Figure 7.

Tumor-derived ITGA6-high exosomes are predominantly uptaken by ovarian fibroblasts. (A) Schematic representation of the experimental design. Exosomes derived from mouse forestomach carcinoma (MFC) cells (50 µg) were co-cultured with 2×10⁵ primary ovary cells. (B) Size distribution of tumor-derived exosomes determined by nanoparticle tracking analysis. (C) WB of ITGA6, ITGB2 and exosome markers expression in tumor-derived exosomes. (D) IF staining of primary ovary cells co-cultured with exosomes from MFC cells with EV or ARHGEF12. Scale bars, 50 µm. (E) Quantification of ovary cells uptake of tumor-derived exosomes in (D). Data shown are from 20 cells counted per condition. (F) WB of ITGA6 and ITGB2 expression in exosomes from MFC cells with EV, ITGA6 or ITGB2. (G) IF staining of primary ovary cells co-cultured with ITGA6/ITGB2-high exosomes. Scale bars, 50 µm. (H) Quantification of ovary cells uptake of tumor-derived exosomes in (G). Data shown are from 20 cells counted per condition. (I) IF staining of primary ovary cells co-cultured with exosomes from MFC cells. Fibroblasts were stained with α-SMA and macrophages were stained with F4/80. Scale bars, 50 µm. (J) Quantification of different types of ovary cells that uptake of tumor-derived exosomes in (I). Data represent the counts of 20 positively stained cells per condition. (K) IF staining of mouse ovarian fibroblasts stimulated with estradiol (E2) and co-cultured with exosomes from MFC cells. Fibroblasts were stained with α-SMA. Scale bars, 50 µm. (L) Quantification of mouse ovarian fibroblasts uptake of tumor-derived exosomes in (K). Data shown are from 20 cells counted per condition. (M) WB of the expression of ITGA6 and cancer-associated fibroblasts (CAF) markers α-SMA, FAP and PDGFRβ in ovarian fibroblasts co-cultured with tumor-derived exosomes. (N) Representative image of tumor masses. MFC cells mixed with ovarian normal fibroblasts (NF) or exosome-educated fibroblasts were implanted into animals (n = 6). (O) Tumor growth curves of xenografts in (N). (P) Quantification of tumor weights in (N). (Q) Concentration of ITGA6 in peripheral blood of GC patients with OM or PM (n = 20). Data are representative of three independent experiments with similar results. Data were presented as mean ± SEM. n.s., no significant, ***P < 0.001, by two-tailed Student's t-test.