Role of Caveolin-3 in lipid uptake and accumulation in muscle cells
Aims: Caveolae are small specialized invaginations of the plasma membrane enriched in proteins of the caveolin family. Caveolin-3 is exclusively expressed in mature skeletal muscle and was shown to interact with proteins fatty acid translocase (FAT/CD36), β-adrenergic or insulin receptor. Intramuscular lipid accumulation is associated with insulin resistance in type 2 diabetes, and high concentrations of extracellular fatty acids. To determine whether caveolin-3 is involved in lipid metabolism in muscle cells, lipid uptake and accumulation was measured before and after in control and cav-3 siRNA transfected myotubes. In the same experiments the fatty acid transporter genes were investigated.
Methods and results: Differentiated C2C12 myotubes were transfected with specific cav-3 siRNA. Both, control and transfected cells were exposed to 400µM palmitate for 24 hours to stimulate lipid droplets (LD) formation. In addition, forskolin-induced lipolysis was investigated. Fatty acid accumulation (Oil Red O fluorescence) and caveolin-3 localization was determined using fluorescence microscopy. Fatty acid uptake was measured by quantitative determination of Non-Estrified Fatty Acids (NEFA) in culture medium. Immunolocalization of cav-3 demonstrated its localization in the plasma membrane. After palmitate stimulation caveolin-3 and FAT/CD36 gene expression did not change. However, stimulation of myotubes with palmitate resulted in a marked translocation of caveolin-3 from the plasma membrane to intracellular compartments. Moreover, knockdown of cav-3 expression with siRNA increased fatty acid accumulation by 7.0- fold, p < 0.05 vs. control. In consonance, forskolin-induced lipolysis was significantly reduced in cav-3 siRNA transfected myotubes.
Conclusions: Taken together, these results support the idea that caveolin-3 is required for the regulation of muscular fatty acid metabolism. Future studies will be necessary to investigate the molecular pathways through which caveolins regulate muscle cell function.