Copper-associated hepatitis in the Labrador retriever : diagnosis, treatment and genetics

Abstract

Copper is an essential trace element and is indispensable for a large number of important biological processes in the body. The liver is important for copper metabolism and is the first organ to be affected in copper overload. Hepatic copper accumulation results from excessive intake and/or from inherited aberrant copper metabolism. Hereditary disorders of copper metabolism are observed in humans, dogs and other mammals.

Copper-associated hepatitis is diagnosed with increasing frequency in the Labrador retriever population. The disease is characterized by a long subclinical phase which hampers early diagnosis and effective treatment. The diagnosis is made by histological evaluation of a liver biopsy. In the search for a non-invasive alternative to liver biopsies, copper- and zinc urinalysis was evaluated for its utility in identifying high hepatic copper concentration in dogs. Urinary copper/zinc ratio in a single urine sample in Labrador retrievers was significantly associated with hepatic copper concentration , but cannot be used as a single test to predict hepatic copper concentration in Labrador retrievers due to overlap between normal and affected dogs.

Treatment for copper-associated hepatitis consists of copper chelation therapy and restriction of dietary copper intake. Based on estimates from modelling studies in Labrador retrievers treated with D-penicillamine, optimal duration treatment can now be predicted and depends on severity of copper accumulation. Lifelong continuous D-penicillamine treatment is not recommended in this breed, as there is a risk for copper and zinc deficiency.

Dietary copper and zinc levels are important in the pathogenesis of copper-associated hepatitis. Hepatic copper concentrations were respectively positively and negatively correlated with dietary copper and zinc intake. Therefore, an adjusted diet (low-copper, high-zinc) in the long-term management of affected dogs after initial chelation therapy was investigated. A wide variation in re-accumulation rate of hepatic copper concentration was observed. The majority of dogs could be maintained on a safe hepatic copper concentration with dietary adaption alone, whereas some dogs re-accumulated within 6 months. Therefore, control liver biopsies should be performed for evaluation of effect of diet. Dietary adaption to a low-copper/high zinc diet was investigated in Labrador retrievers with subclinical hepatic copper accumulation. In approximately half of the dogs, hepatic copper could be normalized with diet alone, whereas other dogs continued to accumulate copper or failed to respond to the diet. This response seemed to be in part depending on the family history of individual dogs.

Finally a genome wide association analysis was used to identify genes involved in hepatic copper accumulation in the Labrador retriever. Two chromosome regions were associated with variation in hepatic copper levels. Subsequent DNA sequence analysis identified missense mutations in two candidate genes that were validated in an independent replication cohort. Functional follow-up studies of the 2 mutations proved their involvement in the disease pathogenesis.

In conclusion, results described in this thesis contribute to better diagnostic and therapeutic protocols for copper-associated hepatitis in Labrador retrievers. The identification of two genes will have major implications for understanding the biological background of the disease and for development of new therapies.