Fig. 4. Effect of palmitate and/or unsaturated fatty acids on mitochondrial respiratory function. L6 myotubes were incubated for 16 h in media containing D-glucose (5 mM) supplemented with or without palmitate (PA, 0.4 mM) or unsaturated fatty acids palmitoleate (PO, C16:1), Oleate (OL, C18:1) or linoleate (LO, C18:2) at 0.4 mM as indicated prior to analysis of real-time cellular respiration using a Seahorse XF24 analyser. Oligomycin (1 μM), FCCP (1 μM) and a rotenone (1 μM)/antimycin-A (2 μM) mix were added at times indicated by the dotted lines. (A) shows a representative readout of oxygen consumption rate (OCR) from a single experiment with each data point representing the mean ± SD of a triplicate measurement. (B) The graphical data shows basal respiration, ATP-linked respiration and maximal respiration (OCR after FCCP stimulation). (C) Mitochondrial DNA quantification by qPCR. Data in (B) and (C) are mean ± SEM from 5 and 3 independent experiments, respectively. The asterisk indicates a significant change (P<0.05) to the control condition (lacking any fatty acid additions).