AUTREV-01485; No of Pages 6 Autoimmunity Reviews xxx (2014) xxx–xxx

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Autoimmunity Reviews journal homepage: www.elsevier.com/locate/autrev

Review

Diagnostic criteria of autoimmune hepatitis Rodrigo Liberal a,b, Charlotte R. Grant a, Maria Serena Longhi a, Giorgina Mieli-Vergani a, Diego Vergani a,⁎ a b

Paediatric Liver, GI & Nutrition Centre and Institute of Liver Studies, King's College London School of Medicine at King's College Hospital, London, UK Faculty of Medicine, University of Porto, Porto, Portugal

a r t i c l e

i n f o

Article history: Accepted 13 November 2013 Available online xxxx

a b s t r a c t Autoimmune hepatitis (AIH) is a chronic immune-mediated liver disorder characterised by female preponderance, elevated transaminase and immunoglobulin G levels, seropositivity for autoantibodies and interface hepatitis. Presentation is highly variable, therefore AIH should be considered during the diagnostic workup of any increase in liver enzyme levels. A set of inclusion and exclusion criteria for the diagnosis of AIH have been established by the International Autoimmune Hepatitis Group (IAIHG). There are two main types of AIH: type 1, positive for anti-nuclear (ANA) and/or anti-smooth muscle antibodies (SMAs) and type 2, defined by the presence of anti-liver kidney microsomal antibody type 1 (LKM-1) and/or anti-liver cytosol type 1 (LC-1) autoantibodies. The central role of autoantibodies in the diagnosis of AIH has led the IAIHG to produce a consensus statement detailing appropriate and effective methods for their detection. Autoantibodies should be tested by indirect immunofluorescence at an initial dilution of 1/40 in adults and 1/10 in children on a freshly prepared rodent substrate that includes kidney, liver and stomach sections to allow for the simultaneous detection of all reactivities relevant to AIH. Anti-LKM-1 is often confused with anti-mitochondrial antibody (AMA) if rodent kidney is used as the sole immunofluorescence substrate. The identification of the molecular targets of anti-LKM-1 and AMA has led to the establishment of immuno-assays based on the use of the recombinant or purified autoantigens. Perinuclear anti-nuclear neutrophil antibody (p-ANNA) is an additional marker of AIH-1; anti soluble liver antigen (SLA) antibodies are specific for autoimmune liver disease, can be present in AIH-1 and AIH-2 and are associated with a more severe clinical course. Anti-SLA are detectable by ELISA or radio-immuno-assays, but not by immunofluorescence. AIH is exquisitely responsive to immunosuppressive treatment, which should be instituted promptly to prevent rapid deterioration and promote remission and long-term survival. © 2014 Elsevier B.V. All rights reserved.

Contents 1. 2. 3. 4.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . Epidemiology . . . . . . . . . . . . . . . . . . . . . . . . . . . Clinical presentation and natural history . . . . . . . . . . . . . . . Diagnosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.1. Scoring systems . . . . . . . . . . . . . . . . . . . . . . 4.2. Laboratory abnormalities . . . . . . . . . . . . . . . . . . 4.3. Diagnostic autoantibodies . . . . . . . . . . . . . . . . . . 4.3.1. Anti-nuclear antibodies . . . . . . . . . . . . . . . 4.3.2. Anti-smooth muscle antibodies . . . . . . . . . . . 4.3.3. Anti-liver-kidney-microsomal type 1 antibodies . . . . 4.3.4. Variant liver microsomal antibodies . . . . . . . . . 4.3.5. Anti-liver cytosol type 1 antibodies . . . . . . . . . 4.3.6. Anti-soluble liver antigen/liver-pancreas antibodies . . 4.3.7. Anti-neutrophil cytoplasmic antibodies . . . . . . . . 4.3.8. Anti-asialoglycoprotein receptor antibodies (anti-ASGPR) 4.4. Histology . . . . . . . . . . . . . . . . . . . . . . . . . 5. Treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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⁎ Corresponding author at: Institute of Liver Studies, King's College Hospital, Denmark Hill, London SE5 9RS, UK. Tel.: +44 203 2993357; fax: +44 203 2994224. E-mail address: [email protected] (D. Vergani). 1568-9972/$ – see front matter © 2014 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.autrev.2013.11.009

