Wilson Disease Workup

The presence of Kayser-Fleischer rings and ceruloplasmin levels of less than 20 mg/dL in a patient with neurologic signs or symptoms suggest a diagnosis of Wilson disease. If a patient is asymptomatic, exhibits isolated liver disease, and lacks corneal rings, the coexistence of a hepatic copper concentration of more than 250 mg/g of dry weight and a low serum ceruloplasmin level is sufficient to establish a diagnosis. Therefore, in the absence of Kayser-Fleischer rings or neurologic abnormalities, a liver biopsy for quantitative copper determination is essential to establish the diagnosis of Wilson disease.

Genetic diagnosis

Linkage analysis has been used in family studies for presymptomatic testing; however, the multiplicity of mutations (>200 mutations of ATP7B have been identified) that require screening in individuals without affected family members is large, making such analysis impractical. Therefore, the use of molecular testing is currently limited to screening of family members for an identified mutation detected in the index patient.

Abdominal imaging

Computed tomography (CT) scanning, magnetic resonance imaging (MRI), ultrasonography, and nuclear medicine studies of the liver have been uninformative, with findings neither specific nor sensitive for Wilson disease.

Electrocardiography

Resting electrocardiographic abnormalities include left ventricular or biventricular hypertrophy, early repolarization, ST segment depression, T-wave inversion, and various arrhythmias.

Serum ceruloplasmin levels are low in newborns and gradually rise within the first 2 years of life. Approximately 90% of all patients with Wilson disease have ceruloplasmin levels of less than 20 mg/dL (reference range, 20-40 mg/dL). (Ceruloplasmin is an acute phase reactant and may be increased in response to hepatic inflammation, pregnancy, estrogen use, or infection.)

Falsely low ceruloplasmin levels may be observed in any protein deficiency state, including nephrotic syndrome, malabsorption, protein-losing enteropathy, and malnutrition. Ceruloplasmin levels may also be decreased in 10-20% of Wilson Disease gene heterozygotes, who do not develop Wilson disease and do not require treatment.

Urinary copper excretion

The urinary copper excretion rate is greater than 100 mg/d (reference range, < 40 mg/d) in most patients with symptomatic Wilson disease. The rate may also be elevated in other cholestatic liver diseases.

The sensitivity and the specificity of this test are suboptimal for use as a screening test; however, it may be useful to confirm the diagnosis and to evaluate the response to chelation therapy.

Hepatic copper concentration

This test is regarded as the criterion standard for diagnosis of Wilson disease. A liver biopsy with sufficient tissue reveals levels of more than 250 mcg/g of dry weight even in asymptomatic patients. Special collection vials are available to help avoid contamination.

A normal hepatic copper concentration (reference range, 15-55 mcg/g) effectively excludes the diagnosis of untreated Wilson disease. An elevated hepatic copper concentration may be found in other chronic hepatic (mostly cholestatic) disorders.

Mutation analysis is an especially valuable diagnostic strategy for certain well-defined populations exhibiting a limited spectrum of ATP7B mutations. Pedigree analysis using haplotypes based on polymorphisms surrounding the the ATP7B gene are also commercially available from specific clinical laboratories.^[7]

Radiolabeled copper testing directly assays hepatic copper metabolism. Blood is collected at 1, 2, 4, 24, and 48 hours after oral ingestion of radiolabeled copper (⁶⁴ Cu or⁶⁷ Cu) for radioactivity in serum. In all individuals, radioactivity promptly appears after absorption, followed by hepatic clearance. In healthy people, reappearance of the radioactivity in serum occurs as the labeled copper is incorporated into newly synthesized ceruloplasmin and released into the circulation.

Heterozygotes exhibit a slow, lower-level reappearance of radioactivity rather than the continued fall in radioactivity seen in persons with Wilson disease, but there may be considerable overlap between the 2 types of patients. Patients with Wilson disease, even those with normal ceruloplasmin levels, do not exhibit the secondary rise in radioactivity.

An algorithm for the diagnosis of Wilson disease adapted from the American Association for the Study of Liver Diseases (AASLD) Practice Guidelines is outlined below.

The cranial lesions observed on CT scans are typically bilateral and are classified into 2 general categories: (1) well-defined, slitlike, low-attenuation foci involving the basal ganglia, particularly the putamen, and (2) larger regions of low attenuation in the basal ganglia, thalamus, or dentate nucleus.

Widening of the frontal horns of the lateral ventricles and diffuse cerebral and cerebellar atrophy, which correlate histologically with widespread neuronal loss, have also been described. (See the image below.)

MRI of the brain appears to be more sensitive than CT scanning in detecting early lesions of Wilson disease. MRI studies have identified focal abnormalities in the white matter, pons, and deep cerebellar nuclei. These lesions, measuring 3-15 mm in diameter, are typically bilateral, appearing with low signal intensity on T1-weighted images and with high signal intensity on T2-weighted images, representing cell loss and gliosis. Other studies describe decreased signal intensity in the putamen and other parts of the basal ganglia, which may represent either copper or iron ferritin deposition.

