Fig. 5

IL30-dependent regulation of inflammation and immunity gene expression in murine and human PC cells. A Mouse Cancer Inflammation & Immunity Crosstalk PCR Array. Fold differences of mRNAs of inflammation and immunity-related genes between rmIL30-treated (red bars) and untreated wild-type TRAMP-C1 cells. A significant threshold of twofold change in gene expression corresponded to p < 0.001. Only genes with a fold change > 2 are shown. Experiments were performed in duplicate. The stripped box represents a scale break. The dashed line represents the twofold change cutoff. B Human Cancer Inflammation & Immunity Crosstalk PCR Array. Fold differences of mRNAs of inflammation and immunity-related genes between IL30KO-DU145 cells (blue bars) and wild-type cells. Results obtained from control NTgRNA-treated DU145 cells were comparable to those from wild type cells. A significant threshold of a twofold change in gene expression corresponded to p < 0.001. Only genes with a fold change > 2 are shown. Experiments were performed in duplicate. The stripped box represents a scale break. The dashed line represents the twofold change cutoff. C Human Cancer Inflammation & Immunity Crosstalk PCR Array. Fold differences of mRNAs of inflammation and immunity-related genes between IL30KO-PC3 cells (blue bars) and wild-type cells. Results obtained from control NTgRNA-treated PC3 cells were comparable to those from wild-type cells. A significant threshold of a twofold change in gene expression corresponded to p < 0.001. Only genes with a fold change > 2 are shown. Experiments were performed in duplicate. Stripped boxes represent scale breaks. The dashed lines represent the twofold change cutoff. D Venn diagram representing the genes that govern “Cancer Inflammation & Immunity Crosstalk” which are up- and/or downregulated by IL30 (treatment with rmIL30 or human IL30 gene knockout) in TRAMP-C1 (purple circle), DU145 (green circle) and PC3 cells (blue circle). Overlapping circles illustrate the sharing of IL30-regulated genes between different cell lines. E, F Western blot analysis of the expression of STAT3 α and β isoforms in EV-, IL30 gene-, NTgRNA-treated and IL30KO-DU145 (E) and PC3 (F) cells. Results obtained from control NTgRNA-treated and EV-transfected cells were comparable to those from wild-type cells. G Immunostaining of STAT3 in tumors developed in NSG mice, after s.c. implantation of IL30 gene-transfected, or deleted, DU145 cells, revealed that the cytoplasmic and nuclear expression of STAT3 was strong in IL30-overexpressing tumors, scanty in IL30-deficient tumors and moderate in wild-type tumors. Immunostaining of control tumors, developed after implantation of EV-transfected or NTgRNA-treated cells, was comparable to that of wild-type tumors. Magnification: × 400. H Graphic representation of the suppression of cytokine and chemokine expression in DU145 and PC3 cells following IL30 gene deletion by CRISPR/Cas9 technology. The most downregulated molecular mediators are represented with a larger font size. Both IL23A and IL12B, which form the heterodimeric cytokine IL23, were suppressed