I.G. McFarlane

Hepatitis C virus antibodies in chronic active hepatitis: pathogenetic factor or false-positive result?

An enzyme immunoassay (Ortho-HCV ELISA) for antibodies against the hepatitis C virus (HCV) was used to test 143 serum samples from 53 patients with autoimmune chronic active hepatitis (AI-CAH). Optical density (OD) values in the assay correlated closely with serum globulin (r = 0.8846, p much less than 0.0005) and IgG (r = 0.6281, p less than 0.0005) concentrations but not with immunosuppressant therapy. OD values were positive in 20 (65%) of 31 with active disease and in only 1 (5%) of 22 in remission (p less than 0.0005). The association between positive results and active disease and high serum globulin levels was confirmed by serial studies in 6 of the patients. In contrast, none of 31 patients with primary biliary cirrhosis and only 2 of 24 with non-hepatic disorders associated with high IgG concentrations were positive, and these controls showed no correlation between OD values and serum globulins or IgG. The findings suggest that serum from AI-CAH patients may contain a component that gives false-positive results in the assay.

Antibodies to hepatitis C virus in autoimmune liver disease: evidence for geographical heterogeneity

To resolve conflicting reports about the occurrence of antibodies against hepatitis C virus (HCV) in patients with autoimmune chronic active hepatitis (AI-CAH), sera from UK and Italian patients were tested with the original anti-HCV assay (Ortho) and a novel anti-HCV assay (UBI) based entirely on synthetic HCV peptides. 28 (60%) of 47 Italian patients with type-1 AI-CAH were anti-HCV-positive by Ortho ELISA, 25 of whom were also strongly positive by the UBI assay. 15 (60%) of 25 UK patients with type-1 AI-CAH were HCV-positive by Ortho ELISA but only 2 were positive by the UBI assay. Similarly, 29 (88%) of 33 Italian patients with type-2 AI-CAH, but 0 of 10 UK patients, were very strongly anti-HCV-positive with the UBI assay. Italian patients with AI-CAH appear to have a high frequency of genuine exposure to HCV, whereas seropositivity by the Ortho HCV ELISA in UK patients is likely to represent a false-positive result. These findings indicate important geographical and/or genetic influences in autoimmune liver disease among different populations.

Serum autoantibodies reacting with the hepatic asialoglycoprotein receptor protein (hepatic lectin) in acute and chronic liver disorders

Circulating autoantibodies reacting with affinity-purified, hepatic asialoglycoprotein receptor protein, hepatic lectin (HL), were detected by radioimmunoassay in 15 (83%) of 18 patients with autoimmune chronic active hepatitis (AI-CAH) who had active disease, at titres that showed a positive correlation (P less than 0.05) with severity of periportal inflammation assessed histologically. In contrast, 10 AI-CAH patients whose disease was in remission were all anti-HL seronegative. Anti-HL was also detected in 16 (73%) of 22 patients with hepatitis B virus-related CAH-a similar frequency to that in active AI-CAH but at significantly lower (P less than 0.005) titres. Only 1 of 8 patients with chronic active liver disease due to presumed non-A, non-B (NANB) viral infection and 5 (22%) of 23 with primary biliary cirrhosis were anti-HL seropositive (P less than 0.001 vs active AI-CAH and HBV-CAH) and there was no correlation with severity of periportal inflammation. Anti-HL antibodies were also found in sera from 7 (35%) of 20 patients with acute virus B hepatitis (AVH-B) but were not detected in 10 patients with AVH-A nor in 12 with AVH due to presumed NANB infection. Anti-HL was not found in sera of 12 patients with autoimmune thyroid disease. Hepatic lectin, a highly purifiable, liver-specific cell surface component, by analogy with the acetylcholine and thyrotropin receptors which are, respectively, targets of the pathogenetically-related autoimmune reactions in myasthenia gravis and autoimmune thyroid disease, may be an important target of autoreactions in liver disease.

ANTIBODIES TO LIVER-SPECIFIC PROTEIN PREDICT OUTCOME OF TREATMENT WITHDRAWAL IN AUTOIMMUNE CHRONIC ACTIVE HEPATITIS

The value of serum autoantibodies against the liver-specific membrane lipoprotein (LSP) complex in predicting the outcome of treatment withdrawal was assessed in 22 patients with autoimmune chronic active hepatitis who had been maintained in remission on azathioprine and/or prednisolone. At the start of treatment withdrawal 8 patients had anti-LSP antibodies. Within 7-20 weeks (median 12 weeks) all 8 showed biochemical (aminotransferase greater than 200 IU/l) amd histological (piecemeal necrosis) evidence of reactivation of disease. Of the remaining 14 patients, who did not have anti-LSP antibodies at the start of treatment withdrawal, 8 became seropositive within 2-13 weeks (median 6 weeks) and all 8 had relapsed by 21 weeks. In 7 of these eight, anti-LSP first appeared 3 to 10 weeks (median 6 weeks) before and, in the eighth, coincident with biochemical evidence of relapse. The remaining 6 patients continued to be anti-LSP negative and showed no evidence of reactivation of disease throughout the study period (median 29 weeks, range 26-32 weeks). The results show that the presence of anti-LSP antibodies in patients who are apparently in complete remission is invariably associated with reactivation of disease during treatment withdrawal.

Lymphocyte cytotoxicity for isolated hepatocytes in alcoholic liver disease.

