Copper-associated hepatitis in dogs; pathogenesis, diagnosis and treatment

Abstract

Copper is an essential trace element for living organisms, but can have deleterious consequences when present in excess. Because the liver has a central role in copper metabolism, this is the predominant organ affected. Copper-accumulating disorders are recognized as hereditary diseases in man and other mammals including several dog breeds. In dogs, copper-associated hepatitis is one of the most common causes of primary hepatitis. Due to a long subclinical stage of disease, dogs usually present late in disease when liver function is severely impaired and treatment is less effective. One of the affected dog breeds, studied in the current thesis, was the Labrador retriever. Labradors with high hepatic copper concentrations but without appreciable histopathological signs of liver injury already had differential expression of multiple signalling pathways, including those associated with inflammation. This underlines the primary role of copper in the disease progress. The diagnosis of copper-associated hepatitis and of parenchymal liver diseases in general, is made by histological evaluation of a liver biopsy. Sensitivity of liver enzymes, alanine aminotransferase (ALT) and alkaline phosphatase, were found to be too low to adequately detect dogs with primary liver disease. Therefore new biomarkers, such as serum microRNAs, were studied for their ability to detect liver injury. Levels of a liver specific microRNA were increased in Labradors with liver injury but normal ALT and sensitivity to detect liver injury was higher compared to that of ALT. However, the microRNAs under study were not able to point toward the underlying liver disease. Therefore, the next step was to characterize a microRNA profile that can help to distinguish between several canine hepatobiliary diseases. When measuring six different microRNAs in dogs with parenchymal, biliary, vascular, neoplastic hepatobiliary diseases, and in healthy dogs it was shown that each disease group had different levels of these microRNAs. These results confirm that different microRNA biomarkers can be used to detect a specific hepatobiliary disease. This is the first step in developing a microRNA profile that can be used in the clinic. Because copper accumulation was not correlated with liver enzyme activity or with the existence of hepatocellular injury, copper specific biomarkers still need to be identified. The copper binding proteins CCS and SOD1 in erythrocytes were evaluated in Labrador retrievers for the association with hepatic copper concentrations. Both CCS and CCS/SOD1 ratio were significantly lower in Labrador retrievers with high hepatic copper concentrations compared to Labradors with normal copper concentrations. CCS and SOD1 are new biomarkers with the potential to identify dogs at risk in an early stage of disease and to monitor copper levels during treatment. Once dogs with hepatic copper accumulation have been identified, treatment is aimed at lowering intrahepatic copper levels by copper chelation therapy with D-penicillamine and dietary copper restriction. Based on Labrador retrievers treated with D-penicillamine, necessary treatment duration to obtain normalization of hepatic copper concentrations can now be predicted for individual dogs. In Labrador retrievers lifelong continuous D-penicillamine treatment is not recommended, as they face the risk of hepatic copper and zinc deficiency.