Autoimmune Thyroiditis Associated with Autoimmune Hepatitis

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Autoimmune hepatitis (AIH) is a disease of unknown etiology in which progressive destruction of the hepatic parenchyma occurs leading to cirrhosis and liver fail ...
THYROID Volume 15, Number 10, 2005 © Mary Ann Liebert, Inc.

Case History Autoimmune Thyroiditis Associated with Autoimmune Hepatitis Valerio Nobili,1 Christos Liaskos,2 Giovannelli Luigi,3 Roberto Guidi,3 Paola Francalanci,4 and Matilde Marcellini1

Introduction

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(AITD) are organ-specific autoimmune disorders where both genetic and environmental factors are thought to play a role. The subjects affected by AITD are prone to develop other autoimmune disorders (1–4). Autoimmune hepatitis (AIH) is a disease of unknown etiology in which progressive destruction of the hepatic parenchyma occurs leading to cirrhosis and liver failure (5). We report an unusual case of severe AIH in a girl with autoimmune thyroiditis. Following this case we suggest that, when abnormalities in aminotransferase levels persist after correcting the thyroid dysfunction, a liver biopsy is mandatory in order to differentiate AIH from liver involvement secondary to thyroid disease and to promptly start an appropriate therapy. UTOIMMUNE THYROID DISEASES

Case Report A 14-year-old girl was referred to our Paediatric Endocrinology Clinic in July 2003 with symptoms of irritability and increased sweating for assessment of possible thyroiditis. At examination her thyroid gland was diffusely enlarged, firm, and nontender. The level of thyrotropin (TSH) was slightly elevated (6.80 IU/mL; normal values, 0.40–5.50 IU/mL) and there was a decrease in the levels of both free thyroxine (FT4; 0.50 ng/dL; normal values, 0.70–1.80 ng/mL) and free triiodothyronine (FT3; 1.70 pg/mL; 2.20–5.10 pg/mL). The patient had serum antibodies to thyroid microsomal antigens, whereas the antithyroglobulin test was negative. Thyroid ultrasound revealed an enlarged, hypoechoic gland with a fine micronodular pattern. The girl was discharged after 4 days in the hospital with a diagnosis of Hashimoto’s thyroiditis and treatment with sodium levothyroxine was started. At diagnosis as well as during 2 months of follow-up, persistently increased levels of aminotransferases (alanine aminotransferase [ALT] 86 IU/L; normal values, 5–40 UI/L; aspartate aminotransferase [AST] 113 IU/L; normal values, 5–40 UI/L) were noted.

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Three months after discharge, clinical improvement continued but aminotransferases (ALT, 98 IU/L; AST, 101 IU/L) were still increased, whereas total bilirubin and serum alkaline phosphatase levels were normal, as well as immunoglobulins (Ig) G, A, and M levels. Hepatitis A, B, C, D, E, cytomegalovirus, and Epstein-Barr virus infections were ruled out by appropriate tests, which included serum HBV-DNA and HCV-RNA. Antinuclear antibodies (ANA), antiliver/kidney microsomal antibodies (LKM-1), antismooth muscle antibodies (ASMA), and antimitochondrial antibodies (AMA) were all negative by indirect immunofluorescence. The girl’s HLA haplotype was A*1, B*8, DRB1*04, DQB1*02, and the family history was unremarkable for autoimmune diseases. Ultrasound-guided percutaneous liver biopsy is the most commonly used procedure to obtain tissue for histopathologic assessment of the nature and severity of a liver disease. When performed by a experienced operator, it is a safe, relatively simple and effective technique and it is considered the gold standard to diagnose AIH (6). The demonstration of pathologic liver histology (interface hepatitis, predominant lymphoplasmacytic infiltrate) or the exclusion of important diagnostic considerations are sometimes as important in the management of the patient and for the family’s peace of mind as any positive finding. A percutaneous liver biopsy was performed and revealed portal and periportal hepatitis with plasma cells at the interface. A dilated portal vein and multinucleated giant hepatocytes were also seen (Fig. 1). A diagnosis of AIH was made (7). Treatment with prednisone (2 mg/kg daily) and azathioprine (2 mg/kg daily) was started and AST and ALT concentrations returned to normal in 6 weeks. Autoantibody Detection Indirect immunofluorescence The serum sample was tested for conventional antibodies by indirect immunofluorescence (IIFL) on 5-m frozen sections of a composite substrate containing rat liver, kidney,

Liver Disease, Research Institute, Bambino Gesu’ Children’s Hospital, Rome, Italy. Medicine, University of Thessay, Larisa, Greece. Laboratory, Research Institute, Bambino Gesu’ Children’s Hospital, Rome, Italy. Pathology, Research Institute, Bambino Gesu’ Children’s Hospital, Rome, Italy.