Please cite this article as: Liberal R, et al, Diagnostic criteria of autoimmune hepatitis, Autoimmun Rev (2014), http://dx.doi.org/10.1016/ j.autrev.2013.11.009

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R. Liberal et al. / Autoimmunity Reviews xxx (2014) xxx–xxx

1. Introduction Autoimmune hepatitis (AIH), a progressive inflammatory hepatopathy and important cause of end-stage liver disease, was first described in a group of young women in 1950 by the Swedish professor Jan Waldenström. The patients were afflicted by a severe form of un-resolving hepatitis characterised by markedly elevated serum immunoglobulin levels [1]. The observation that a proportion of patients in similar cohorts were positive for lupus erythematosus cells and anti-nuclear autoantibodies (ANA), led to the hypothesis that a loss of immune tolerance was at the basis of this condition [2]. The most typical features of AIH are female preponderance, hypergammaglobulinaemia, seropositivity for circulating autoantibodies and a picture of interface hepatitis on histology [3]. AIH responds to immunosuppressive treatment in the majority of cases. Treatment should be instituted promptly upon diagnosis. If left untreated, AIH usually progresses to liver failure requiring transplantation. Two types of AIH are distinguished according to serological profile: type 1 AIH (AIH-1) patients are positive for ANA and/or anti-smooth muscle antibody (SMA), and type 2 AIH (AIH-2) patients are defined by the positivity for anti-liver kidney microsomal type 1 antibody (anti-LKM-1) and/or for anti-liver cytosol type 1 antibody (anti-LC-1) [4]. 2. Epidemiology AIH occurs globally, affecting children and adults of all ages [5,6] and both sexes, although it is more commonly found in females [7]. Estimates of the prevalence of AIH were mired by a lack of standardization before the introduction of the International Autoimmune Hepatitis Group (IAIHG) Scoring System in the early 1990s [8,9]. Moreover, early studies did not exclude patients with hepatitis C virus infection. The first study to utilize the IAIHG Scoring System reported a prevalence of definite AIH of 34.5 cases per 100,000 in Alaskan natives [10]. In a more recent epidemiological study using the standardized inclusion criteria, an annual incidence of 2.0 cases per 100.000 and a point prevalence of 24.5 cases per 100,000 were observed in New Zealand [11]. Importantly, the prevalence of AIH-2 is largely unknown because diagnosis is still often overlooked. At the King's College Hospital tertiary paediatric hepatology referral centre, there has been a seven-fold increase in the incidence of both AIH-1 and AIH-2 over the last decade. AIH represents approximately 10% of some 400 new referrals per year, with two thirds of cases diagnosed with AIH-1 and one-third with AIH-2, a disease that typically affects children. 3. Clinical presentation and natural history The mode of presentation of AIH is diverse and highly variable [12], although the majority of adult patients manifest with an insidious onset characterised by progressive fatigue, relapsing jaundice, amenorrhea, weight loss and occasionally arthralgia [13]. Complications of portal hypertension, such as gastrointestinal bleeding or hypersplenism can sometimes be apparent [14]. Approximately 25% of patients are completely asymptomatic and are diagnosed after incidental discovery of abnormal liver function tests. Finally, 30 to 40% of patients, particularly children and young adults, present with symptoms and signs mimicking those of acute hepatitis due to other causes. Although rarely, AIH patients can also present with fulminant hepatic failure [15]. Irrespective of the mode of presentation, histological evidence of cirrhosis is found in at least 30% of patients, indicating that subclinical disease has been present for some time. Indeed, advanced fibrosis or cirrhosis can often be found in patients presenting acutely [3]. AIH can occasionally develop during pregnancy [16]. Appearance of AIH post-partum and exacerbations of existing disease in patients whose condition improved during pregnancy have also been described [17].

