Fig. 5

HIF-1α is directly involved in the regulation of chemotherapeutic drug-induced, ROS-mediated ACE2 expression. A Western blot and qRT-PCR analysis showed HIF-1α expression in four breast cancer cells after exposure to 0.05 µM EPI, 500 µM NAC, 0.05 µM EPI + 500 µM NAC for 72 h or 100 µM H₂O₂ for 4 h. B Western blotting analysis of ACE2 and HIF-1α expression in parental and resistant breast cancer cells cultured under normoxic or hypoxic (1% O₂ for 24 h) conditions. The relative expression of proteins is quantified by grayscale analysis and is shown below the bands. C Knockdown of HIF-1α in MDA-468 cells or D inhibition of HIF-1α by YC-1 inhibitor blocked the EPI-induced elevation of ACE2 in breast cancer cells under normoxia or hypoxia. The relative expression of proteins is quantified by grayscale analysis and is shown below the bands. E Western blot analysis showed that both knockdown and inhibition of HIF-1α expression significantly decreased ACE2 expression in drug-resistant breast cancer cells (468/EPR cells). β-actin was used as an internal reference protein. F Western blot analysis showed no change in the protein level of HIF-1α after knockdown of ACE2 in 468/EPR cells and overexpression of ACE2 in MDA-468 cells. β-actin was used as a loading control