To determine whether an autoimmune reaction to liver-specific proteins occurs in alcoholic liver disease, the cytotoxic effect of lymphocytes on isolated hepatocytes was determined in 27 alcoholic patients. Cytotoxicity was demonstrated in 15 of 17 patients with a histological diagnosis of alcoholic hepatitis, but in none of 10 with other forms of alcoholic liver disease. The ability of a liver-specific membrane lipoprotein to block the reaction suggests that sensitization to this antigen is responsible for the cytotoxicity. The demonstration of cytotoxicity using T cell-depleted, but not B and K cell-depleted, lymphocytes suggests an antibody-dependent cell-mediated reaction. There was a close correlation between cytotoxicity and the presence of alcoholic hyalin, liver cell necrosis, and piecemeal necrosis, but not with other histological features, immunoglobulines, autoantibodies, or standard liver function tests. The persistence of this autoimmune response, induced in some way by alcohol or one of its metabolites, may be important in the progression of acute alcoholic hepatitis to chronic liver disease.

Cell-mediated Immunity to Human Tamm-Horsfall Glycoprotein in Autoimmune Liver Disease with Renal Tubular Acidosis

Cell-mediated immune responses to Tamm-Horsfall glycoprotein isolated from human urine were investigated using the leucocyte migration test. Abnormal responses were found in 91% of patients with active chronic hepatitis or primary biliary cirrhosis with an associated renal tubular acidosis (R.T.A.) but in only 19% of those without R.T.A. In nearly all of a group of patients without autoimmune liver disease and in a control group of normal subjects results were within normal limits. In addition, using an immunofluorescent technique with rabbit antibody to human Tamm-Horsfall glycoprotein, it was possible to show the presence in human liver cell membrane of material reacting immunologically as Tamm-Horsfall. These findings suggest that the development of an immune response to this glycoprotein, initiated by release of cross-reacting antigens from damaged hepatocytes, could be the mechanism underlying the occurrence of R.T.A. in some patients with autoimmune liver disease.

Cell-mediated Immune Response in Primary Biliary Cirrhosis to a Protein Fraction from Human Bile

Cell-mediated immune responses to a protein fraction of human bile have been demonstrated, using the leucocyte migration test, in eight out of 10 patients with primary biliary cirrhosis but in only three out of nine with active chronic hepatitis. In the latter condition sensitization to a liver-specific hepatocellular antigen was found more frequently (five out of nine patients) than in primary biliary cirrhosis (two out of 10). These results, as well as the granuloma formation observed histologically, suggest that the initial bile duct lesion in primary biliary cirrhosis may be associated with a cell-mediated response to antigens—perhaps derived from bile duct epithelial cells—which may be normal constituents of hepatic bile.

T-lymphocyte sensitization to hepatocyte antigens in autoimmune chronic active hepatitis and primary biliary cirrhosis

Cultured with a liver-derived lipoprotein complex, T lymphocytes from 42 of 45 patients with autoimmune chronic active hepatitis generated migration inhibitory factor compared with 16 of 33 patients with primary biliary cirrhosis. Unlike T lymphocytes from patients with primary biliary cirrhosis, the T-cell reactivity of patients with chronic active hepatitis was always suppressed by T cells from normal subjects and, with two exceptions, by T cells from patients with primary biliary cirrhosis, even when these latter cells exhibited sensitization to this same antigen complex. Using a component of the whole complex, the asialoglycoprotein receptor as antigen, migration inhibitory factor was invariably released by T cells from patients with autoimmune chronic active hepatitis, but from only 2 of 8 patients with primary biliary cirrhosis sensitized to the whole complex. Thus, in autoimmune chronic active hepatitis, but not in primary biliary cirrhosis, the asialoglycoprotein receptor is invariably a target for cellular immune reactions and is associated with a suppressor T-cell defect for hepatocyte antigens.

A radioimmunoassay for detection of circulating antibodies reacting with the hepatic asialoglycoprotein receptor protein

To determine whether circulating antibodies against the liver-specific membrane lipoprotein complex (LSP), which occur in a number of liver disorders, are directed at the hepatic asialoglycoprotein receptor protein (hepatic lectin)--recently shown to be a constituent of LSP--a radioimmunoassay for anti-hepatic lectin antibodies was developed based on the use of protein A in situ on staphylococcal cells to precipitate 125I-labelled rabbit hepatic lectin/anti-lectin immune complexes. The assay was used to test sera from 30 patients (15 anti-LSP-positive and 15 anti-LSP-negative) with autoimmune chronic active hepatitis (CAH), hepatitis B virus-positive CAH or primary biliary cirrhosis. Anti-lectin antibodies (at titres of 1:200 to 1:1600) were found in all 15 anti-LSP-positive sera and were not detected in the 15 anti-LSP-negative patients. Hepatic lectin is a highly purifiable, liver-specific, hepatocellular surface component and the anti-lectin assay (which proved sensitive, reliable and easy to perform) will permit further exploration of the role of autoimmune mechanisms in the pathogenesis of various liver diseases.

T-CELL INDUCERS OF SUPPRESSOR LYMPHOCYTES CONTROL LIVER-DIRECTED AUTOREACTIVITY

The sensitisation of helper T cells of patients with autoimmune chronic active hepatitis to a liver-cell membrane-expressed asialoglycoprotein receptor protein is shown to be associated with a defect of T cells that specifically induce suppressor lymphocytes. These lymphocytes are found in an activated state in the peripheral blood of healthy people and may form part of an immunoregulatory network which actively prevents autoimmunity.