Approximately 40% of AIH patients have a family history of autoimmune disease and at least 20% have concomitant autoimmune diseases or will develop them during follow-up [14]. In line with other chronic liver diseases, AIH can ultimately progress to cirrhosis and occasionally hepatocellular carcinoma (HCC) despite the use of immunosuppressive therapy. In one study, of 243 AIH patients followed up for 16 years, 15 cases of HCC with underlying cirrhosis, were reported [18]. In another study, 6/322 AIH patients, again with cirrhosis, developed HCC within 10 years of follow-up [19]. Surveillance for HCC is therefore warranted [20]. 4. Diagnosis 4.1. Scoring systems The diagnosis of AIH is based on the presence of elevated serum transaminase and IgG levels, autoantibody seropositivity and histological evidence of interface hepatitis. This tenet has been factored into the IAIHG diagnostic criteria for AIH [8,9], which were originally developed for use in the research setting. This scoring system incorporates a number of positive and negative scores, enabling the researcher to grade clinical, laboratory and histological features of AIH. The score has also proved useful in the clinical context when assessing patients with few or atypical features of the disease. The distinction between a definite and probable diagnosis of AIH predominantly relates to the extent of the increase in serum gamma-globulin/IgG or autoantibody titre, as well as exposure to alcohol, hepatotoxic medication or infection. Laboratory and histological features associated with cholestasis carry a negative score. In the rare instances where conventional autoantibodies are not detected, the presence of anti-asialoglycoprotein receptor (anti-ASGPR), anti soluble liver antigen/liver pancreas (anti-SLA/LP) or atypical perinuclear antineutrophil cytoplasmic antibodies (atypical pANCA, currently better referred to as pANNA) weigh towards a probable diagnosis of AIH. The scoring system also incorporates response to corticosteroids, with a definite diagnosis before steroid treatment requiring a score higher than 15, and a definite diagnosis after treatment institution requiring a score greater than 17 (Table 1) [9]. It is important to note that because healthy children are very rarely autoantibody positive, in the paediatric setting, lower autoantibody titres contribute to the diagnoses of AIH. These titres are 1:20 for ANA and SMA and 1:10 for anti-LKM-1. A simplified scoring system, for use in clinical practice, has recently been proposed by the IAIHG (Table 2) [21]. This system is based on only four criteria: positivity for autoantibodies, elevated IgG levels, histological evidence of interface hepatitis and the exclusion of viral hepatitis. Neither scoring system is immediately applicable to the diagnosis of the juvenile form of AIH. 4.2. Laboratory abnormalities The most frequently observed laboratory abnormalities associated with AIH include elevated levels of aspartate (AST) and alanine (ALT) aminotransferases. Less striking are increases in serum bilirubin or alkaline phosphatase (AP) levels. Cholestasis, evident in a minority of patients, warrants consideration of extra-hepatic obstruction and cholestatic forms of viral hepatitis, drug-induced disease, primary biliary cirrhosis (PBC), primary sclerosing cholangitis (PSC), and overlap syndromes [12]. AIH is also often associated with a generalised elevation of serum globulins, particularly gamma globulins, mainly due to an increase in IgG. The serum autoantibodies typically present include ANA, SMA, anti-LKM-1, and anti-LC-1. Anti-mitochondrial antibodies (AMAs), typically associated with PBC, are occasionally found in AIH. The isolated occurrence of autoantibody positivity does not equate to a diagnosis

Please cite this article as: Liberal R, et al, Diagnostic criteria of autoimmune hepatitis, Autoimmun Rev (2014), http://dx.doi.org/10.1016/ j.autrev.2013.11.009

R. Liberal et al. / Autoimmunity Reviews xxx (2014) xxx–xxx Table 1 International autoimmune hepatitis group revised diagnostic scoring system. Adapted from Alvarez F, Berg PA et al. J Hepatol 1999; 31: 929:938.

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Table 2 Simplified criteria for the diagnosis of autoimmune hepatitis. Adapted from Hennes EM, Zeniya M et al. Hepatology 2008; 48: 169–176.

