Role of selenium supplementation in the prevention of copper-induced hepatocellular damage in Fischer 344 rats

Copper (Cu) toxicosis associated with consumption of excess dietary Cu is a well recognized condition in sheep and may have a role in Cu-associated liver diseases in children. The pathogenesis of Cu-induced hepatocellular damage is unknown. Three experiments were conducted in the Fischer 344 rat, a strain that has been proposed as a suitable model to investigate Cu-induced liver disease. The first experiment was designed to investigate the hepatotoxic effects of long-term exposure to excess dietary Cu and the role of selenium (Se) supplementation in the prevention of lipid peroxidation. Weanling male Fischer 344 rats were fed a diet containing 500 ppm Cu with normal Se levels (0.35 ppm), supplemented with 2 ppm Se, or in a Se-deficient diet (<0.2 ppm). Controls included normal diet (18 ppm Cu), Se-supplemented diet, and Se-deficient diet. Five rats per group were killed after 3, 6, and 12 months and their livers were removed for histology, histochemistry, Cu and Se analysis, glutathione peroxidase measurement, and quantification of malondialdehyde (MDA). Copper accumulation was low and morphologic changes were mild in all Cu-loaded rats. Evidence of lipid peroxidation was only observed as an age-related change in all rats after 12 months. In the second experiment, male Fischer 344 rats were fed diets containing 18 (control), 750, 1000, 1250, 1500, and 2000 ppm Cu and killed after 3 months in order to determine the amount of excess dietary Cu necessary to induce hepatocellular injury in this strain. Livers were removed and processed for histology, histochemistry, Cu analysis, and MDA measurement. Hepatic Cu levels were significantly higher in rats receiving more than 1000 ppm. The severity of the histological lesions became more prominent as levels of dietary Cu increased, being significant (p < 0.01) in rats receiving 1,250 ppm Cu and over. However, no significant differences were found for MDA levels between groups. The third experiment was developed to determine the optimal sampling time for evaluating liver damage, and the role of Se supplementation in the prevention of lipid peroxidation. Rats receiving excess (2,000 ppm) dietary Cu with normal Se levels, supplemented with 2 ppm Se, and Se-deficient diet were killed after 4, 8 and 12 weeks. Controls (normal Cu and Se, Se-supplemented and Se-deficient) were also included. Significant morphologic changes (necroinflammatory foci) were observed in all Cu-loaded rats. Apoptosis, which may be related to DNA damage, was a prominent feature in all Cu-loaded rats. These studies demonstrate that compared to other species, very high levels of excess dietary copper are needed to induce liver injury in Fischer 344 rats. This may be due to Cu resistance. Fischer 344 rats fed 1,250 to 2,000 ppm Cu developed morphologic evidence of Cu toxicity without increase in hepatic MDA content suggesting that lipid peroxidation does not play a major role in the pathogenesis of Cu-induced liver injury in the Fischer 344 model. Selenium supplementation did not prevent hepatocellular damage. Further studies are needed to investigate the major mechanisms by which Cu induces hepatocellular damage and to clarify the role of DNA damage in the pathogenesis of Cu-associated liver diseases.