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. Author manuscript; available in PMC: 2017 Sep 15.
Published in final edited form as: Inflamm Bowel Dis. 2016 Jan;22(1):55–67. doi: 10.1097/MIB.0000000000000592

FIGURE 6.

FIGURE 6

FL3 and FL37 preserve mitochondrial function during TNFα treatment. Caco2-BBE cells were pretreated with 10 nM FL3 or FL37 for 1 hour, followed by treatment with 10 ng/mL TNFα or 50 ng/mL IFNγ for 16 hours. A, ATP concentration. B, Mitochondrial ROS levels were measured using mitoSOX dye; n = 8 per treatment across 2 separate experiments. C, 15 minutes before collection, cells were treated with 5 μM rotenone, a complex I inhibitor, or 1 μM antimycin A, a complex III inhibitor. Mitochondrial ROS levels were measured using mitoSOX dye; n = 8 per treatment. D, Mitochondrial complex I activity was determined using the Mitochondrial Dipstick Assay kit. Activity of no tx + vehicle control cells was set to 100%. E, Before pretreatment with FL3 or FL37, cells were transfected with siPHB or siNegative control (siNC) for 72 hours. Mitochondrial ROS levels were measured using mitoSOX dye. Total protein was analyzed by Western blotting to ensure efficiency of PHB knockdown by siPHB; *P < 0.05 versus no tx + vehicle; #P < 0.01 versus TNFα + vehicle; †P < 0.05 versus TNFα + vehicle + rotenone.