Parameter

Feature

Score

Variable

Cut-off

Points

Sex ALP: AST (or ALT) ratio

Female N3 1.5–3 b1.5 N2.0

+2 −2 0 +2 +3

ANA or SMA ANA or SMA or anti-LKM-1 or SLA IgG

1 2a

1.5–2.0 1.0–1.5 b1.0 N1:80 1:80 1:40 b1:40 Positive Positive Negative Yes No b25 g/day N60 g/day Interface hepatitis Plasma cells Rosettes None of the above Biliary changes a Atypical changes b Thyroiditis, colitis, other DR3 or DR4 Anti-SLA/LP, actin, ASGPR, pANNA Remission Relapse

+2 +1 0 +3 +2 +1 0 −4 −3 +3 −4 +2 +2 −2 +3 +1 +1 −5 −3 −3 +2 +1 +2

Liver histology

≥1:40 ≥1:80 ≥1:40 Positive NUpper limit of normal N1.10 times upper limit of normal Compatible with AIH Typical of AIH Yes

Serum globulins or IgG (times above normal)

ANA, SMA or anti-LKM-1 titres

AMA Viral markers of active infection Hepatotoxic drug history Average alcohol Histological features

Immune diseases HLA Seropositivity for other autoantibodies Response to therapy

+2 +3

Pre-treatment score N15: definite AIH; 10–15: probable AIH; Post-treatment score N17: definite AIH; 12–17: probable AIH. ALP, alkaline phosphatase; AST, aspartate aminotransferase; ALT, alanine aminotransferase; IgG, immunoglobulin G; ANA, anti-nuclear antibody; SMA, anti-smooth muscle antibody; anti- LKM-1, anti-liver kidney microsomal type 1 antibodies; AMA, anti-mitochondrial antibodies; SLA/LP, soluble liver antigen/liver pancreas; ASGPR, asialoglycoprotein receptor; p-ANNA, peripheral anti-nuclear neutrophil antibody; and HLA, human leukocyte antigen. a Including granulomatous cholangitis, concentric periductal fibrosis, ductopenia, marginal bile duct proliferation and cholangiolitis. b Any other prominent feature suggesting a different aetiology.

of AIH because autoantibodies have also been reported in other liver diseases. 4.3. Diagnostic autoantibodies Autoantibody detection should proceed as soon as a diagnosis of AIH is suspected [4]. Not only does positivity weigh towards AIH diagnosis, but autoantibody profiles enable the distinction between AIH type 1 and AIH type 2. ANA and/or SMA, characterising AIH-1, and antiLKM-1 and/or anti-LC-1, defining AIH-2, are rarely detected in combination (Fig. 1) [4], and in those cases in which they are present simultaneously, the clinical course is similar to that of AIH-2. Antibodies to LC-1, SLA/LP, and to neutrophil cytoplasmic antigens may assist in diagnosing patients negative for the defining AIH autoantibodies; for this reason, they have been incorporated in the original and revised IAIHG diagnostic scoring systems [8,9]. Recognition and interpretation of immunofluorescence (IFL) patterns can be challenging, and reporting errors are not infrequent due to the inherent operator-dependency of the technique and the relative rarity of AIH. Clinical interpretation of laboratory results can also be problematic, partly owing to insufficient test standardization but also because some clinicians can be unfamiliar with the disease spectrum of AIH. In 2004, the IAIHG Committee for Autoimmune Serology drew up a consensus statement containing guidelines for appropriate and

Absence of viral hepatitis

1 2 1 2 2

Score ≥ 6: probable AIH; ≥7: definite AIH. ANA, anti-nuclear antibody; SMA, anti-smooth muscle antibody; anti-LKM-1, anti-liver kidney microsomal antibody type 1; SLA, soluble liver antigen; IgG, immunoglobulin G; and AIH, autoimmune hepatitis. a Addition of points achieved for all autoantibodies cannot exceed a maximum of 2 points.

effective autoantibody testing in AIH [4]. The document contained strong recommendations that first line screening should consist of indirect IFL on fresh, multi-organ rodent sections (usually rat liver, kidney and stomach) to enable the concomitant detection of a range of autoantibodies relevant to liver disease: SMA, ANA, anti-LKM-1, anti-LC-1 and AMA (Table 3). The committee also detailed the correct methodology for substrate preparation, dilution and application of serum samples, dilution of fluorochrome-labelled revealing agents, appropriate selection of controls, as well as accurate identification of diagnostically relevant staining patterns [4]. Of note, tissue sections should be air-dried and used without further fixation. Commercially available sections vary considerably, due to the use of fixatives, which inevitably increase background staining and interfere with autoantibody detection [4]. Diagnostic screening requires careful planning and orientation of kidney sections to ensure that they contain both proximal and distal tubules. This is because, although anti-LKM-1 and AMA both stain the renal tubules, AMA has greater affinity for the mitochondria-reach distal tubules, whereas anti-LKM-1 stains the third portion of the larger proximal tubules. The use of multi-tissue substrate also facilitates distinction between these autoantibodies, because AMA, but not antiLKM-1, stains the gastric parietal cells.

