Portal tract eosinophils and hepatocyte cytokeratin 7 immunoreactivity helps distinguish early-stage, mildly active primary biliary cirrhosis and autoimmune hepatitis.

We studied nondiagnostic liver biopsy specimens from 20 patients with definite primary biliary cirrhosis (PBC) and 18 with definite autoimmune hepatitis (AIH) to identify distinguishing features. All patients had early-stage disease; biopsy specimens were devoid of granulomas or diagnostic features of PBC or AIH. Diagnoses were based on serologic and clinical variables. Sixteen specimens from each group were immunostained with cytokeratin 7. The density of portal tract eosinophils and number with cytokeratin 7-reactive periportal hepatocytes were quantified. Sixteen of 18 patients with AIH and 13 of 20 with PBC had no or minimal bile duct injury. Histologic activity index scores were 5.8 in AIH and 5.7 in PBC. The mean portal eosinophil score was greater in PBC than in AIH. Cytokeratin 7 identified many central bile ducts that were obscured by portal inflammation. The mean periportal cytokeratin 7-reactive hepatocyte score was greater in PBC than in AIH. Portal eosinophils and cytokeratin 7 reactivity in periportal hepatocytes are supportive of PBC rather than AIH. No morphologic features were supportive of AIH. Cytokeratin 7 reactivity in periportal hepatocytes may be an early response to PBC-induced biliary obstruction in other regions of the liver.     

Increased expression of nuclear envelope gp210 antigen in small bile ducts in primary biliary cirrhosis

The sustained antibody response to nuclear envelope gp210 antigen indicates a group of primary biliary cirrhosis (PBC) patients at high risk for the progression to end-stage hepatic failure. To address this issue, we immunohistochemically studied the expression of gp210 antigen in needle liver biopsy specimens from PBC patients using a monoclonal antibody specific for gp210 antigen. The specimens from autoimmune hepatitis (AIH), chronic viral hepatitis B (CHB) and C (CHC) patients served as disease controls. The expression of gp210 antigen was apparently increased on the nuclear envelope of biliary epithelial cells (BECs) of small bile ducts in almost all specimens from PBC. In contrast, the expression of gp210 antigen was negative in BECs of small bile ducts in normal liver, while relatively weak anti-gp210 immunostaining was observed in AIH, CHC and CHB. In addition, the degree of gp210 expression in BECs of small bile ducts was positively correlated to that of portal inflammation, interface hepatitis and lobular inflammation in PBC. These results indicate that the increased expression of gp210 in small bile ducts, which is probably associated with damage to BECs by inflammation, is possibly involved in autoimmune response to gp210 leading to the progression to end-stage hepatic failure in PBC.     

Histological grading and staging in chronic hepatitis: its practical correlation

Although the histological features of various causes of chronic liver disease have been well described, usually the inflammatory activity of the disease is important after the cause has been established. Some patients have co-infection or concomitant liver disease and on occasion it is difficult to decide the treatment. In order to clarify the histological differences, we investigated the inflammatory activity among autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC), chronic hepatitis C (CHC) and chronic hepatitis B (CHB) in a standardized way using the modified histological activity index (HAI). According to the modified HAI, inflammatory activity is divided into four categories; categories A/D explains portal/periportal inflammation and categories B/C explains lobular activity. The inflammatory score of AIH tended to be greater in all categories from the early stage of fibrosis, whereas scores of PBC were lower, except for portal inflammation. Chronic hepatitis C patients had portal or periportal inflammation, and their inflammatory scores were linked to the development of fibrosis. Chronic hepatitis B patients tended to have severe lobular injury, but did not have a relationship between the inflammatory score and their stage. To know the distribution of inflammation using the modified HAI scoring system may be helpful and convenient in evaluating patients with chronic inflammatory liver disease.     

cDNA microarray analysis of autoimmune hepatitis, primary biliary cirrhosis and consecutive disease manifestation

While autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) usually have distinct clinical manifestations, some patients present with features of both conditions. Using cDNA microarrays, we analyzed and compared gene expression profiles in 8 patients with AIH, 9 with PBC, 8 with chronic hepatitis C (CHC), 8 with non-alcoholic steatohepatitis (NASH) and 9 with normal livers. We subsequently applied this method to a tissue sample from a 61-year-old woman with overlapping features of both AIH and PBC. A 61-year-old woman was admitted to our hospital for evaluation of elevated serum alkaline phosphatase. A liver biopsy showed accumulation of mononuclear cells around the bile duct cells, a feature characteristic of chronic non-suppurative destructive cholangitis (CNSDC). Three years later, her serum alanine aminotransferase (ALT) level had increased, and a liver biopsy demonstrated evidence of a severe form of hepatitis. A cDNA microarray analysis of both biopsies identified the molecular events associated with her altered histology. The expression profile of this patient, which was originally different from that of the other PBC patients, changed to an AIH pattern. Our results suggest that this patient has characteristics of both AIH and PBC.     

