Section 3 -

Chronic Hepatitis Linda D. Ferrell University of California San Francisco, CA

Hepatitis B In chronic HBV infections, ground glass cells containing hepatitis B surface antigen (HBsAg) can be seen on H&E stained sections. The cytoplasm of these cells have a homogeneous lightly-staining, eosinophilic appearance and often resembles a cytoplasmic inclusion body displacing the nucleus to the periphery of the cell. The presence of HBsAg can be confirmed by positive staining with an immunoperoxidase or orcein stain; the former is the more sensitive technique and easier to interpret. HBV can also be identified by immunohistochemical staining for hepatitis B core antigen (HBcAg). HbsAg has a blue or green hue on trichrome stain (depending on the blue or green collagen stain used). These stains can be variably positive, depending on sample size and stage of disease. In acute HBV infections, surface antigen staining is usually nega­tive as the infected cells are effectively cleared from the liver [1]. The positive staining is, instead, usually seen in chronic hepatitis where the viral proliferation outstrips the immune clearance. In late-stage chronic hepatitis or in long-term healthy carriers, HBsAg can be present, but not HBcAg. This is thought to be due to viral genomic incorporation into hepatocytic nuclear DNA so that the HBsAg is coded for, but viral DNA is not [1]. HBcAg positivity can often be found in immuno­compromised patients and cytoplasmic core antigen may suggest active replication of HBV. Staining for HBcAg is much more common in the nucleus than in the cytoplasm. Hepatitis C Hepatitis C virus seems to induce a milder form of hepatitis than HBV and some typical histological differences between the two entities have been noted other than the presence of ground glass cells in HBV. Acute hepatitis C can occur, but in only very rare instances, and essentially cannot be differentiated histologically from other forms of acute hepatitis. In chronic HCV, lymphocyte infiltration is greater and reactive changes in the duct epithelium are often seen, but the duct damage is not extensive or severe enough to cause ductopenia. Lymphocytic infiltrates in the sinusoids are often prominent and lym­pho­cytic aggregates in the portal zones are typical findings. Fatty change of hepatocytes is greater than that seen in HBV [2]. No currently commercially available immunohistochemical method of identifying HCV in liver biopsies appears to be consistently reliable [3, 4] (personal observation and communications from other pathologists). Thus, PCR technology still serves as the gold standard for viral identification and to determine viral load. Aggressive Variants of HBV and HCV Both HBV and HCV can cause more aggressive and cholestatic forms of hepatitis in the setting of an immunocompromised patient. For HBV, this distinct variant was originally described in HBV-infected recipients of liver allografts [7, 8]. Since then, this aggressive form has also been described in other patients who are immunocompromised but do not have a liver allograft [9]. The clinical course is characterized by the rapid progression to liver failure and death. The microscopic picture is characterized by periportal fibrosis, cholestasis, hepatocyte ballooning, and a relatively scant inflammatory infiltrate [7, 8]. The hepatitis B viral core and surface antigens are overexpressed in these cases [7] and the transcription of HBV DNA has been shown to be enhanced [10]. Immunohistochemical staining for HB core and surface antigen show marked positivity, involving the vast majority of the hepatocytes. Likewise, some patients with HCV who are immunocompromised can develop an aggressive cholestatic form, which frequently results in rapidly progressive liver failure [11]. These patients also have the cholestasis and ballooned hepatocytes, usually with scant inflammatory infiltrate, with or without the fibrosis that is characteristic of the HBV variant. Hepatocyte dropout/necrosis may be prominent. In our experience, these patients generally have high viral titers by PCR analysis. It has been speculated that in these patients, the immunosuppression enhances the dominance of viral quasispecies of enhanced pathogenic potential, leading to the severe recurrent disease. Both of these aggressive forms typically lead to liver failure within a year or so of onset, so the outcome is dismal. Table 1: Non-viral Causes of Chronic Viral-like Hepatitis TYPE CLINICAL PRESENTATION HISTOLOGY Autoimmune Active chronic hepatitis; can be rapidly progressive Similar to hepatotropic viruses, plasma cells may be prominent Steatohepatitis, generally Nonalcoholic type Mild chronic hepatitis Mild chronic hepatitis-like changes, with ballooned hepatocytes, fat, centrizonal fibrosis, possible Mallory hyaline Wilsons disease Acute, including fulminant; chronic hepatitis Similar to hepatotropic viruses; copper deposits Alpha-1-antitrypsin deficiency Chronic hepatitis Similar to hepatotropic viruses; AAT globules Drug reactions Mostly acute, rarely chronic hepatitis Similar to hepatotropic viruses, may see prominence of eosinophils, granulomas, or cholestasis Autoimmune Hepatitis Autoimmune hepatitis (AIH), (no longer called "lupoid hepatitis" or used with the term "chronic" has been subclassified into three forms characterized by differences in clinical presentations or the serum antibodies identified. Most patients respond favorably to immunosuppressive therapy (prednisone), with a resultant decrease in the activity of the disease, so their distinction from viral hepatitis or chronic hepatitis due to other causes is important. ClassicalType 1 AIH presents as hepatitis that primarily affects females in the age ranges of 2-15 and 35-70 years. These patients have positive antinuclear antibody (ANA) titers and some association (in about 10-20%) with other autoimmune disorders. Type 2 AIH seems to present more in children (girls more than boys, age less than 15 years) and is frequently associated with other autoimmune dis­orders (about 20%) as well. The anti-liver-kidney microsomal (LKM-1) antibody is present in these patients, who often present with the clinical picture of an acute or fulminant hepatitis. In these cases, it is often difficult to determine whether a preexisting chronic hepatitis was present before the acute phase, so the lesions may be truly more acute in nature. The type 2 variant in adult patients has also been associated with coinfection with hepatitis C virus. In this group, some patients have been reported to respond favorably to interferon therapy while others have been reported to show flares of hepatitis during interferon therapy, and then, subsequently, respond to steroid therapy. It is generally thought that infection with hepatitis C may, in fact, trigger an autoimmune reaction to the LKM-1 target antigen in patients who have a genetic predisposition as evidenced by a high incidence of certain HLA antigens. The most recently described variant, Type 3AIH, also occurs mostly in women but with a later onset (mean age 37). About 25% of these patients will display the anti-soluble liver antigen (SLA) only. These patients are seronegative for ANA and liver-kidney-microsomal antibody, but 75% will have anti-smooth muscle antibody (SMA) or liver membrane antibody (LMA). In contrast to types 1 and 2, systemic autoimmune manifestations are not typical. Diagnostic Considerations AIH is often characterized by prominent plasma cell infiltrates not typically seen in HCV or HBV hepatitis, but may display the prominent lymphoid aggregates and duct damage seen with HCV hepatitis. However, patients with AIH seem to have more diffuse and severe piecemeal necrosis (interface hepatitis), an increased incidence of bridging and confluent necrosis, and more rapid progression to cirrhosis than patients with HCV. In addition, infiltration of mononuclear inflammatory cells tends to be diffuse with AIH and focal with chronic HCV. Multinucleate hepatocytic giant cells have also been reported to occur in autoimmune hepatitis. Giant Cell Hepatitis Recently, the ultrastructural analysis of a severe hepatitis with giant cells associated with the presence of paramyxovirus has been described in children and adults. This hepatitis has been termed "syncytial giant cell hepatitis", but this viral etiology is currently questioned. In addition, giant cell hepatidites have also been described as rare forms of viral, drug, autoimmune, or post-transplantation hepatitis; many of these may have a more chronic type of course. Wilsons Disease This disease is included in those entities that can cause chronic hepatitis, but the more complete picture