Fig. 5

Gastric cancer cell-derived exosomes induced neutrophil activation through interaction of HMGB1 and TLR4. a. LC-MS/MS proteomic analyses for exosomes from gastric cancer cell lines (BGC-823, MGC80-3, and SGC-7901). b. The expression of HMGB1 in gastric cancer cell-derived exosomes was verified by western blot. c-h. Neutrophils were pre-treated with HMGB1 antagonist (Gly) or TLR4 inhibitor (TAK) followed by treatment with BGC-Ex. Western blot assays for the expression of LC3-II in neutrophils (c). The expression of ATG7 and BECN1 genes in neutrophils was determined by qRT-PCR (d). Flow cytometric analyses for apoptosis and CD11b expression in neutrophils (e). The expression of phosphorylated NF-κB, STAT3, and ERK in neutrophils was determined by western blot (f). QRT-PCR analyses of pro-inflammatory factor gene expression (IL-1β, IL-6, IL-8, OSM, and TNFα) in neutrophils (g).Transwell migration assays for gastric cancer cells following treatment with supernatant from neutrophils (h). ^**P<0.01 and ^*P<0.05 compared to control (Ctrl); ^##P<0.01 and ^#P<0.05 compared to BGC-Ex