Breast cancer cells exposed to hypoxia release more exosomes than cells under normoxic conditions. This study demonstrates that hypoxia enhances exosome release, which may be mediated by the HIF-1α oxygen sensing pathway. Exosomes are nanovesicles secreted by tumor cells that play a role in paracrine signaling during tumor progression, including tumor-stromal interactions, activation of proliferative pathways, and immunosuppression. Hypoxia, a key feature of solid tumors, promotes tumor progression, angiogenesis, and metastasis, potentially through exosome-mediated signaling. Breast cancer cell lines were cultured under moderate (1% O₂) or severe (0.1% O₂) hypoxia. Exosomes were isolated from conditioned media and quantified using nanoparticle tracking analysis (NTA) and immunoblotting for CD63, an exosomal marker. Hypoxic exosome fractions were analyzed for miR-210, a miRNA that is upregulated under hypoxia. The results showed that hypoxia significantly increased the number of exosomes in conditioned media. Activation of hypoxic signaling by the HIF hydroxylase inhibitor DMOG increased exosome release, while transfection with HIF-1α siRNA prevented this enhancement. Hypoxically regulated miR-210 was found at elevated levels in hypoxic exosome fractions. These findings suggest that hypoxia promotes exosome release by breast cancer cells, which may contribute to tumor progression by releasing more exosomes into the tumor microenvironment to support their survival and invasion. The study highlights the potential role of hypoxia in modulating exosome release and the importance of understanding hypoxic tumor phenotypes. The results have implications for understanding how hypoxia influences tumor progression through exosome-mediated signaling.Breast cancer cells exposed to hypoxia release more exosomes than cells under normoxic conditions. This study demonstrates that hypoxia enhances exosome release, which may be mediated by the HIF-1α oxygen sensing pathway. Exosomes are nanovesicles secreted by tumor cells that play a role in paracrine signaling during tumor progression, including tumor-stromal interactions, activation of proliferative pathways, and immunosuppression. Hypoxia, a key feature of solid tumors, promotes tumor progression, angiogenesis, and metastasis, potentially through exosome-mediated signaling. Breast cancer cell lines were cultured under moderate (1% O₂) or severe (0.1% O₂) hypoxia. Exosomes were isolated from conditioned media and quantified using nanoparticle tracking analysis (NTA) and immunoblotting for CD63, an exosomal marker. Hypoxic exosome fractions were analyzed for miR-210, a miRNA that is upregulated under hypoxia. The results showed that hypoxia significantly increased the number of exosomes in conditioned media. Activation of hypoxic signaling by the HIF hydroxylase inhibitor DMOG increased exosome release, while transfection with HIF-1α siRNA prevented this enhancement. Hypoxically regulated miR-210 was found at elevated levels in hypoxic exosome fractions. These findings suggest that hypoxia promotes exosome release by breast cancer cells, which may contribute to tumor progression by releasing more exosomes into the tumor microenvironment to support their survival and invasion. The study highlights the potential role of hypoxia in modulating exosome release and the importance of understanding hypoxic tumor phenotypes. The results have implications for understanding how hypoxia influences tumor progression through exosome-mediated signaling.