. CAFR-derived IL8 activated the NF-κB signaling pathway to upregulate UPK1A-AS1.

Fig. 4

A Top 20 upregulated pathways were plotted based on the enriched gene ratio and p value in Panc-1 cells treated with CAFR1-CM compared to CAFS1-CM. B UPK1A-AS1 expression in Panc-1 and MiaPaCa-2 cells treated with IL8 (100 ng/ml) alone or IL8 and CAPE (2 μM) together for 3 days. C Western blot analysis of IκBα, p-IκBα, p65, and p-p65 protein expression in Panc-1 and MiaPaCa-2 cells treated with CAFR1-CM or IL8. A neutralizing antibody against IL8 was used to deplete IL8 in CAFR1-CM. D Western blot analysis of p65, and p-p65 protein expression in PDAC tissues from platinum-resistant patients and platinum-sensitive patients. E Luciferase reporter assays of Panc-1 cells transfected with a reporter plasmid containing the UPK1A-AS1 promoter, and treated with IL8 or p65 depletion. F, G A conserved p65-binding motif was predicted by JASPAR and schematic images of the potential p65 motif binding sites in the promoter of UPK1A-AS1 are shown. H ChIP analysis of the p65 occupancy at the promoter of UPK1A-AS1 in Panc-1 cells. I Luciferase reporter assays of Panc-1 cells treated with IL8 and transfected with reporter plasmids containing P1, P2 and P3 deletions in the UPK1A-AS1 promoter. The results are presented as the mean ± SD of three technical replicates. **P < 0.01; ***P < 0.001; ns no significance.