Hepatitis B polypeptide vaccine preparation in micelle form.

The immunoprophylaxis of hepatitis B is hampered by the lack of a technique for growing hepatitis B virus (HBV) in tissue culture. Plasma from persistently infected individuals, one source of viral antigen, contains characteristic 22-nm spherical particles which share a common antigen (the hepatitis B surface antigen or HBsAg) with the outer envelope of the 42-nm double-shelled DNA virus. Highly purified inactivated 22-nm particles have been shown to be safe and to confer protective immunity against HBV in a recent large-scale clinical trial.

Ultrastructural changes in the liver in experimental non-A, non-B hepatitis.

Factor VIII, a blood-clotting derivative prepared from pooled human plasma, previously shown to cause a short-incubation-period non-A, non-B hepatitis in patients and in chimpanzees, was studied further to establish the nature of the pathological lesions associated with this infection. Percutaneous liver biopsy specimens were examined in detail. Cytoplasmic changes in the hepatocytes were observed on the 7th day after inoculation in one instance and after 13 days in the second, and persisted in the biopsy specimens for 13 weeks after infection.

The use of markers in immune electron microscopy.

Immune electron microscopy (IEM) cannot be used successfully for structures that do not have recognisable morphology. However, at least some of these structures or components are related antigenically to recognisable antigens or viruses. We have therefore mixed unknown antigens with known markers and looked for the presence of mixed aggregates.

Specificity of an immunoprecipitin test for non-A non-B hepatitis.

An immunodiffusion system detecting an antigen showing immunological identity with international laboratory reference sera was developed by use of acute-phase and recovery sera from patients with transmission-proven non-A, non-B acute hepatitis. In other liver diseases the antigen was also present in a high proportion of patients and there were similar findings in conditions with high levels of circulating immune complexes in the serum. Fractionation of antigen-containing sera by column chromatography, polyethylene glycol treatment, and reduction and alkylation also suggested that immune complexes may be responsible for antigenic activity.

Induction of cell-mediated immunity to HBsAg polypeptides.

Adult male guinea pigs were immunized with hepatitis B virus polypeptides prepared by Triton X-100 solubilization of purified 22-nm hepatitis B surface antigen (HBsAg) particles. A virus-specific subunit containing both the 28,000 molecular weight glycoprotein and the 23,000 molecular weight protein stimulated both cell-mediated and humoral immunity. A whole-blood-cell transformation assay additionally showed that the 64,000 molecular weight component of HBsAg, previously shown to contain host-specific antigens, also stimulated a cellular response to purified intact HBsAg particles, suggesting the additional presence of virus-specific material in this fraction.

Nondetection of infectious hepatitis B virus in a human hepatoma cell line producing hepatitis B surface antigen.

The PLC/PRF/5 human hepatoma cell line producing hepatitis B surface antigen (HBsAg) was studied to determine whether infectious hepatitis B virus (HBV) was also being produced. 2 chimpanzees with no previous exposure to HBV and no serologic markers of past or active HBV infection were inoculated intravenously with 50 ml of either tissue culture supernatant fluid (357 ng/ml HBsAg) or a suspension of cells disrupted by repeated freeze-thaw cycles (57 ng/ml HBsAg). No evidence of HBV infection was detected in either chimpanzee during 6 months of evaluation.

Detection of HBsAg in a clone derived from the PLC/PRF/5 human hepatoma cell line.

A total of 28 clones were established from the PLC/PRF/5 hepatoma cell line by a plating procedure. All clones were found to secrete HBsAg into the supernatant culture fluids. Of these, one clone (No.

Establishment of a human hepatocellular carcinoma cell line releasing hepatitis B virus surface antigen.

A new hepatocellular carcinoma cell line, DELSH-5, derived from operative wedge biospy from a HBsAg sero- and tissue-positive patient, has been continuously propagated in vitro for nearly 22 months. The cells not only resemble hepatocytes on light and electron microscopic examination but also possess biosynthetic markers of the latter such as albumin and alpha-foetoprotein which were demonstrated in the supernatant medium as well as in the tumour cell cytoplasm. Karyology of cloned cells shows moderate aneuploidy, the major model chromosome number being 61.

Non-A/non-B hepatitis in experimentally infected chimpanzees: cross-challenge and electron microscopic studies.

Inoculation of eight chimpanzees with factor VIII, factor IX, or "H" strain plasma resulted in enzymatic and histopathologic evidence of non-A/non-B hepatitis in all eight animals. Challenge of two chimpanzees convalescent from factor VIII-induced disease with either factor IX or "H" strain plasma resulted in non-A/non-B hepatitis only in the animal inoculated with factor IX materials. Reciprocal cross-challenge of a chimpanzee convalescent from factor IX-induced disease with factor VIII also produced unequivocal enzymatic and histopathologic evidence of non-A/non-B hepatitis.

Further evidence of cellular changes associated with non-A, non-B hepatitis.

Two distinct types of ultrastructural changes were found in the hepatocytes of chimpanzees infected with two forms of non-A, non-B hepatitis. In the type of infection that was of long incubation, there was a marked cytoplasmic derangement of the endoplasmic reticulum, with the formation of tubules, but no pathological changes in the nuclei. In the short-incubation type of non-A, non-B hepatitis, induced experimentally in a chimpanzee that had recovered from the long-incubation type of infection, nuclear alterations were found together with the presence of aggregates of particles measuring 15--27 nm in diameter.

Hepatitis B surface antigen production as a growth cycle-related terminal event in PLC/PRF/5 hepatoma cells.

The dynamics of hepatitis B surface antigen production of PLC/PRF/5 hepatoma cells were studied using a quantitative radioimmunoassay method. Maximum rates of antigen production were found in nutrient-depleted, non-dividing cultures and were temporally related to cytological changes preceding cell death. These results indicate that antigen production may be cell-cycle related and represent a terminal event.

De novo acute infection and reactivation of hepatitis B virus in established cirrhosis.

Five patients with cirrhosis proved by biopsy had clinical, biochemical, and serological evidence of an acute hepatitis B infection. In two the illness was fulminant and led to death. Only one patient completely recovered.

Analysis of hepatitis B surface antigen components solubilized with Triton X-100.

Three glycoproteins of intact hepatitis B surface antigen (HBsAg) with mol. wt. of 32 000, 30 000 and 28 000 respectively were identified by reaction with 125I-concanavalin A (Con A) after separation by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE).