Irradiation leads to mitochondrial dysfunction in Caco-2 cells. Caco-2 cells were irradiated at the indicated doses, cultured for an additional 24 hours, and subjected to various mitochondrial functional assays. (A) A representative Seahorse plot of the Mito Stress assay and bar graphs of the respiratory parameters normalized within the experiment for difference in the total protein content between the groups (n = 6). (B) The median fluorescence intensity (MFI) of cells stained with MitoSOX and analyzed by flow cytometry is shown as relative fold inductions in relation to control nonirradiated cells (CT; n = 3). Cells treated with 5 μM antimycin-A for 3 hours served as a positive control for the assay. (C) ATP production in 1 μg of isolated mitochondria from nonirradiated (CT) and irradiated (10 Gy) Caco-2 cells when provided with pyruvate and malate as substrates. A representative plot of luciferase activity recorded over time (left) and the ATP concentration calculated are shown as relative fold change in comparison to CT (n = 4). ∗P < .05, ∗∗P < .01, ∗∗∗P < .001, and ∗∗∗∗P < .0001; n, number of independent experiments.