N-Acetylcysteine, Dichloroacetate, and Metformin Restore Mitochondrial Homeostasis by Counteracting Oxidative Stress and Fusion-Fission Imbalance in Palmitic Acid-Induced Lipotoxicity.

Background

Palmitic acid (PA)-driven lipotoxicity in skeletal muscle is associated with excessive reactive oxygen species (ROS) and disturbed mitochondrial dynamics. This study aimed to characterize PA-induced alterations in oxidative status and fusion-fission balance in C2C12 skeletal muscle cells and to test whether N-acetylcysteine (NAC), dichloroacetate (DCA), or metformin mitigate these changes.

Methods

Differentiated C2C12 myotubes were exposed to PA under conditions detailed in the Methods. Intracellular ROS was quantified, antioxidant defenses were assessed by activities of catalase (CAT), glutathione peroxidase (GPx), and superoxide dismutase (SOD), and mitochondrial dynamics were evaluated by expression of the fusion protein mitofusin 1 (MFN1) and the fission protein dynamin-related protein 1 (DRP1), together with morphological assessment of mitochondrial fragmentation.

Results

PA exposure increased ROS and was accompanied by decreases in CAT, GPx, and SOD activities. PA shifted mitochondrial dynamics toward fission, with reduced MFN1, elevated DRP1, and increased mitochondrial fragmentation. Co-treatment with NAC, DCA, or metformin attenuated PA-induced ROS accumulation, improved antioxidant enzyme activities relative to PA alone, and partially normalized MFN1 and DRP1 expression, with reduced fragmentation.

Conclusions

In an in vitro C2C12 model, PA-induced lipotoxicity is associated with oxidative stress and a fusion-fission imbalance favoring mitochondrial fragmentation. NAC, DCA, and metformin mitigate these alterations and help preserve mitochondrial homeostasis. These findings support targeting oxidative stress and mitochondrial dynamics as a potential approach to counteract mitochondrial dysfunction under lipotoxic conditions.