4.3.1. Anti-nuclear antibodies ANA produce a clear and easily detectable nuclear IFL pattern on kidney, stomach and liver sections. In AIH, a homogenous staining pattern is most common, particularly in the liver, with coarsely or finely speckled patterns visualised less frequently [4]. In AIH, clinically relevant ANA titres are considered 1/40 in adults and 1/20 in children, in whom titres correlate with disease activity. In order for clearer definition of the nuclear pattern, human epithelial type 2 (HEp2) cells can be utilised owing to their prominent nuclei. These cells are, however, less than ideal for screening purposes, due to a high positivity rate in healthy subjects. It is important to note that ANA can also be identified in up to 52% of patients with PBC. In contrast to AIH, however, PBC-specific ANA is highly diagnostic for this condition, with multiple nuclear dot or rim-like membranous patterns. This pattern is recognised by IFL when HEp-2 or HeLa cells are used as substrate. Moreover, ANA are present in other autoimmune disorders, such as SLE, Sjögren syndrome and systemic sclerosis, as well as non-autoimmune conditions, like viral hepatitis, drug-induced hepatitis, and alcoholic and non-alcoholic fatty liver diseases [22]. The target antigens of ANA in AIH are heterogeneous and incompletely defined, although ANA have been shown to react with singleand double-stranded deoxyribonucleic acid (DNA), small nuclear ribonucleoproteins (sn-RNPs), centromeres, histones, chromatin and

Please cite this article as: Liberal R, et al, Diagnostic criteria of autoimmune hepatitis, Autoimmun Rev (2014), http://dx.doi.org/10.1016/ j.autrev.2013.11.009

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R. Liberal et al. / Autoimmunity Reviews xxx (2014) xxx–xxx

Fig. 1. Indirect immunofluorescence appearance on rodent tissue substrates of the autoantibodies diagnostic of autoimmune hepatitis (AIH). Left AIH type 1 (AIH-1): top anti-nuclear antibody (ANA), bottom anti-smooth muscle antibody (SMA) staining a vessel (V pattern) and a glomerulus (G pattern). Right AIH type 2 (AIH-2): top anti-liver kidney microsomal type 1 (LKM-1) antibody, bottom anti-liver cytosol type 1 (LC-1) antibody.

cyclin A. A better definition of the ANA target antigens will likely follow the development of new techniques using recombinant nuclear antigens and immuno-assays. The mechanisms leading to the production of ANA in AIH are not understood, although the release of nuclear components resulting from hepatocyte injury and/or the loss of B cell tolerance to several nuclear components are possible explanations [22].

4.3.2. Anti-smooth muscle antibodies IFL patterns characteristic of SMA positivity can be visualised on kidney, stomach and liver sections. SMAs target the artery walls and, in the stomach substrate they also bind the muscularis mucosa and the lamina propria. In the kidney, SMA from AIH typically stains the smooth muscle of the vessels, glomeruli and tubules (VGT pattern). The VG and VGT IFL patterns are considerably more specific for AIH than the solo V pattern, which has also been observed in liver disease of other aetiologies, infectious diseases and rheumatic disorders [4]. The titres of ANA in AIH usually

equate to or exceed 1/80, although very young patients may have titres as low as 1/20. The first targets of AIH-specific SMA to be recognised were constituents of actin. Later, SMAs were also shown to be directed against other components of the cytoskeleton such as tubulin, vimentin, desmin, and skeletin [23]. The AIH-1-specific target of SMA responsible for the VGT pattern remains elusive, however, several studies point to actin in its filamentous form. Despite the VGT pattern being highly specific for AIH-1, it is absent in approximately 20% of SMA-positive patients. Moreover, when molecular assays using purified F-actin are employed, some AIH VGT positive cases are negative, while anti-F-actin positivity is reported in diseases distinct from AIH-1 [4,23]. 4.3.3. Anti-liver-kidney-microsomal type 1 antibodies Anti-LKM-1, the hallmark of AIH-2, stains the hepatocellular cytoplasm and the P3 portion of the renal tubules. As described previously, the IFL patterns of anti-LKM-1 and AMA can become confused because both autoantibodies stain the liver as well as the kidney. However, AMA stains the liver more faintly than anti-LKM-1, and marks the renal

