Extracellular O₂ level and pH modulation affected the human breast cancer stem cells’ survival and stemness

Introduction: It has been reported that cancer stem cells could maintain their viability and stemness under certain extracellular changes. Therefore, efforts that modulate tumor hypoxia and acidity should be compelled in order to impede the growth of cancer stem cells. This study was aimed to analyze the effect of extracellular pH and O₂ level modulation on viability, apoptosis, and stemness of the human CD24−/CD44+ breast cancer stem cells (BCSCs). Methods: The primary BCSCs (CD24−/CD44+ cells) were cultured under hypoxia (1% O₂) or under supplementation of sodium bicarbonate (100 mM) for various periods. After each incubation time, cell viability was determined by trypan blue exclusion assay and apoptosis was examined using flow cytometry with Annexin V/PI assay. Furthermore, total RNA was isolated for qRT-PCR analysis of HIF1, ALDH1 and Klf4 mRNA expression. Results: This study demonstrated that hypoxia could suppress BCSC proliferation, but inhibit the cell apoptosis. Alkaline pH could also suppress BCSC proliferation and promote early apoptosis at the same time. Interestingly, the expressions of ALDH1 and KLF4 were downregulated in hypoxia-treated BCSCs which might be regulated through the increase of HIF1. Conversely, ALDH1 and Klf4 expressions were upregulated in sodium bicarbonate-treated BCSCs under alkaline pH. Conclusion: The modulation of extracellular pH and the O₂ level has diverse effects on the viability, apoptosis, dan stemness of BCSCs. Therefore, we suggest that targeting tumor hypoxia and acidity may be a prospective therapeutic strategy to eradicate BCSCs.