Alterations in hepatocyte lysosomes in experimental hepatic copper overload in rats

Background: Although Wilson's disease is characterized by an accumulation of copper within hepatocyte lysosomes, the effects of excess copper on hepatic lysosomes are unknown. We studied the effects of excess copper on the structure, physicochemical properties, and pH of hepatocyte lysosomes using a rodent model. Methods: Rats were copper loaded with 0.125% copper acetate in water for 6 weeks. Copper was measured by atomic absorption spectrophotometry. Morphology was studied by electron microscopy. Lysosomal membrane fluidity was studied by fluorescence polarization, and lipid composition was determined by gas chromatography. Hepatocyte lysosomal pH was determined by flow cytometry. Results: Copper overload resulted in a 10-fold increase in hepatic copper. Hepatocyte lysosomes were enlarged and abnormally shaped with a 27-fold increase in copper, increased in vitro fragility, and decreased lysosomal membrane fluidity. Thiobarbituric acid reactive substances, a measure of lipid peroxidation, doubled in isolated lysosomal membranes. Polyunsaturated fatty acids increased, saturated fatty acids decreased, and membrane content of selected fatty acids was modified after copper overload. Lysosomal pH increased from 4.67 ± 0.02 to 4.87 ± 0.02. Conclusions: Copper overload causes alterations in lysosomal morphology, increases lysosomal fragility, decreases membrane fluidity, alters membrane fatty acid composition, and increases lysosomal pH. Copper catalyzed lipid peroxidation represents the likely mechanism for these alterations.