Essential trace elements, including chromium, copper, manganese, molybdenum, selenium, and zinc, are required for nutrition (1). Chronic liver diseases may alter trace element contents in some organs, including the brain. Among these trace elements, manganese is a ubiquitous transition metal excreted by the liver into the bile (2, 3). During cholestasis, blood manganese levels may rise with resultant toxicity. Indeed, neuropsychiatric symptoms are prominent in jaundiced patients. In addition, phenothiazine-derived drugs may potentiate manganese toxicity (3).
Biliary atresia is an idiopathic, localized, complete obliteration or discontinuity of the hepatic or common bile ducts at any point from the porta hepatis to the duodenum (4). Obstruction of bile flow leads to cholestasis, progressive fibrosis, and ultimately, cirrhosis. Hence, biliary atresia once was uniformly fatal. Now liver transplantation has provided an opportunity for a more felicitous outcome (5, 6). However, there is little information on manganese kinetics within the brains of patients with biliary atresia before and after hepatic transplantation. The present study was undertaken to investigate brain manganese accumulation in patients with biliary atresia before and after hepatic transplantation.
MATERIALS AND METHODS
Eight patients were diagnosed with biliary atresia and underwent hepatic portoenterostomy. The mean age at the time of diagnosis and surgery was 1.7 months. Clinical characteristics are shown in Table 1. The study patients were consecutive cases. No patients received intravenous nutrition. Neuropsychiatric symptoms were evaluated when the magnetic resonance imaging (MRI) studies were performed and after informed consent was obtained.
Measurement of serum aspartate transaminase (AST), alanine transaminase (ALT), lactate dehydrogenase, and total bilirubin.
Blood samples were examined for serum AST, ALT, lactate dehydrogenase, and total bilirubin levels. All levels were measured with a multianalyzer (Clinilizer, JCA-VX 1000; Nippon Denshi Co., Tokyo, Japan).
Measurement of blood manganese concentrations.
Blood concentrations of manganese were determined by graphite furnace atomic absorption spectroscopy (7). The normal range of blood manganese was <2.5 μg/dL.
MRI of the brain.
Standardized T1-weighted MRI scans (Magnetom Vision 1.5 tesla; Siemens, Munich, Germany) of the brain were performed in transaxial and sagittal planes. The signal intensity of the globus pallidus was calculated as the pallidal index, a percentile ratio of signal intensity in the globus pallidus to the subcortical frontal white-matter signal intensity in sagittal T1-weighted MRI planes (8). The pallidal index was also obtained from healthy volunteers as controls.
This study was approved by the investigation and ethics committee of the Kumamoto University Hospital. Informed consent after Institutional Review Board (IRB) approval was obtained from each subject.
Living-related reduced-size hepatic transplantation.
Cases 7 and 8 underwent hepatic transplantation with left lateral liver segments as grafts. Procurement of the graft was performed using an ultrasonic aspirator and bipolar electrocautery without blood vessel clamping and without graft manipulation. The partial liver graft was transplanted into the recipient, who underwent total hepatectomy with preservation of the inferior vena cava using a vascular side clamp. The hepatic vein of the liver graft was anastomosed to the inferior vena cava in the recipient. Microsurgery was used for hepatic artery reconstruction. Biliary reconstruction utilized a Roux-en-Y limb or interposed a previously existing jejunal conduit (9).
This regimen consisted of tacrolimus (FK506) with low-dose steroids. Methylprednisolone 10 mg/kg was given intravenously immediately after reperfusion. FK506 was begun orally at 0.075 mg/kg/day from the day before transplantation (10).
Living-related hepatic transplantation for biliary atresia.
Transplant case 1 (number 7): An 8-year-old girl was diagnosed with biliary atresia and underwent hepatic portoenterostomy 2 months after birth. She developed cirrhosis and pancytopenia due to hypersplenism. Liver enzymes included AST, 231 U/L (normal range, 1∼30 U/L); ALT, 209 U/L (1∼30 U/L); and gamma-glutamyl transpeptidase, 201 U/L (<45 IU/L). Complete blood count showed red blood cells, 342×104/mm3 (375∼507×104/mm3); hemoglobin, 8.5 mg/dL (11.4∼15.6 mg/dL); white blood cells, 1700/mm3 (3500∼8500/mm3); and platelets, 7.5×104/mm3 (13.6∼35.2×104/mm3). Blood manganese concentration was 6.8 μg/dL (<2.5 μg/dL). She underwent living-related hepatic transplantation without complications. The blood manganese concentration was reduced to 2.0 μg/dL at 7 months after transplantation.
Transplant case 2 (number 8): A 2-year-old girl was diagnosed with biliary atresia complicated by intracranial hemorrhage due to vitamin K deficiency and underwent hepatic portoenterostomy and craniotomy 2 months after birth. Elevated total and direct bilirubin concentrations were detected: 4.5 mg/dL (0.2∼1.2 mg/dL) and 2.5 mg/dL (0.0∼0.3 mg/dL), respectively. Liver enzymes were elevated: AST, 5707 U/L (1∼30 U/L); ALT, 2411 U/L (1∼30 U/L); and gamma-glutamyl transpeptidase, 292 U/L (<45 IU/L). She underwent living-related reduced-size hepatic transplantation without complications. The blood manganese concentration fell to 2.5 μg/dL at 2 months after transplantation.
Serum concentrations of total bilirubin were variable. Elevated blood manganese concentrations were detected in most patients (Table 1). One case (number 4) had depressive symptoms and dyskinesia. She recovered after oral administration of the dopamine precursor, l-DOPA. Cases 7 and 8 underwent living-related reduced-size hepatic transplantation.
