Fig 1: Tumor-secreted PLAU promoted the formation of inflammatory CAFs, which secreted IL8 via phosphorylation of Akt/NF-?B.A Transcriptional profile of NFs and CAFs with or without recombinant PLAU treatment. B Recombinant PLAU promoted activation of cytokine, chemokine, and interleukins pathways and NF-?B canonical pathways as demonstrated by GSEA. C RT-qPCR detection of markers of iCAFs and myCAFs in CAFs with or without recombinant PLAU treatment. D A 48-cytokine panel was used to characterize the CM of CAFs with or without recombinant PLAU treatment. E, F Detection of IL8 expression in CM of CAFs with or without recombinant PLAU treatment in the presence or absence of IPR-803, a uPA-uPAR inhibitor, by ELISA (E) and western blot (F). G–J Experimental scheme (G) and detection of IL8 expression in CM of indicated CAFs cocultured with PLAU overexpressing and vector KYSE-450 cells (H, J) or treated with CM of PLAU overexpressing and vector KYSE-450 cells (I, J), in the presence or absence of IPR-803 as assessed by ELISA (H–I) and western blot (J). The CM used for western blots was concentrated 40-fold. K Top 20 predicted transcription factors of CXCL1 predicted by the Cistome website. L Detection expression of uPAR, total and phosphorylated Akt and p65 of indicated CAFs. Three biological replicates were performed for in vitro assays. Data in bar charts are presented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (Student’s t-test).
Fig 2: IL8 secreted by CAFs upregulated PLAU expression in tumor cells.A, B Detection of PLAU expression of WT KYSE-30 and KYSE-450 cells cocultured with CAFs treated with recombinant uPA in the presence or absence of IPR-803, a uPA-uPAR inhibitor, as assessed by western blot (A) and RT-qPCR (B). C–F Western blot and RT-qPCR were performed to detect PLAU expression of WT KYSE-30 and KYSE-450 cells exposed to recombinant IL8 treatments at different times (C, E) and concentrations (D, F). G–I Detection of PLAU expression in WT KYSE-30 and KYSE-450 cells with recombinant IL8 treatment in presence or absence of SB225005, a CXCR2 inhibitor, by western blot (G), RT-qPCR (H), and ELISA (I). J Correlation analysis of IL8 and PLAU expression based on GSE53625 data. Three biological replicates were performed for in vitro assays. Data in bar charts are presented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (Student’s t test).
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