Insulin transport to mitochondria

PASSICOT G; CAMBEROS MC; CRESTO JC

Octubre 13-17, Mar del Plata, Argentina

Congreso; XIX Congreso Anual de SLEP; 2007

Sociedad Latinoamericana de endocrinología pediátrica

Objectives: Mitochondria are insulin-dependent organelles because insulin regulates the amount required of oxidative substrates and control the synthesis and activity of some key enzymes. We previously found the insulin-degrading enzyme (IDE) in the mitochondrial matrix (MM). Our objective was to determine if IDE facilitates the insulin transport to MM. Methods: IDE was extracted from rat skeletal muscle homogenates after successive chromatographic steps. Active mitochondria from hepatic source were isolated with Parson’s method.  Mitochondria were recovered and studied at equilibrium at different times with 100% oxygen and insulin.  Substrates or inhibitors of IDE were added in agreement with the experimental design. The reaction was stopped at 2 ºC with 1 mM N-ethylmaleimide. The MM was isolated after digestion with digitonin. Insulin transport and degradation were studied by electrophoresis in SDS-PAGE, gel chromatography, inmunoblot and autoradiography. Results: Insulin incorporation in MM by IDE is doses/dependent. Dinitrophenol and vanadate decreases this incorporation in MM and succinate+ADP slightly decrease it. The insulin degradation by IDE is unapparent and the degraded insulin (pg/sec/mg of protein; control: 0.87; IDE: 0.88) is produced by mitochondrial proteases. The chromatography and autoradiography of  MM showed insulin incorporation through the mitochondrial transport system, acting IDE as transporter. The results strongly suggest that insulin is actively transported to MM. Conclusions: IDE induce the insulin transport to MM.