Immunocytochemical and electron microscopic study of hepatitis B virus antigen and complete particle production in hepatitis B virus DNA transfected HepG2 cells.
The relationship between the presence of hepatitis B virus antigens, their localization and hepatitis B virus replication was studied in different clones of cultured HepG2 hepatoblastoma cells transfected with cloned hepatitis B virus DNA. Intracellular hepatitis B virus antigens were detected by immunofluorescence. The production of these antigens was evaluated in the culture media by enzyme-linked immunoassay. Hepatitis B virus DNA was detected using dot-blot hybridization. Three types of HBcAg staining were observed in transfected HepG2 cells: (a) cells with nuclear HBcAg, (b) cells with cytoplasmic HBcAg and (c) cells with both nuclear and cytoplasmic HBcAg. Cell types b and c also expressed hepatitis B virus DNA in their culture media. Our results suggest that cytoplasmic HBcAg may be more involved than nuclear HBcAg in hepatitis B virus replication. The site of hepatitis B virus formation in hepatocytes was studied by electron microscopic examination of a specific hepatitis B virus producer clone, thereby allowing detection of intracellular Dane particles more easily than liver biopsy samples from infected patients. Dane particles and HBsAg filaments were found in large, dilated structures probably related to the endoplasmic reticulum. Budding of core particles into cisternae of endoplasmic reticulum-related structures appears to be a possible mechanism for hepatitis B virus formation; our results suggest that the exocytosis of cisternae to extracellular spaces may be a mechanism for release of hepatitis B virus particles.
Department of Cancer Biology, Harvard School of Public Health, Boston, Massachusetts 02115.
P
P
Roingeard
S L
SL
Lu
C
C
Sureau
M
M
Freschlin
B
B
Arbeille
M
M
Essex
J L
JL
Romet-Lemonne
D015139:Blotting, Southern; D003593:Cytoplasm / Q000648:ultrastructure; D004279:DNA, Viral / Q000235:genetics*; D005455:Fluorescent Antibody Technique; D006512:Hepatitis B Core Antigens / Q000235:genetics* / Q000276:immunology; D006514:Hepatitis B Surface Antigens / Q000235:genetics* / Q000276:immunology; D006515:Hepatitis B virus / Q000235:genetics* / Q000254:growth & development / Q000648:ultrastructure; D006801:Humans; D008099:Liver / Q000648:ultrastructure; D008854:Microscopy, Electron; D014162:Transfection; D014407:Tumor Cells, Cultured; D014779:Virus Replication
D004279:DNA, Viral; D006512:Hepatitis B Core Antigens; D006514:Hepatitis B Surface Antigens