Table 3 Autoantibodies and their targets in autoimmune liver diseases. Autoantibody

Target antigen(s)

Liver disease

Value in AIH

Conventional method of detection

Molecular based assays

ANA

AIH PBC PSC Drug-induced Chronic hepatitis C Chronic hepatitis B NAFLD Same as ANA

Diagnostic of AIH-1

IIF

ELISA, IB, LIA

Diagnostic of AIH-1

IIF

ELISA

Anti-LKM1

Chromatin Histones Centromeres Cyclin A Ribonuclueoproteins Double stranded DNA Single stranded DNA Microfilaments (filamentous actin) Intermediate filaments (vimentin, desmin) Cytochrome P4502D6

Diagnostic of AIH-2

IIF

ELISA, IB, LIA, RIA

Anti-LC1

Forminino-transferase cyclodeaminase

ELISA, LIA, RIA

tRNP(Ser)Sec

Inhibition ELISA

ELISA, IB, RIA

pANNA

Nuclear lamina proteins

Diagnostic of AIH-2 Prognosis of severe disease Diagnostic of AIH Prognostic of severe disease, relapse and treatment dependence Point towards diagnosis of AIH

IIF, DID, CIE

SLA/LP

AIH-2 Chronic hepatitis C AIH-2 Chronic hepatitis C AIH Chronic hepatitis C

IIF

N/A

AMA

E2 subunits of 2-oxoacid dehydrogenase complexes, particularly PDC-E2

Against diagnosis of AIH

IIF

ELISA, IB, RIA

SMA

AIH PSC/ASC PBC

ANA, anti-nuclear antibodies; SMA, anti-smooth muscle antibodies; anti-LKM1, anti-liver kidney microsomal antibody type 1; anti-LC1, anti-liver cytosol antibody type 1; SLA/LP, soluble liver antigen/liver pancreas; pANNA, peripheral anti-nuclear neutrophil antibodies; AMA, anti-mitochondrial antibodies; AIH, autoimmune hepatitis; PBC, primary biliary cirrhosis; PSC, primary sclerosing cholangitis; NAFLD, non-alcoholic fatty liver disease; IIF, indirect immunofluorescence; DID, double-dimension immune-diffusion; CIE, counter-immune-electrophoresis; ELISA, enzyme-linked immunosorbent assay; IB, immunoblot; LIA, line-immuno-assay; RIA, radio-immune-precipitation assay; and N/A, not applicable.

Please cite this article as: Liberal R, et al, Diagnostic criteria of autoimmune hepatitis, Autoimmun Rev (2014), http://dx.doi.org/10.1016/ j.autrev.2013.11.009

R. Liberal et al. / Autoimmunity Reviews xxx (2014) xxx–xxx

tubules more diffusely, while accentuating the distal tubules. Additionally, AMA stain gastric parietal cells while anti-LKM-1 does not [4,23]. Clinically relevant anti-LKM-1 titres are considered to be 1/40 in adults and 1/10 in patients under 18 years of age; the titre of this autoantibody is associated with disease activity [4]. Interestingly, anti-LKM-1 is also detected in some 5–10% of patients with chronic hepatitis C virus (HCV) infection. Cytochrome P450-2D6 (CYP2D6) is the molecular target of anti-LKM-1. Since the molecular targets of AMA – enzymes of the 2-oxo-acid dehydrogenase complexes – have also been identified, immune-assays based on the use of recombinant or purified antiLKM-1 and AMA autoantigens have been developed. Commercially available enzyme-linked immunosorbent assays (ELISAs) accurately detect anti-LKM-1, at least in the context of AIH-2, and detect AMA reasonably accurately. These assays can therefore be utilised when doubt about IFL patterns arises [4]. 4.3.4. Variant liver microsomal antibodies Variant liver microsomal antibodies are most commonly directed against non-2D6 isoforms of cytochrome-P450. Antiliver microsol antibodies only stain the liver cytoplasm, react with liver specific cytochrome P4501A2 and occur in dihydralazine induced hepatitis and in hepatitis associated with autoimmune polyendocrinopathycandidiasis-ectodermal dystrophy (APECED) [24]. Antibodies specific for P4502A6, which are found in APECED and occasionally hepatitis C patients, have an anti-LKM-1 like pattern on immunofluorescence. The term anti-LKM-2 was coined to describe anti-LKM-1-like microsomal antibodies directed against cytochrome P4502C9 that are produced during hepatitis induced by the anti-hypertensive tienilic acid, which is no longer marketed [25]. Anti-LKM-3, targeting members of the family 1 UDP glucuronosyltransferases (UGT), also give an immunofluorescent pattern similar to anti-LKM-1, but this autoantibody is found mainly in hepatitis D (delta) [26] and in some patients with AIH [27]. 4.3.5. Anti-liver cytosol type 1 antibodies Anti-LC-1 was originally described alone or in combination with anti-LKM-1 to define AIH-2. Although anti-LC-1 has also been detected in patients positive for serological markers associated with AIH-1, and in patients with chronic HCV infection, anti-LC-1 in isolation scores positively towards AIH-2 diagnosis [9]. Anti-LC-1 positivity can be used as a marker of hepatocellular inflammation in AIH, since the presence and titre of this antibody correlate with disease activity [23]. Anti-LC-1 binds the folate-metabolising enzyme formimino transferase cyclodeaminase (FTCD), which is present at high density in the liver. In terms of IFL, anti-LC-1 stains the liver cell cytoplasm with relative sparing of the centrilobular area. When present together, anti-LC-1 pattern is usually obscured by the anti-LKM-1 pattern. To overcome this, anti-LC-1 can be detected in isolation using liver cytosol in double-dimension immunodiffusion or counterimmunoelectrophoresis, with the use of a positive reference serum, or by ELISA detecting reactivity to its target FTCD [4]. 4.3.6. Anti-soluble liver antigen/liver-pancreas antibodies Originally believed to be distinct autoantibodies, anti-SLA and antiLP are now known to be the same entity. Anti-SLA/LP is routinely detected by radioimmunoassay or ELISA [4], while newly developed diagnostic assays based on the molecular target are being fully evaluated. The observation that anti-SLA/LP could be detected in the absence of seropositivity for conventional AIH autoantibodies, led to the suggestion of a third type of AIH positive only for this autoantibody. However, these early reports used a particularly high cut-off point for conventional autoantibody screening; therefore this distinct group of patients has not been accepted by the IAIHG. Though anti-SLA/LP have occasionally been reported in the context of anti-LKM-1 positive HCV infection, their presence is highly specific for the diagnosis of AIH, and its

