Fig. 5
Validation assays show that TAb2 and TCh3 tumors upregulated distinct factors. A Representative images of cytokine array analysis of supernatants of TCh3 (left) and TAb2 (right). Red boxes indicate the cytokines that were visually different between TCh3 and TAb2 tumors. B Quantification of signal differences in cytokine array analysis indicated by the red boxes in (A). Left panel: bar graph showing the signal differences between TCh3 and TAb2 tumors. Cytokines shown vertically on the y-axis and signal intensity shown horizontally on the x-axis (positive values for upregulation in TAb2 and negative values for upregulation in TCh3, respectively). Right panel: volcano plot showing adj. p-value of the differences on the y-axis and signal intensity shown horizontally on the x-axis. C Validation of cytokine expression by ELISA. Expression of CXCL16, CXCL17, CSF1 (a.k.a. MCSF), HGF, and CXCL12 from TAb2 or TCh3 cell lysate or supernatant were detected by ELISA. The optical density (OD) at 450 nm is shown on the y-axis and the dilution factor (5-fold serial dilution) is shown on the x-axis. Results are representative of experiments done in duplicates. D TAb2 tumors increased the expression of phosphorylated STAT3 (p-STAT3). Western blotting analysis of total STAT3 and p-STAT3 (Tyr705) in TAb2 and TCh3 cell lysates. GAPDH was used as loading control. Data are representative of at least three independent experiments. E Kaplan-Meier plots of 10-year survival in PIK3CAAmpTP53Mutated HNSCC patients (n = 300) expressing different levels of VEGF-C or both CSF1 and VEGF-C. Patients were grouped into high-expression group or low-expression group based on gene expression as described in Methods