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. Author manuscript; available in PMC: 2022 Feb 1.
Published in final edited form as: Cell Signal. 2020 Nov 17;78:109838. doi: 10.1016/j.cellsig.2020.109838

Fig. 6. Effect of SACLAC exposure on mitochondrial bioenergetics in drug-resistant leukemia.

Fig. 6.

HL-60/dnr cells (resistant to daunorubicin) (800,000/ml dnr-free, RPMI-1640 medium, 10% FBS) were exposed to 10 μM SACLAC (DMSO vehicle) or DMSO (control) for 24 hr. Cells were then harvested by centrifugation, washed three times in PBS, and subjected to bioenergetic characterization. For respiration experiments, cells were suspended in a potassium-based respiration buffer, permeabilized with digitonin (10μg/mL), and energized with various carbon substrates and respiratory stimuli/inhibitors. All data were normalized to live cell count. A. Cell viability. B. Respiration under basal conditions, as well as in response to digitonin (Digi, 10μg/mL), pyruvate/malate (Pyr/M, 5mM/1mM), cytochrome c (Cyt C, 10μM), CK clamp (CK 20U/mL; phosphocreatine, PCR, 1mM; ATP, 5mM), octanoyl-carnitine/glutamate/succinate (O/G/S, 0.2mM/5mM/5mM); and multiple PCR additions to titrate ATP free energy (ΔGATP) across a physiological span. C. Relationship between oxygen consumption (JO2) and ATP free energy (ΔGATP). Calculation of ΔGATP done using the online resource https://dmpio.github.io/bioenergetic-calculators/ck_clamp/. D. Respiration under basal conditions, as well as in response to digitonin (Digi, 10μg/mL), Pyr/M/O/G/S (Multi), cytochrome c (Cyt C, 10μM), and FCCP titration (0.5–3.0μM). Rotenone (0.5μM) and antimycin A (0.5μM) were added at the end to control from any non-mitochondrial respiration. Data are mean ± SEM, N=3/group, *P<0.05.