including acute hepatitis presentation was discussed in the section on Acute Hepatitis. See that syllabus section for details on clinical considerations, pathologic findings, and use of special stains for identification of copper. Additional clinical screening criteria, testing standards, and genetic characteristics are additionally discussed below. Hepatic Presentation Guidelines as proposed by Brewer, Fink, and Hedera [28]. Viral-negative hepatitis. Always screen for WD Viral-positive hepatitis. Screen for WD if there are aspects of the disease (such as neurological or behavioral symptoms) that do not fit a viral etiology Cirrhosis, under the age of 40. Always screen for WD Cirrhosis, age 40-60. Screen for Wilsons unless a definite etiology can be established Acute, unexplained liver failure. Always screen for WD Recommended screening methods for patients with hepatic presentation (Brewer et al, ref 28) Serum ceruloplasmin (Cp). Easy to do, and low serum Cp will point to diagnosis in 75%. However, Cp levels can be normal as noted above, and many heterozygotes have an intermediately low value. Thus, low Cp needs further workup. 24-hour urine copper. This value will be >100µg in symptomatic WD. A lower value in symptomatic, untreated patients excludes the diagnosis. If the liver disease is long-standing and has an obstructive component, the urine copper can be elevated in the absence of Wilson's disease. Slit-lamp examination for Kayser-Fleisher (KF) rings is close to 100% diagnostic for patients with neurological or psychiatric symptoms, but not nearly as helpful for hepatic presentations. In fact, patients with obstructive liver disease may also develop KF rings. Quantitative copper assay on liver biopsy. This is the gold standard for diagnosis over 200 µg/g dry wight of liver (normal 20-50 µg/g ). However, these levels are not specific as patients with obstructive liver disease (such as primary biliary cirrhosis, primary sclerosing cholangitis) can have copper levels similar to those seen in Wilson's disease. Heterozygotes may have some increase in levels, but typically not over 125µg/g. The liver biopsy can be negative by staining for copper and positive by quantitative analysis. Gene for Wilsons disease The gene responsible for WD has been identified and designated as ATP7B. This gene codes for a copper-binding, membrane bound ATPase. The mutations in this gene identified are numerous, and no one mutation accounts for more than 30% of the total group of mutations, so most patients are compound heterozygotes. Thus, it is also not practical to develop a DNA test for diagnosis. Perhaps the large number of mutations may account for some of the variability in clinical and pathological presentations. Alpha-1-Antitrypsin Deficiency Clinical Considerations Alpha-1-antitrypsin (AAT) deficiency is characterized by the accumulation of AAT in hepatocytes. Numerous alleles for the AAT gene have been described, but the most common allele is PiM. Homozygotes MM have normal levels of AAT. The most common alleles resulting in AAT deficiency are PiZ and PiS, with liver pathology occurring predominantly in patients with at least one Z allele. Homozygotes for the Z allele may arise in patients of all ages (from neonates to adults) for unknown reasons, while MZ phenotypes present in adults. Neonates present with cholestatic jaundice, and may develop serious liver disease with cirrhosis and death during childhood. Those with the most severe course also have a paucity of bile ducts. Other poor prognostic factors include persistence of jaundice beyond one year of age and fibrosis on liver biopsy. When the lesion presents in adulthood, the patients tend to present with cryptogenic cirrhosis, with the AAT globules identified on liver biopsy. Diagnostic Considerations The characteristic microscopic finding is the presence of PASD positive cytoplasmic globules found mainly in the periportal zone or periphery of the cirrhotic nodule. The globules are usually present in neonates as well as in adults. These globules will vary in size, and their exact composition as AAT should be confirmed by immunohistochemical means. Typically stained larger globules show a distinct outer dark ring with a pale center. Ultrastructural examination reveals dilated cisternae that contain flocculent or granular to fibrillar