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FIG. 1. Autoimmune hepatitis: Portal and periportal hepatitis with plasma cells at the interface. A dilated portal vein and multinucleated giant hepatocytes were also seen.

and stomach using as revealing reagent an antitotal human IgG-fluorescein isothiocyanate conjugate (Dako Ltd., High Wycombe, Bucks, UK) at an initial dilution of 1/10 as proposed by the recent consensus statement from the committee for autoimmune serology of the International Autoimmune Hepatitis Group (8). In order to investigate also antinuclear reactivity, the serum sample was also evaluated using commercially available human epithelial cells, HEp2010 (Euroimmun, Lübeck, Germany) at a 1/40 dilution. Each slide was examined by an independent observer using an Olympus BX40 (Olympus Optical, London, UK) fluorescence microscope. Immunobloting using human liver antigen and HEp-2 cell extract Autoantibody binding was evaluated further by immunobloting using strips precoated with either electrophoretically separated microsomal, cytosolic and mitochondrial fractions of human liver antigen or HEp-2 cell extract, according to the manufacturer’s instructions (Euroimmun). Strips were incubated for 1 hour with individual serum samples at a dilution of 1/100 and after three 5-minutes washes, they were incubated for 1 hour with horseradish peroxidase (HRP)-conjugated anti-human IgG (1/2000) (Dako). After washing, the blots were developed with diaminobenzidine at 0.25 mg/mL in phosphate-buffered saline (PBS) with 0.1% H2O2 as substrate (Euroimmun). Two reference strips from the same lots with immunofixed bands of known microsomal, mitochondrial, and cytosolic autoantigens or known antinuclear reactivities were used for the interpretation of the results of individual strips, according to the manufacturer instructions. Commercially available enzyme-linked immunosorbent assays (ELISAs; (Euroimmun) were used for the detection of anticytochrome P-450IID6 (CYP2D6), the target of liver kid-

ney microsomal type 1 (LKM1), antibodies; anti-formiminotransferase cyclodeaminase (FTCD), the molecular target of liver cytosol-1 (LC1), antibodies; and anti-UGA suppressor tRNA-associated antigenic protein (tRNP(Ser)Sec, the molecular target of soluble liver antigen (SLA), according to the manufacturer’s instructions. Results The serum sample immunofixed seven bands, with molecular weights ranging from 37–74 kd, on electrophoretically separated human liver homogenate. Two of these bands have the molecular weight of cytokeratin 8 and 18, described as autoantigens in patients with autoimmune hepatitis (9). Another band has a molecular weight of 50 kd. This is the molecular weight of cytochrome P450IID6, the target of LKM-1; however, when the serum was tested for antiCYP2D6 reactivity against the recombinant enzyme, it was found to be negative confirming the negativity for LKM-1 by immunofluorescence. The identity of this 50-kd autoantigen therefore remains to be defined. The remaining four bands do not correspond to defined autoantigenic targets, even though an especially evident band was observed at 40 kd. As alluded to above, LKM-1 was negative by immunofluorescence as were all conventional autoantibodies detected by this technique, including ANA and SMA. ANA reactivities were also negative by immunoblot using a nuclear preparation from HEp-2 cell as substrate. LC1 and SLA reactivities tested by ELISA were negative. Discussion Mild increases in aminotransferases are common in children with autoimmune thyroiditis and often regarded as of little clinical importance (10,11). Occasionally, however, such

AUTOIMMUNE THYROIDITIS ASSOCIATED WITH AUTOIMMUNE HEPATITIS increases may indicate an underlying autoimmune hepatitis, as we report in the present paper. Several authors have described altered liver function, such as elevations of aminotransferase and alkaline phosphatase levels, in association with AITD. Altered LFTs return to normal once the primary thyroid pathology is adequately treated, without residual liver damage (12,13). The persistence of elevated aminotransferase levels needs further investigation. In our own patient, the aminotransferase elevation preceded the diagnosis of thyroiditis and continued during the treatment with antithyroid hormones. This led us to investigate autoimmune serology that demonstrated the presence of several autoantibodies directed to antigens contained in a preparation of human liver homogenate. The targets of these autoantibodies were defined by their molecular weight, which ranged from 37–74 kd, but not corresponding to the conventional autoantibodies of AIH. Seronegativity for the conventional autoantibodies at presentation does not, however, exclude the diagnosis of AIH (14). A biopsy was then performed revealing lesions compatible with AIH. The classical treatment of AIH, with corticosteroid and azathioprine, was promptly instituted leading to normalization of the aminotransferase levels within 2 months. These levels remain normal 8 months after starting the immunosuppressive treatment that currently consists of AZT and steroid. Of note, the immunogenetic make up of our patients A*1, B*8, DRB1*04, DQB1*02 is known to predispose to AIH. Taken together our data suggest that in patients with AITD in whom aminotransferase levels are elevated and persist elevated the diagnosis of autoimmune hepatitis should be considered. Appropriate investigations should include the study of the autoimmune serology and of the liver histology: if the diagnosis of AIH is made the prompt institution of the correct treatment will control the disease, as our case clearly demonstrates.

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Address reprint requests to: Valerio Nobili Department of Liver Disease Bambino Gesu’ Children’s Hospital S. Onofrio 4 Square 000165 Rome Italy E-mail: [email protected]