Figure 2
From: The CPT1a inhibitor, etomoxir induces severe oxidative stress at commonly used concentrations
ETO inhibits oxidative glutamine metabolism in CD28-costimulated T cells. (A) Activated T cells (day 3) were switched to cell culture medium containing varying levels of glucose and glutamine. After 48 hrs, live cells were enumerated using bead-based flow cytometry, as described in the materials and methods. Representative values from two independent experiments with separate donors are shown. (B) T cells were expanded following stimulation with dynabeads. After 7 days, the cells were transferred to bicarbonate-free XF assay medium containing varying concentrations of glutamine. Metabolic parameters were measured by extracellular flux assay (Seahorse). The oxygen consumption rate (OCR) was measured at baseline and following the addition of 1.5 μM oligomycin, 1.5 μM FCCP, and 50 μM ETO. Values are means ± S.E.M. from 3 independent experiments. (C) T cells undergoing logarithmic expansion (day 7) were treated with either 10 mM 13C6 glucose or 2 mM 13C5 glutamine for 1 hr. Mass isotopomer data for acetyl-CoA are shown. Values represent means ± S.E.M. from 3 independent experiments. (D) T cells undergoing logarithmic expansion (day 7) were treated with 13C16 palmitate +/−10 μM BPTES for 2 hrs. Mass isotopomer data for acetyl-CoA are shown. Values represent means ± S.E.M. from 3 independent experiments.