material. Large numbers of AAT globules have also been noted in alcohol associated disease and other entities, so phenotyp­ing is necessary for definitive diagnosis. Drug Hepatitis Drug reactions must often be considered in the differential diagnosis of hepatitis of various types, but drugs only rarely truly cause a chronic hepatitis. Such agents can include etretinate, lisinopril, sulphonamides, and trazadone. Some of these chronic syndromes overlap features of autoimmune hepatitis, including the presence of autoimmune antibodies (SMA, ANA, anti-DNA), as in the case of minocycline, nitrofurantoin, and oxyphenisatin Biliary Disorders A hepatitis-like mononuclear infiltrate that could be confused for hepatitis may be frequently seen in primary biliary cirrhosis, especially in the early stages. The infiltrate consists of increased sinusoidal lymphocytes and interface hepatitis, but the hepatocytes in the areas of infiltrate often do not show much evidence of injury, in contrast to chronic hepatitis. However, spotty hepatocyte necrosis (apoptosis) can be seen quite frequently, and rarely, centrilobular necrosis with venulitis can be noted. In such cases, clinical correlation is important and examination for early changes of ductopenia or granulomatous lesions is essential. Primary sclerosing cholangitis can also show some degree of similar hepatitis-like infiltrates, although it does not tend to have the spotty necrosis. Chronic Hepatitis - Grading and Staging Table 1: Grading and Staging - Portal and Lobular Inflammatory Activity - Grade* Grade Pathology 0 None or minimal portal inflammation, no hepatocyte necrosis 1 Portal and/or lobular inflammation without significant evidence of hepatocyte necrosis 2 Mild interface hepatitis (piecemeal necrosis and disruption of limiting plate) and/or focal lobular hepatocyte necrosis 3 Moderate limiting plate (interface hepatitis) and/or severe focal cell necrosis (confluent necrosis) 4 Severe limiting plate and/or bridging necrosis Fibrosis - Stage Stage Pathology 0 No fibrosis 1 Mildly enlarged portal tracts (fibrous portal expansion) 2 Periportal or portal-portal septa (not prominent) with intact architecture, or periportal fibrosis 3 Septal fibrosis with architectural distortion or bridging fibrosis; no obvious cirrhosis 4 Probable or definite cirrhosis *Grade 1 inflammatory changes were formerly termed chronic persistent hepatitis; Grade 2-mild, 3-moderate, and 4-severe chronic active hepatitis. Table 2-A complex grading and staging system (modified from Ishak et al 1995) A. Periportal or periseptal interface hepatitis (piecemeal necrosis) Absent 0 Mild (focal, few portal areas) 1 Mild/moderate (focal, most portal areas 2 Moderate (continuous around <50% of tracts or septa) 3 Severe (continuous around >50% of tracts or septa) 4 B. Confluent necrosis Absent 0 Focal confluent necrosis 1 Zone 3 necrosis in some areas 2 Zone 3 necrosis in most areas 3 Zone 3 necrosis + occasional portal-central (P-C) bridging 4 Zone 3 necrosis + multiple P-C bridging 5 Panacinar or multiacinar necrosis 6 C. Focal (spotty) lytic necrosis, apoptosis and focal inflammation* Absent 0 One focus or less per 10x objective 1 Two to four foci per 10x objective 2 Five to ten foci per 10x objective 3 More than ten foci per 10x objective 4 D. Portal inflammation None 0 Mild, some or all portal areas 1 Moderate, some or all portal areas 2 Moderate/marked, all portal areas 3 Marked, all portal areas 4 * Does not include diffuse sinusoidal infiltration by inflammatory cells N.B. Other features such as bile duct damage should be noted but not scored Architectural changes, fibrosis and cirrhosis No fibrosis 0 Fibrous expansion of some portal areas, with or without short fibrous septa 1 Fibrous expansion of most portal areas, with or without short fibrous septa 2 Fibrous expansion of most portal areas with occasional portal to portal (P-P) bridging 3 Fibrous expansion of portal areas with marked bridging (portal to portal (P-P) as well as portal to central (P-C) 4 Marked bridging (P-P and/or P-C) with occasional nodules (incomplete cirrhosis) 5 Cirrhosis, probable or definite References for HBV, HCV, grade/stage Gerber M, Thung S. The diagnostic value of immunohistochemical demonstration of hepatitis viral antigens in the liver. 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