The Effects of Extracellular Vesicles From Cancer Cells in Acidosis or ER Stress to Endothelial Cells: Angiogenesis and Endothelial Activation.

Tumor-derived extracellular vesicles (EVs) facilitate metastatic progression by transferring bioactive cargo to recipient cells, thereby increasing endothelial permeability and promoting angiogenesis. The harsh conditions of the tumor microenvironment, such as extracellular acidosis, can alter the content of EVs, leading to changes in recipient cells, including endoplasmic reticulum (ER) stress. This study aimed to investigate whether EVs released by tumor cells adapted to acidosis or ER stress can alter endothelial functional properties. ER stress was induced in MDA-MB-231 breast cancer cells with tunicamycin, and adaptation to acidosis was achieved by culturing at pH 6.5 for 30 days. Large extracellular vesicles (LEVs) in these conditions were isolated by centrifugation. LEVs were characterized by nanoparticle tracking analysis (NTA), immunogold, Western blot, and high-performance liquid chromatography (HPLC). ER stress and autophagy markers were quantified by Western blot; cancer cells' adhesion to endothelial cells was assessed by adhesion assay; barrier permeability by dextran diffusion in an organ-on-a-chip model; migration by wound healing assay, and angiogenesis by tube formation assay. Tunicamycin was not detected in LEV from tunicamycin-treated cells (LEV T). LEV T increased the migration of endothelial cells, while not inducing ER stress, nor altering adhesion, angiogenesis, or permeability. Acidosis-adapted cells did not have greater UPR activation, but their LEVs (LEV 6.5) induced ER stress, reduced adhesion and angiogenesis, and increased permeability and migration. These findings indicate that LEVs from acidosis-adapted tumor cells reprogram endothelial physiology, supporting a potential role in metastatic progression.