A prospective clinical trial of D-penicillamine in the treatment of primary biliary cirrhosis.

We conducted a prospective clinical trial to assess the relative efficacy and safety of high- vs. low-dose D-penicillamine in patients with primary biliary cirrhosis. Following clinical tests and liver biopsy diagnostic of primary biliary cirrhosis, 56 patients were randomized to receive either 250 or 750 mg D-penicillamine daily. Patients were monitored with clinical tests and annual liver biopsy. Randomization produced two groups without differences in demographic, clinical or histologic characteristics. During the trial, no differences were seen between the mean change in liver test results in patients in either treatment group. The 11% per year rise of bilirubin in the 750 mg dose group during the first 3 years was not significantly different from the 18% per year rise in the 250 mg dose group. No patient showed improvement on liver biopsy although patients on 750 mg D-penicillamine deteriorated more slowly. Side effects, particularly rash and dysgeusia, were more common in the 750 mg dose group. The frequency and severity of side effects were responsible for the early conclusion of our trial. Twenty-six patients experienced side effects necessitating discontinuation of D-penicillamine. No evidence of increased efficacy was demonstrated by high-dose D-penicillamine therapy, and side effects were observed in patients on 250 mg D-penicillamine daily. With the severity of adverse effects and continued progression of disease, D-penicillamine is not a clinically useful therapy in primary biliary cirrhosis.

Detection of two forms of HBeAg (free-and IgG-bound HBeAg) in patients with HBe antigenemia using staphylococcus bearing protein A.

Protein A-bearing Staphylococcus aureus organisms (STA) were used to separate free HBeAg from IgG-bound HBeAg. Free HBeAg was detected in the supernate while IgG-bound HBeAg could be liberated from the pellets using MgCl2 or a glycine buffer. HBeAg was determined by radioimmunoassay and the results expressed as patient's cpm/normal control's cpm ratio (S/N ratio). This ratio was demonstrated to be proportionate to the antigen concentration and used as a titer of HBeAg. Sera of 40 HBsAg-negative healthy volunteers and 82 HBeAg-positive patients who were either asymptomatic HBsAg carriers or had various diseases including chronic persistent hepatitis (CPH), chronic active hepatitis (CAH), and renal disease undergoing hemodialysis, were tested for free HBeAg and IgG-bound HBeAg. Patients with CAH from two different countries were compared. Free HBeAg was detected in all patients but one, IgG-bound HBeAg was detected with similar prevalences (from 56% to 67%) in HBsAg asymptomatic carriers, hemodialysis patients, CPH and Italian CAH patients. In contrast, all CAH patients from New York, who had frequently been exposed to HBV infection, had detectable levels of IgG-bound HBeAg, with the highest S/N ratios observed in the study, and frequently showed an unfavorable outcome. In AVH due to HBV and delta agent co-infection, IgG-bound HBeAg was detected in two of four patients only in the initial stage of the disease. The data reported indicate that a separate determination of free- and IgG-bound HBeAg may have clinical value.

Randomized controlled trial of quinacrine for the treatment of HBsAg-positive chronic hepatitis.

Several drugs which react with DNA decrease hepatitis B viral (HBV) DNA polymerase activity in vitro. Because such an alteration of viral replication, if produced in patients with hepatitis B surface antigen (HBsAg)-positive chronic hepatitis, may lead to elimination of viral infection, we conducted a controlled trial of the use of the intercalating agent, quinacrine hydrochloride, in treatment of HBsAg-positive chronic hepatitis. No patient converted from HBsAg positive to negative during the trial and no consistent effect on HBV DNA polymerase activity was noted. Following treatment, elevated transaminase values and alterations of HBV markers were observed in several patients. Fluctuations of transaminase values and HBV markers may reflect alterations in host immunity and viral replication. Quinacrine alone is ineffective in therapy of chronic HBV infection. Additional study with intercalating agents, perhaps in conjunction with other drugs, is suggested.

Circulating complement fixing immune complexes in chronic hepatitis. Use of anti-C3 enzyme immunoassay to define antibody class and nature of antigen.

An anti-C3 enzyme immunoassay (anti-C3 EIA) was used to identify the antigen and antibody of immune complexes (IC). HBsAg, liver cell membrane antigens (LSP and LP-2), IgA, IgG and IgM were determined in IC of 258 patients with chronic active hepatitis (CAH) and 31 patients with chronic persistent hepatitis (CPH). IC that contained an antibody of either IgG, IgM or IgA class were detected in all types of CAH and CPH investigated. IC were seen more frequently in patients with cirrhosis than in CAH patients without cirrhosis and patients with CPH. LSP and LP-2 in IC were detected in 50% of patients with CAH and, infrequently, in CPH patients. In a group of 56 patients with CAH, followed for two to nine years, those patients treated with steroids showed a tendency to clear IC, whereas no variation was seen in untreated patients. While the presence of IC seems to indicate a decreased clearance of IC, a damaging activity of IC on the liver or the immune system remains to be demonstrated.

