Electrical Pulse Stimulation Partially Restores Insulin Signaling in Palmitate-Induced Insulin-Resistant Skeletal Muscle Cells via Mitochondrial Remodeling and Contractile Activity-Dependent Secretome.

Lipid oversupply induces insulin resistance in skeletal muscle (SkM), partly by altering mitochondrial function and dynamics. Although electrical pulse stimulation (EPS) improves insulin signaling in myotubes, the mechanisms contributing to this effect under lipotoxic conditions remain unclear. Thus, we investigated whether EPS restores lipid-induced insulin resistance in association with changes in mitochondrial respiration and dynamics, lipotoxic intermediates and evaluated the contribution of contraction-induced secreted factors, including small extracellular vesicles (sEV). Murine C2C12 myotubes were treated with palmitate (PA, 0.3 mmol/l) and subjected to EPS (1 Hz, 11.5 V) for 24 h. Insulin signaling, mitochondrial dynamics, respiratory capacity and lipotoxic intermediates were assessed by Western blotting, high-resolution respirometry and liquid chromatography-tandem mass spectrometry in control (CON) and PA-treated cells before and after EPS. Additionally, CON and PA-treated cells were incubated for 24 h with conditioned media and sEV from unstimulated or EPS-stimulated CON and PA-treated cells to evaluate the effect of the myocellular secretome on AKT signaling. PA reduced insulin-stimulated AKT(Ser473) and (Thr308) phosphorylation, which was partially rescued by EPS. In PA cells, EPS decreased the fusion proteins mitofusin 1 and 2 (MFN1, MFN2) and increased the fission marker dynamin-related protein 1 (DRP1). However, mitochondrial respiration and lipid accumulation were unaffected by EPS. Furthermore, conditioned media and sEV from EPS-stimulated cells enhanced AKT phosphorylation in PA-treated cells. These findings demonstrate that attenuation of lipid-induced insulin resistance is independent of mitochondrial respiratory capacity and lipid accumulation, but it is associated with alterations in proteins involved in mitochondrial dynamics and is likely mediated by exercise-derived sEV.