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detection at the time of diagnosis identifies patients with a more severe disease and a worse prognosis [28]. The molecular target of anti-SLA/LP is a UGA tRNA suppressorassociated antigenic protein (tRNP (ser)sec ), more specifically O-Phosphoseryl-tRNA:selenocysteinyl-tRNA synthase (SepSecS) [29]. 4.3.7. Anti-neutrophil cytoplasmic antibodies ANCAs react to constituents of the cytoplasm of neutrophils to give a perinuclear (pANCA) or cytoplasmic (cANCA) IFL pattern. pANCAs are frequently detected in AIH-1, although the IFL pattern is somewhat atypical. Staining is associated with peripheral nuclear membrane components, hence the name of peripheral anti-nuclear neutrophil antibody (pANNA). Positivity for pANNA is very rare in AIH-2. In AIH-1, however, its detection can facilitate diagnosis, particularly when other autoantibodies are absent [9]. The proposed target of pANNA is a 50 kDa neutrophil-specific nuclear protein belonging to the nuclear pore complex, potentially the tubulin β chain 5 [23]. Conventional p-ANCA binds myeloperoxidase and is most commonly found in microscopic polyangiitis. The predominant target of c-ANCA, which is frequently detected in Wegener's granulomatosis, is proteinase 3. 4.3.8. Anti-asialoglycoprotein receptor antibodies (anti-ASGPR) The ASGPR – a type II transmembrane glycoprotein also known as hepatic lectin – is the only liver-specific autoantigen to be identified thus far. It was described in an attempt to identify possible autoantigenic targets expressed on hepatocytes in AIH, and is a component of the crude liver extract preparation known as lover specific protein (LSP). Almost 90% of AIH patients are anti-ASGPR seropositive; anti-ASGPR are found in combination with ANA, SMA and anti-LKM-1. Since titres correlate with indicators of inflammatory activity, anti-ASGPR can be used to monitor the efficacy of treatment. However, the detection of anti-ASGPR requires either purified or recombinant antigen, and the development of reliable molecular assays has been difficult, therefore their applicability to clinical practice is limited. Moreover, since these autoantibodies have also been detected in patients with viral hepatitis, drug-induced hepatitis and PBC, they are not disease specific [23]. 4.4. Histology Liver biopsy is essential in order to confirm the diagnosis of AIH and evaluate the severity of liver damage. Hepatitis at the portal– parenchymal interface, known as interface hepatitis (Fig. 2), is typical, but not exclusive, of AIH [30]. It is characterised by portal and periportal lymphocytic or lymphoplasmacytic infiltration, hepatocyte swelling and/or pycnotic necrosis. Lymphocytes, plasma cells and histiocytes surround individual dying hepatocytes at the portal/parenchymal interface and in the lobule. Though plasma cells are usually abundant at the interface and throughout the lobule, their presence in low number does not preclude the diagnosis of AIH. In AIH presenting acutely or during episodes of relapse, panlobular hepatitis is often present, and is associated with bridging necrosis and, in the case of fulminant presentation, massive necrosis. Though sampling variation is possible in needle biopsy specimens, especially when cirrhosis is present, the severity of the histological appearance is usually of prognostic value. Inflammatory changes surrounding the bile ducts are apparent in a small proportion of patients with AIH. This observation, suggestive of an overlap with sclerosing cholangitis, is more frequently reported in the paediatric setting [31]. 5. Treatment With the exception of fulminant presentation with encephalopathy, AIH responds to immunosuppressive treatment whatever the degree of liver impairment [20].

Please cite this article as: Liberal R, et al, Diagnostic criteria of autoimmune hepatitis, Autoimmun Rev (2014), http://dx.doi.org/10.1016/ j.autrev.2013.11.009

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R. Liberal et al. / Autoimmunity Reviews xxx (2014) xxx–xxx

Fig. 2. Portal and periportal cellular inflammation in autoimmune hepatitis: lymphocytes, monocytes/macrophages and plasma cells infiltrate the portal tracts and invade the surrounding parenchyma, resulting in the characteristic picture of interface hepatitis. Haematoxylin and eosin staining.

Standard treatment, consisting of prednisolone alone or in combination with azathioprine, induces remission (i.e. normal transaminases and IgG levels) in around 80% of the patients, including those with cirrhosis [20,32]. Once achieved, remission can be maintained with azathioprine alone [20]. In treatment failure, other drugs have been anecdotally tried including methotrexate, cyclophosphamide, tacrolimus, ursodeoxycholic acid, cyclosporine and mycophenolate mofetil [33], the latter two being those most frequently reported as alternative medications. Although some encouraging results have been described, the progress with these nonstandard treatments has evolved slowly. Liver transplantation is the ultimate treatment for AIH patients presenting with acute liver failure, or end-stage chronic liver disease and for those with hepatocellular carcinoma that meet the transplant criteria [34]. Although liver transplantation for AIH is very successful, it should be noted that AIH may recur after transplant [35]. Finally we note a dedicated issue devoted entirely to autoimmune liver disease, particularly autoimmune hepatitis and note that the liver, which is essential to immune tolerance, can itself become a victim of autoimmunity [36–40].

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Please cite this article as: Liberal R, et al, Diagnostic criteria of autoimmune hepatitis, Autoimmun Rev (2014), http://dx.doi.org/10.1016/ j.autrev.2013.11.009