In vitro synthesis of IgG and IgM in patients with HBsAg-positive and HBsAg-negative chronic active hepatitis.

Spontaneous and PWM-driven IgG and IgM synthesis was investigated in the PBMC of 15 patients with HBsAg-negative CAH and six HBsAg-positive patients with CAH. PBMC from patients with HBsAg-positive CAH show an impaired IgG synthesis upon stimulation with PWM but an IgM increase similar to that of control subjects. In contrast, PBMC from HBsAg-negative patients with CAH show a trend to spontaneous increased synthesis of IgG and a decrease or lack of IgG and IgM synthesis upon PWM stimulation. Steroid treatment seems to ameliorate these alterations. No differences were found among the three groups of HBsAg-negative chronic active hepatitis (autoimmune, HBsAg-related and cryptogenic). These results indicate that differences of B-cell functions may exist in the two groups of CAH patients, not only in spontaneous B-cell activation or PWM-induced Ig synthesis but also in the different classes of Ig.

Geographic differences in HBV related, autoimmune, and cryptogenic chronic active hepatitis.

To clarify the importance of ethnic and geographic factors in chronic active hepatitis (CAH), HBV markers and autoantibodies (AMA, ANA, SMA), have been compared in 158 patients with biopsy-proven CAH from New York City and in 92 patients with CAH from Milan. HBsAg-positive CAH was more frequently observed in Milan (49%) than in New York City (27%). However, among HBsAg-positive patients, HBcAg, HBeAg, and epidemiologic risk factors for acquisition of HBV infection were more frequently found in New York than in Italy. The prevalence of HBsAg-negative, anti-HBc-positive CAH and cryptogenic CAH was similar in the two cities, while autoimmune CAH was more frequently observed in New York (20%) than in Milan (2%). In particular, the prevalence of autoimmune hepatitis was higher among Jewish patients than among patients of Anglo-Saxon or Latin ethnic background in New York. Thus, environmental and/or ethnic factors may influence the prevalence of the four major types of CAH.

Radioimmunoassay for hepatitis B core antigen.

Serum hepatitis B core antigen (HBcAg) is an important marker of hepatitis B virus replication. We describe an easy, sensitive radioimmunoassay for determination of HBcAg in detergent-treated serum pellets containing Dane particles. Components of a commercial kits for anti-core determination are used, and HBcAG is measured by competitive inhibition of binding of 125I-labeled antibodies to HBcAg with HBcAg-coated beads. We assayed of HBcAG in the sera of 49 patients with hepatitis B surface antigen (HBsAg)-positive chronic hepatitis, 50 patients with HBsAg-negative chronic hepatitis, and 30 healthy volunteers. HBcAg was detected in 41% of patients with HBsAg-positive chronic hepatitis but not in patients with HBsAg-negative chronic hepatitis. Hepatitis Be antigen (an antigen closely associated with the core of Dane particles) determined in the same sera by radioimmunoassay, was not detected in 50% of HBcAg-positive sera.

Cell-mediated immunity to HBcAg and HBsAg in patients with chronic hepatitis.

The leukocyte adherence technique (LAT) has been utilized to assess cell-mediated immunity (CMI) to HBcAg and HBsAg in patients with chronic (CH) and acute viral (AVH) hepatitis. All patients with AVH type B and 91.6% of patients with HBV-related CH displayed reactivity to both HBcAg and HBsAg, whereas healthy controls and patients with liver disease not related to HBV failed to show reactivity to these antigens. Four of 6 laboratory workers who had been exposed to HBsAg and had no signs of hepatitis and 2 of 17 patients with CH unrelated to HBV who received multiple transfusions exhibited reactivity to HBsAg, while reactivity to HBcAg was seen only in 2 laboratory workers and in 1 patient with CH unrelated to HBV. These results suggest that according to the LAT, reactivity to HBcAg is present in patients with AVH and CH and may be related to the etiology of the disease, whereas reactivity to HBsAg alone indicates previous exposure to HBV.

A radioisotopic leukocyte adherence test.

