Role of Drp1 in cigarette smoke extract-induced C2C12 myoblast senescence and mitochondrial dysfunction

Sarcopenia is a frequent comorbidity of chronic obstructive pulmonary disease (COPD), which is strongly associated with prolonged cigarette smoke exposure. This study investigated the effects of cigarette smoke extract (CSE) on an immortalized murine C2C12 myoblast model, focusing on apoptosis, proliferation, cellular senescence, and mitochondrial function. C2C12 myoblasts were exposed to graded concentrations of CSE (0.75%, 1.5%, and 3%). Cell viability and proliferation were assessed by Cell Counting Kit-8 (CCK-8) and ethynyl-2'-deoxyuridine (EdU) assays, respectively. Apoptosis was quantified by flow cytometry, and cellular senescence was evaluated by SA-β-galactosidase staining. Mitochondrial membrane potential (ΔΨm) was assessed by tetramethylrhodamine ethyl ester (TMRE) staining, and mitochondrial fluorescence was evaluated using MitoTracker staining. Drp1 protein expression was determined by Western blotting, while Drp1, Fis1, Mff, and Mitofusin 1/2 mRNA levels were analyzed by RT-qPCR. To assess the contribution of Drp1 expression, a rescue experiment was performed using Drp1 overexpression. CSE exposure reduced C2C12 cell viability and proliferation, increased apoptosis and SA-β-Gal-positive cells, lowered ΔΨm, and reduced MitoTracker-positive fluorescence. Drp1 protein expression and Drp1, Fis1, and Mff mRNA levels decreased, whereas Mitofusin 1/2 mRNA levels increased. Drp1 overexpression attenuated CSE-associated senescence and mitochondrial dysfunction. These findings indicate that altered Drp1 expression is associated with CSE-induced myoblast senescence and mitochondrial homeostatic disruption in vitro.