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. 2020 Dec 9;21(24):9367. doi: 10.3390/ijms21249367

Figure 3.

Figure 3

DCA induces ROS production in hypoxic cells by overloading the mitochondrial metabolism. EMT6 and 4T1 cells were treated with DCA overnight at indicated concentrations, and N-acetyl-cysteine (NAC) (10 mM) was added 1 h prior to and during treatment. ROS generation was measured by flow cytometry using CM-H2DCFDA probe under aerobic (a) and hypoxic (b) conditions. (c) Oxygen consumption rate (OCR) measurements were obtained over time by a Seahorse analyzer using the mitochondrial stress test. To extract detailed information on the electron transport chain in mitochondria, specific inhibitors consisting of oligomycin, FCCP, rotenone and antimycin A were added sequentially. Basal mitochondrial OCR, ATP-linked OCR and maximal OCR were calculated from these results. (d) NAD(P)H was measured by flow cytometry using a JZL1707 NAD(P)H sensor under aerobic and hypoxic conditions. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.