A characteristic "face of the giant panda" sign has been described, formed by high signal intensity in the tegmentum (except for the red nucleus), preserved signal intensity of the lateral portion of the pars reticulata of the substantia nigra, and hypointensity of the superior colliculus.

Results from a study by Tarnacka et al indicated that relative to the thalamus, the basal ganglia are more sensitive to ongoing degenerative changes and portal-systemic encephalopathy in Wilson disease. The authors used proton magnetic resonance spectroscopy (MRS) in 37 patients with newly diagnosed Wilson disease to identify the pathomechanism of the disease's cerebral pathology, specifically looking at the globus pallidus and thalamus to assess cerebral metabolic changes in myoinositol, choline, creatine, N-acetyl-aspartate, lipid, glutamine, and glutamate levels and ratios.^[12]

The investigators speculated that N-acetyl-aspartate/creatine ratio reductions seen in hepatically and neurologically impaired patients in the study may have indicated an association between neurodegeneration and all presentations of Wilson disease. In addition, they suggested that observed decreases in myoinositol and choline and an increase in neurologic glutamate may have been due to portosystemic shunting.^[12]

Positron emission tomography (PET) scanning reveals a significantly reduced regional cerebral metabolic rate of glucose consumption in the cerebellum, striatum, and, to a lesser extent, in the cortex and thalamus.

PET scan analyses of patients with Wilson disease have also demonstrated a marked reduction in the activity of dopa-decarboxylase, indicative of impaired function of the nigrostriatal dopaminergic pathway.

These abnormalities improve with chelation therapy, indicating a reversible component of striatal neuron injury.

Electron microscopic studies on ultrathin sections reveal numerous electron-dense lysosomes and residual bodies. The elemental analysis in transmission electron microscopy with electron energy loss spectroscopy, and in scanning electron microscopy with energy dispersive x-ray analysis, shows copper-specific signals of electron-dense accumulations inside these dark lysosomes and residual bodies.

The electron microscopic detection of copper-containing hepatocytic lysosomes is helpful in the diagnosis of the early stages of Wilson disease, in addition to the quantification of hepatic copper by atomic absorption spectrophotometry.

Hepatic

The earliest changes detectable with light microscopy include glycogen deposition in the nuclei of periportal hepatocytes and moderate fatty infiltration. The lipid droplets, which are composed of triglycerides, progressively increase in number and size, sometimes resembling the steatosis induced by ethanol. Hepatocyte mitochondria typically exhibit heterogeneity in size and shape, with increased matrix density, separation of the normally apposed inner and outer mitochondrial membranes, widened intercristal spaces, and an array of vacuolated and crystalline inclusions within the matrix. With progression of disease, copper protein is sequestered in lysosomes and is visible as electron-dense pericanalicular granules.

Despite consistently elevated hepatic copper levels in patients with Wilson disease, histochemical staining of liver biopsy specimens for copper is of little diagnostic value. Early in the disease, copper distribution is primarily cytoplasmic and is not readily apparent with rhodamine or rubeanic acid staining.

The rate of progression of the liver histology from fatty infiltration to cirrhosis is variable, although it tends to occur by 1 of 2 general processes, either with or without hepatic inflammation. The histologic picture may be histologically indistinguishable from that of chronic active hepatitis. Pathologic features include mononuclear cell infiltrates, which consist mainly of lymphocytes and plasma cells; piecemeal necrosis extending beyond the limiting plate; parenchymal collapse; bridging hepatic necrosis; and fibrosis.

The histologic pattern is one of a macronodular or mixed micro-macronodular cirrhosis, with fibrous septa (containing predominantly types I and III collagen), bile ductule proliferation, and variable septal round cell infiltration. Hepatocytes at the periphery of the nodules frequently contain Mallory hyalin. One proposed mechanism implicates copper as the inducer of fibrogenesis.

Interestingly, hepatocellular carcinoma is exceedingly rare in patients with Wilson disease compared with patients with hemochromatosis. This may be attributable to the significantly shortened life expectancy in untreated patients with Wilson disease, which does not allow time for carcinoma to develop. An increasing number of case reports suggest that the incidence will likely increase with improved survival. It has been proposed that the diminished cancer risk is due to the relatively low inflammatory component in the pathogenesis of Wilson disease.

Neurologic

Observed gross anatomical changes include degeneration and cavitation, primarily involving the putamen, globus pallidus, caudate nucleus, and thalamus. Little correlation has been observed between the degree of neurologic impairment and the neuropathologic findings. The affected areas of the brain do not possess higher copper concentrations than the unaffected portions.