Increase of chemokine interferon-inducible protein-10 (IP-10) in the serum of patients with autoimmune liver diseases and increase of its mRNA expression in hepatocytes

To clarify the role of IP-10 in autoimmune liver diseases, we studied the serum levels of IP-10 in 14 patients with autoimmune hepatitis (AIH), 23 patients with primary biliary cirrhosis (PBC), and 65 patients with chronic viral hepatitis (20 type B and 45 type C). The hepatic expression of IP-10 mRNA and the correlation between the serum levels of IP-10 and clinical parameters were also evaluated. In addition to 20 healthy controls, 16 rheumatoid arthritis (RA) patients were included as an extrahepatic inflammatory disease. The serum level of IP-10 was significantly (P < 0.02) higher in patients with AIH, PBC, and chronic hepatitis B and C than in healthy controls, and it was significantly correlated (P < 0.05) with the serum levels of aspartate aminotransferase and alanine aminotransferase in patients with AIH, PBC, and chronic hepatitis B and C. The serum level of IP-10 was not elevated in RA patients. After successful treatment of AIH and chronic hepatitis C, the serum level of IP-10 decreased to the same level as in healthy volunteers. As we previously showed in cases with chronic hepatitis B or C, in situ hybridization in both AIH and PBC cases demonstrated the expression of IP-10 mRNA in hepatocytes around focal or lobular necrosis surrounded by infiltrating mononuclear cells, whereas IP-10 mRNA was not expressed in areas around the damaged bile ducts in PBC cases. The present results suggest that IP-10 is specifically produced by hepatocytes in inflammatory areas irrespective of the aetiology of hepatitis, and that IP-10 may help to recruit T cells to the hepatic lesions in autoimmune liver diseases as well as in chronic viral hepatitis.     

Interferon-alpha therapy does not modulate hepatic expression of classical type I interferon inducible genes

Hepatitis C virus (HCV) infection is a major cause of chronic liver disease. Treatment with interferon-alpha(2) (IFN-alpha(2)) can induce viral clearance and marked biochemical and histological improvement. IFN-alpha(2) treatment has been shown to stimulate the expression of type I IFN regulated genes in peripheral blood mononuclear cells (PBMCs) of hepatitis C patients; however, whether it affects hepatic expression remains unknown. This study thus aimed at comparing hepatic gene expression with particular emphasis on type I IFN inducible genes in patients with chronic hepatitis C before and during an IFN-alpha(2) monotherapy. Responsiveness to IFN-alpha(2) therapy was monitored by determining serum and hepatic viral load. Differential gene expression analysis was performed by two different techniques, namely suppression subtractive hybridization (SSH) and differential display (DD). Expression of two prototype type I IFN regulated genes was quantified in further PBMC and liver samples. Among different genes found to be up-regulated during an effective, that is, virus clearing, IFN-alpha treatment, only a single one was identified which can be accounted to type I IFN responsive genes. Parallel quantitative real time PCR analyses demonstrated significant induction of the type I IFN regulated genes MxA and PKR in PBMC, but not in the liver. Taken together, while IFN-alpha treatment leads to the induction of type I IFN regulated genes in PBMC, such an induction appears not to occur in the liver of hepatitis C patients. The mechanism by which IFN-alpha treatment causes viral clearance might be independent of hepatic activation of type I IFN regulated genes.     

Capillarization of hepatic sinusoid by liver endothelial cell-reactive autoantibodies in patients with cirrhosis and chronic hepatitis

The special features of liver sinusoidal endothelium (LSE) are crucial for normal liver physiology. Cirrhotic livers, especially in primary biliary cirrhosis (PBC), are characterized by transformation of the LSE into a continuous, vascular type. The transformation is important for disease progression and explains some of the pathological hallmarks of the cirrhotic liver. Here, we investigated the presence of liver sinusoidal endothelial cell (LSEC)-reactive autoantibodies (Abs) in the sera of patients with autoimmune liver diseases, and assessed the ability of these Abs to transform LSE into vascular endothelium. Compared to healthy individuals (9%), significantly higher numbers of patients with PBC (59%; P < 0.001) and autoimmune hepatitis (AIH) (32%; P < 0.05) had Abs against LSECs. Incubation of primary LSEC cultures with F(ab')(2) fragments of anti-LSEC Abs isolated from sera of patients with PBC and AIH, induced 1) cell surface expression of vascular endothelium-associated markers, CD31, and factor VIII-related antigen; 2) significant production of fibronectin, laminin and collagen type IV; 3) loss of fenestrae, formation of tight junctions and Weibel-Palade bodies. Deposition of immunoglobulins on LSECs were found in liver biopsies of AIH and PBC patients. Thus, anti-LSEC autoAbs transform LSE into a vascular type and may therefore play an important role in the development of hepatocellular failure and portal hypertension in PBC and AIH patients.     

