This study investigates the impact of hypoxia on exosome release by breast cancer cells. Exosomes, nanovesicles secreted by tumor cells, play a crucial role in paracrine signaling during tumor progression. The study found that breast cancer cell lines (MCF7, SKBR3, and MDA-MB 231) exposed to moderate (1% O2) and severe (0.1% O2) hypoxia showed significant increases in exosome release, as determined by nanoparticle tracking analysis (NTA) and immunoblotting for the exosomal protein CD63. The hypoxic regulation of miR-210 was also identified in hypoxic exosome fractions. The activation of hypoxic signaling by a HIF hydroxylase inhibitor, dimethoxyallylglycine, further increased exosome release, while transfection of HIF-1α siRNA prevented this enhancement. These findings suggest that hypoxia promotes exosome release by breast cancer cells, potentially through the HIF-1α pathway, with implications for tumor progression and invasion.This study investigates the impact of hypoxia on exosome release by breast cancer cells. Exosomes, nanovesicles secreted by tumor cells, play a crucial role in paracrine signaling during tumor progression. The study found that breast cancer cell lines (MCF7, SKBR3, and MDA-MB 231) exposed to moderate (1% O2) and severe (0.1% O2) hypoxia showed significant increases in exosome release, as determined by nanoparticle tracking analysis (NTA) and immunoblotting for the exosomal protein CD63. The hypoxic regulation of miR-210 was also identified in hypoxic exosome fractions. The activation of hypoxic signaling by a HIF hydroxylase inhibitor, dimethoxyallylglycine, further increased exosome release, while transfection of HIF-1α siRNA prevented this enhancement. These findings suggest that hypoxia promotes exosome release by breast cancer cells, potentially through the HIF-1α pathway, with implications for tumor progression and invasion.