T1-weighted MRI scans of the brain in patients with biliary atresia detected hyperintense signals in the globus pallidus. The pallidal index in atresia cases (112.4±2.43, mean ± SE) was significantly greater than in healthy volunteers (100.7±0.88) (P <0.05) (Table 2). Brain T1-weighted MRI demonstrated that the hyperintense signals in the globus pallidus disappeared in both cases within 3 months after transplantation, suggesting clearance of manganese from the brain (Figs. 1 and 2).
Correlations between serum manganese levels and the T1 ratio were assessed using linear regression grades. A significant positive correlation was observed between manganese levels and the T1 ratio (R=0.779, P =0.0006) (Fig. 3).
The present study demonstrates that hepatic transplantation normalizes blood manganese concentrations and reduces pallidal MR signals in patients with biliary atresia. It is known that patients with cirrhosis have significantly higher blood manganese concentrations than controls. In addition, semiquantitative scores of T1-weighted signal hyperintensity on brain MRI correlate with blood manganese concentrations in patients with cirrhosis (11–14). Thus, chronic liver failure is associated with high-intensity signal abnormalities in the basal ganglia on T1-weighted MRI of the brain (15, 16). These abnormalities are strikingly similar to those seen after manganese intoxication (11, 12). Maeda et al. (13) demonstrated that the globus pallidus and putamen show high intensities on T1-weighted MRI in patients with cirrhosis; these findings correspond to atrophy, necrosis, deciduation of nerve cells, and proliferation of glial cells and microglia in the globus pallidus. These findings are also similar to those of chronic manganese poisoning (17, 18). Deposition of manganese in the globus pallidus may cause a high signal on T1-weighted images and nerve cell death. Krieger et al. (8) demonstrated that manganese accumulates within the basal ganglia in patients with cirrhosis. This suggests that manganese has a role in the pathogenesis of chronic hepatic encephalopathy. Chelating agents may prove to be a therapeutic option to prevent or reverse such neuropsychiatric syndromes.
We had not previously examined noncholestatic patients undergoing liver transplantation with MRI before and after transplantation. In addition, we must consider the influence of immunosuppressants on T1 density in the brain. Tacrolimus induces cholestasis by primarily inhibiting biliary excretion of glutathione (19). Furthermore, inhibition of ATP-dependent bile salt transport by cyclosporine may induce cholestasis (20). These data suggest that immunosuppressants may induce cholestasis, and therefore, immunosuppressants are not likely to reduce T1 density in the brains of transplant recipients.
The underlying mechanism(s) of manganese neurotoxicity in patients with end-stage cirrhosis are unresolved. Blood manganese concentrations were elevated in cirrhotic patients and in patients with portacaval anastomoses or transjugular intrahepatic portosystemic shunts. Pallidal signal hyperintensity was also observed in such patients (21). Assessment of extrapyramidal symptoms using the Columbia rating scale revealed a significant incidence of tremor, rigidity, and akinesia in cirrhotic patients (22). Spahr et al. (14) reported that there was no significant correlation between blood manganese levels and extrapyramidal symptoms, although the severity of akinesia was significantly greater in Child-Pugh C patients. Extrapyramidal symptoms may result from a toxic effect of manganese on basal ganglia dopaminergic function, but further studies are required.
Manganese toxicity recently has been recognized in children with hepatic dysfunction on total parenteral nutrition (23). The clinical spectrum includes cholestasis, cholelithiasis, hepatic fibrosis with progression to biliary cirrhosis, and development of portal hypertension and liver failure in a significant number of children who are totally parenterally fed. Fell et al. (24) reported on patients receiving long-term parenteral nutrition and found that cholestatic disease and nervous system disorders were associated with high blood concentrations of manganese. Additionally, MRI with T1-weighted images showed bilateral symmetrically increased signal intensity in the globus pallidus and subthalamic nuclei (24).
Manganese neurotoxicity has been implicated in patients with end-stage cirrhosis and with TPN-induced cholestatic liver disease. However, no apparent neurologic symptoms have been observed in patients with biliary atresia. A relationship between manganese deposition within the brain and neurologic symptomatology in patients with biliary atresia has not been established. Yet deficiency of dopamine within the brain is generally accepted as an expression of manganese toxicity in experimental animals (25). In humans, manganese intoxication produces an early psychotic disorder that is followed by a Parkinson-like syndrome. Autoreceptor presynaptic control of dopamine released from the striatum is lost at the early stages of manganese poisoning (26). The toxicity of manganese may also be related to its ability to accelerate the oxidation of catecholamines (27, 28). Autoxidation of dopamine is catalyzed by manganese through the formation of a highly reactive complex. Manganese participates in a redox cycle, which involves intramolecular electron transfer between manganese and the dopamine ligand (29). This may be a possible insight in manganese-mediated dopaminergic neurotoxicity. It is well known that the developing nervous system is able to compensate for neurochemical changes caused by manganese exposure (30). In addition, an age-related impairment of the neuronal antioxidant system, including dopamine, may play an enabling role in manganese neurotoxicity (16). Only one of our patients had neuropsychiatric symptoms and dyskinesia, but she recovered after oral administration of l-DOPA. This may explain why children with biliary atresia are less susceptible to manganese intoxication than adults with cholestasis due to chronic liver damage.
In conclusion, manganese accumulates within the brains of patients with biliary atresia, but there are few neurologic symptoms. However, manganese deposition within the brain resolves after living-related hepatic transplantation.
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