¹²⁵I-Insulin Internalization by Perfused Rat Liver: Comparison of its Subcellular Distribution with that of a Lysosomally Targeted Molecule, ¹²⁵I-Asialofetuin

Summary

The subcellular distribution of ¹²⁵I-insulin in the perfused rat liver was compared with the subcellular distribution of the lysosomally targeted asialoglycoprotein, ¹²⁵I-asialofetuin. The use of Percoll density gradient medium provided excellent separation of lysosomes from the subcellular membrane fractions. Following perfusion with ¹²⁵I-asialofetuin, a distinct peak of TCA-precipitable radioactivity could be observed in the lysosomal region of the gradient. In contrast, the gradient distribution of TCA-precipitable radioactivity following perfusion with physiological concentrations of ¹²⁵I-insulin was unimodal, the observed peak corresponding to the distribution of intracellular membrane marker enzymes. Leupeptin, an inhibitor of lysosomal proteolysis, inhibited the degradation of ¹²⁵I-asialofetuin but had no effect on ¹²⁵I-insulin degradation. In addition, leupeptin produced a marked increase in TCA-precipitable radioactivity in the lysosome rich region of gradients prepared from livers perfused with ¹²⁵I-asialofetuin. No such effect was observed following perfusion with ¹²⁵I-insulin. These findings are consistent with an initial localization of the internalized insulin molecule with the membraneous system of the liver cell rather than the lysosomal system.

Key-Words:

Insulin Internalization - Perfused Rat Liver - Lysosomes - Asialofetuin - Percoll