Hypoxia promotes the phenotypic change of aldehyde dehydrogenase activity of breast cancer stem cells.

Stable breast cancer cell (BCC) lines are valuable tools for the identification of breast cancer stem cell (BCSC) phenotypes that develop in response to several stimuli as well as for studying the basic mechanisms associated with the initiation and maintenance of BCSCs. However, the characteristics of individual, BCC-derived BCSCs varies and these cells show distinct phenotypes depending on the different BCSC markers used for their isolation. Aldehyde dehydrogenase (ALDH) activity is just such a recognized biomarker of BCSCs with a CD44(+) /CD24(-) phenotype. We isolated BCSCs with high ALDH activity (CD44(+) /CD24(-) /Aldefluor(pos) ) from a primary culture of human breast cancer tissue and observed that the cells had stem cell properties compared to BCSCs with no ALDH activity (CD44(+) /CD24(-) /Aldefluor(neg) ). Moreover, we found Aldefluor(pos) BCSCs had a greater hypoxic response and subsequent induction of HIF-1α expression compared to the Aldefluor(neg) BCSCs. We also found that knocking down HIF-1α, but not HIF-2α, in Aldefluor(pos) BCSCs led to a significant reduction of the stem cell properties through a decrease in the mRNA levels of genes associated with the epithelial-mesenchymal transition. Indeed, HIF-1α overexpression in Aldefluor(neg) BCSCs led to Slug and Snail mRNA increase and the associated repression of E-cadherin and increase in Vimentin. Of note, prolonged hypoxic stimulation promoted the phenotypic changes of Aldefluor(neg) BCSCs including ALDH activity, tumorigenesis and metastasis, suggesting that hypoxia in the tumor environment may influence BCSC fate and breast cancer clinical outcomes.