A modification of the leukocyte adherence inhibition method is described which utilizes 51Cr-labeled blood mononuclear cells placed in microwells. The test is reproducible, objective, employs approximately 2,000 cells per well, and allows multiple replicates of several antigens. With the two antigens tested, SK-SD and PPD, both increases and decreases of leukocyte adherence are observed. Serum proteins are important in the medium and inactivated AB serum gave the best results. A good correlation was observed between this leukocyte adherence test and positivity of skin testing,

Dane particles-associated hepatitis B core antigen in patients with HBsAg-positive chronic hepatitis.

Dane particles-associated hepatitis B core antigen (HBcAg) was determined by radioimmunoassay in 61 patients with hepatitis B surface antigen (HGsAg)-positive chronic hepatitis. HBc antigenemia was observed in 61% of patients, especially in those with epidemiological risk factors. Patients with chronic active hepatitis as well as those with chronic persistent hepatitis may have HBc antigenemia. The highest levels of HBcAg were observed in male homosexuals. Follow-up determinations indicate the general tendency of HBcAg to decrease or disappear. HBcAg-positive patients with chronic active hepatitis had a poor prognosis, whereas HBcAg-negative patients frequently had a favorable clinical course of the disease (P less than 0.001). The assay of HBcAg in the serum of patients with HBsAg-positive chronic active hepatitis is a useful parameter with both clinical and epidemiological importance.

Immune response to hepatitis B virus in children with papular acrodermatitis.

Papular acrodermatitis of childhood (PAC) is characterized by papular eruption of skin, lymphadenopathy, and acute hepatitis B surface antigen (HBsAg)-positive anicteric hepatitis. To study the course of hepatitis B virus infection we followed 16 patients with PAC, 2 to 7 years of age, for periods ranging from 6 to 46 months. All patients tested developed hepatitis B surface antigenemia subtype ay, and produced antibody to hepatitis B core antigen with the highest incidence after 3 to 5 months. Half of the children investigated developed antibody to hepatitis B surface antigen 4 to 18 months (mean, 6.5) after the onset of PAC. At the end of the investigation, 31% of the children were still HBsAg-positive, 50% were antibody to hepatitis B core antigen-positive, and in 43% the activity of serum aminotransferases was abnormal. Liver biopsy repeated in 2 children showed chronic aggressive hepatitis. The pattern of antibody response to hepatitis B virus is similar in both HBsAg-positive hepatitis and PAC. The frequent development of HBSAg carrier state and the high proportion of children with liver abnormalities at the end of the investigation suggest an impaired clearance of hepatitis B virus and a tendency to chronicity.

T and B lymphocytes in patients with chronic active hepatitis (CAH).

Absolute numbers of T and B lymphocytes as well as active E rosette-forming cells were measured in twenty-seven patients with chronic active hepatitis (CAH), and in thirty control patients. In patients with CAH without cirrhosis, active E rosette-forming cells (a subpopulation of T lymphocytes considered to be actively involved in cell-mediated immune reactions) as well as lymphocytes with surface markers for IgA, IgM and IgG were increased. In patients with CAH and cirrhosis, total T lymphocytes were decreased. These results emphasize the significance of lymphocytes in CAH, and suggest the importance of monitoring T- and B-cell populations in patients with this disease.

Antibodies to hepatitis B core antigen in hepatitis B surface antigen-positive and -negative chronic hepatitis.

Antibody to hepatitis B core antigen (anti-HBc) is a sensitive indicator of hepatitis B virus (HBV) infection. However, anti-HBc has been found in only a few patients with chronic hepatitis. Therefore, we tested for anti-HBc in 124 sera from 67 patients with histologically proven chronic hepatitis by the indirect fluorescent antibody technique. All patients, except for one with chronic hepatitis who was seropositive for hepatitis B surface antigen (HBs Ag), had anti-HBc that persisted throughout the follow-up period (three months to three years). Of 33 HBs Ag-seronegative patients, anti-HBc was detected in seven patients and persisted for six months to two years. These findings suggest that in this study 21% of patients with chronic hepatitis with undetectable amounts of HBs Ag in the serum had evidence of recent or continued HBV replication.

Extrahepatic malignancy in chronic liver disease: report of six cases.

Six patients are described with chronic liver disease, 3 with primary biliary cirrhosis and 3 with chronic active hepatitis, all of whom developed malignant disease in organs other than the liver. Two malignancies were mixed histiocytic-lymphocytic lymphomas. All patients had received corticosteroid or azathioprine therapy. Specific lymphocyte counts and immunoglobulin and antibody determinations failed to reveal a consistent pattern of abnormality to recognize the patient at risk. Several diseases in which the immune responsiveness is disturbed seem to have this potentiality in common, especially if therapy that further alters immunological reactivity is given.