The role of complement activation in autoimmune liver disease

IntroductionThe complement system, an essential part of the innate immune system, is involved in various autoimmune diseases. Activation of the complement system by autoantibodies results in immune activation and tissue damage. At the moment little is known about the role of the complement system in autoimmune liver disease, including primary biliary cholangitis (PBC), primary sclerosing cholangitis (PSC) and autoimmune hepatitis (AIH). Since inhibition of the complement system is currently being tested in several autoimmune diseases as a therapeutic option, its role in autoimmune liver disease requires further clarification.MethodsA review of the literature was performed on studies investigating complement activation in PBC, PSC and AIH. Since data on AIH were lacking immunohistochemical staining for IgG, C1q, C3d, C4d and C5b9 was performed on liver tissue of nine AIH patients, two healthy controls and one positive control (acute liver failure caused by paracetamol intoxication).ResultsImmunohistochemical analysis in AIH revealed increased production of C3 and C4 by hepatocytes. Despite a strong staining for IgG in the immune infiltrate in AIH, C3d, C4d and C5b9 deposition was only present in one AIH patient and the deposition was restricted to the interface between portal tracts and liver parenchyma. No deposition was found in all other AIH patients or healthy controls.Literature review showed raised plasma C3 and C4 levels in AIH, PBC and PSC patients compared to healthy controls. For PBC and PSC no complement depositions at the bile ducts were reported.Conclusion and discussionAlthough complement is involved in various autoimmune diseases, the role of complement in autoimmune liver disease seems limited. Therefore it is unlikely that complement inhibition will become a novel treatment option for these diseases.     

Asymptomatic systemic sarcoidosis arising 5 years after IFN-alpha treatment for chronic hepatitis C: a new challenge for clinicians.

Interferon (IFN)-induced sarcoidosis is well documented. Herein, we report the case of a patient with chronic hepatitis C (CHC) who developed IFN-alpha-induced sarcoidosis. The clinical features of this case make it unique among all cases so far described. The patient was, in fact, asymptomatic for sarcoidosis, and the disease, characterized by liver and lung granulomatosis, was discovered by chance during the CHC follow-up. The diagnosis was made 5 years after IFN-alpha discontinuation. A pathogenetic role for IFN-alpha in our patient is supported by a liver biopsy performed before the therapy with IFN-alpha was started, showing no evidence of granulomatous localizations. This case suggests that the incidence of sarcoidosis during IFN-alpha treatment is underestimated. A search for clinical and laboratory findings typical of the disease, as well as a liver biopsy, should always be included in the follow-up of CHC patients undergoing therapy with IFN-alpha.     

A spectrum of histopathologic findings in autoimmune liver disease

We retrospectively studied 42 liver biopsy specimens from 39 patients who met serologic and histologic criteria of autoimmune liver diseases. We found 10 cases of overlap syndrome (OLS), 10 autoimmune cholangitis (AIC), 10 primary biliary cirrhosis (PBC), and 9 autoimmune hepatitis (AIH) type 1. The following results were obtained: (1) Granulomas and biliary duct lesions were more prominent in PBC and AIC than in OLS and AIH. (2) Bile duct loss was not observed in AIH cases. (3) Features of hepatocellular damage such as piecemeal necrosis, spotty lobular necrosis, and confluent necrosis, were much more prevalent in OLS and AIH than in PBC and AIC. (4) HLA-DR antigen expression by hepatocytes was more frequent in AIH and OLS, whereas the expression of the same antigen by the bile duct epithelium was more frequent in PBC and AIC. We conclude there is a morphologic spectrum in autoimmune liver diseases, in which PBC forms one end of the spectrum, AIH the other, OLS the middle but closer clinically and histologically to AIH than to PBC, and AIC, which seems to be an antimitochondrial antibody-negative subtype of PBC.     

Autoimmune hepatitis in primary Sjogren's syndrome: pathological study of the livers and labial salivary glands in 17 patients with primary Sjogren's syndrome

Although primary Sjogren's syndrome (pSS) is an autoimmune exocrinopathy, the involvement of liver has been reported. Because no study focusing on autoimmune hepatitis (AIH) in pSS has been published, the purpose of the present study was to perform a clinical and histological examination of the liver, focusing on AIH, in 17 pSS patients. The patients had liver enzyme abnormalities without hepatitis virus infection. In all cases, biopsied livers were examined, and in 10 cases biopsied labial salivary glands were also examined histologically. Based on the authors' diagnostic criteria for AIH in pSS, the liver diseases consisted of AIH (eight cases, 47%), primary biliary cirrhosis (PBC; six cases, 35%), non-specified chronic hepatitis (two cases, 12%) and acute hepatitis (one case, 6%). Lymphoplasmacytic infiltrate, with predominancy of CD3(+) T cells, was noted in both the liver and salivary glands in the patients with AIH. The patients with AIH with severe interface hepatitis had a good response to immunosuppressive therapy. The comparison of liver histology between the PBC with pSS group and the PBC without pSS group showed that the incidence of lymphoid non-suppurative cholangitis was higher in PBC with pSS. In conclusion, the present study offers new information on the relatively common occurrence, diagnostic criteria and treatment effects of AIH in pSS.     

Intrahepatic expression of the co-stimulatory molecules programmed death-1, and its ligands in autoimmune liver disease

Liver-infiltrating T cells play an essential role in the immunopathogenesis of autoimmune liver disease. Programmed death-1 (PD-1) and its ligands, B7-H1/PD-L1 and B7-DC/PD-L2, are new CD28-B7 family members that are involved in the regulation of immune responses. The ligation of PD-1 inhibits T-cell receptor-mediated T cell proliferation and cytokine production, and PD-1-deficient mice develop various organ-specific autoimmune diseases. To investigate the expressions of PD-1 and its ligands in autoimmune liver disease, in particular autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC), immunohistochemical analysis was performed. Liver biopsy specimens obtained from 17 patients with AIH and PBC were studied. PD-1 was expressed on more than half of the liver-infiltrating T cells within the portal tract. Some of the intrahepatic T cells expressed B7-H1 in patients with AIH and PBC. B7-H1 and B7-DC were mainly expressed on some Kupffer cells (KC) and liver sinusoidal endothelial cells (LSEC) within the sinusoids and their expression was upregulated in autoimmune liver disease. These results suggest that the interaction of PD-1 on T cells with increased expression of B7-H1 and B7-DC on KC and LSEC might be involved in the downregulation of autoreactive lymphocytes and result in the regulation of pathogenesis in autoimmune liver disease.     

The hepatitic/cholestatic "overlap" syndrome: an Italian experience

BACKGROUND: Patients with hepatitic and cholestatic autoimmune liver disease ("overlap syndrome") represent a diagnostic and therapeutic challenge. AIM: To evaluate the prevalence of the "hepatitic/cholestatic overlap" in a large series of consecutive patients with cholestatic autoimmune liver disease. METHODS: We re-evaluated the diagnosis of 235 patients with autoimmune liver disease, including 70 with type 1 autoimmune hepatitis (AIH), 142 with primary biliary cirrhosis (PBC), and 23 with primary sclerosing cholangitis (PSC), using the revised International Autoimmune Hepatitis Group (IAIHG) scoring system. Anti-mitochondrial, anti-nuclear, anti-smooth muscle, anti-liver kidney microsomal type 1, anti-liver cytosol type 1, perinuclear anti-neutrophil nuclear and anti-soluble liver antigen antibodies were evaluated in each patient. RESULTS: Ten patients (3 with a previous diagnosis of PBC and 7 of PSC) scored as "probable" or "definite" AIH. These patients did not have a specific autoantibody profile. CONCLUSIONS: Among patients with PBC, the occurrence of a PBC/AIH overlapping syndrome is rare (2.1%), whereas among patients with PSC an overlap between PSC and AIH is frequent (30.4%). Whether patients with the hepatitic/cholestatic overlap syndrome would benefit from a combination therapy with immunosuppression and ursodeoxycholic acid remains to be established.     

Animal models for autoimmune hepatitis

The liver is the target of adverse immune reactions in three putative autoimmune diseases: autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC). These three diseases can be distinguished by clinical, histological, and immunological features. However, especially on the level of specific antibody formation an overlap can occur, which sometimes complicates diagnosis. In this review, we will concentrate on autoimmune hepatitis and the current state of animal models for this severe disease. AIH is characterized by the presence of interface hepatitis and portal plasma cell infiltration, hypergammaglobulinemia, and autoantibodies. The hallmark of type 2 AIH is the generation of autoantibodies of the LKM-1 type. The major target of these antibodies is the cytochrome P450 isoform 2D6 (CYP2D6). In the past several attempts have been made to develop a reliable animal model that reflects the persistent hepatic destruction that occurs in human AIH. However, most models were only successful in causing a transient form of hepatic damage and often used rather complex ways of disease induction.