Duck cultured hepatocytes from Pekin ducks naturally infected by duck hepatitis B virus can remain functional twice longer if a coculture system with rat liver epithelial cells is used instead of ordinary primary culture. The use of a selective medium in which ornithine and lactate replaced arginine and glucose, respectively, allowed viral replication initiated in vivo to be maintained in the coculture for 2 months.
Several antiviral compounds including the pyrophosphate analog (phosphonoformic acid) or nucleoside analogs (9β-arabinofuranosyl AMP, 1-(2′-deoxy-2′-fluoro-β-D-arabinofuranosyl)-5-iodocytosine, 1,2′-deoxy-2′-fluoro-β-D-arabinofuranosyl-5-ethyluracil and 1,2′-deoxy-2′-fluoro-β-D-arabinofuranosyl thymine were studied in both culture systems for their ability to inhibit duck hepatitis B virus replication. Hepatocytes were treated for 7 days with 1,2′-deoxy-2′-fluoro-β-D-arabinofuranosyl-5-ethyluracil (10 μM) and 1,2′-deoxy-2′-fluoro-β-D-arabinofuranosyl thymine (0.5 μM) or for 14 days with 9β-arabinofuranosyl AMP (90 μM), phosphonoformic acid (100 μM) and 1-(2′-deoxy-2′-fluoro-β-D-arabinofuranosyl)-5-iodocytosine (6 μM). The effects of the drugs on viral replication were monitored by testing for duck hepatitis B virus DNA in the culture supernatant and in the cells by molecular hybridization. All the above-mentioned drugs demonstrated an inhibitory activity in both types of cultures which at the quite distinct doses used was greater for phosphonoformic acid and 1-(2′-deoxy-2′-fluoro-β-D-arabinofuranosyl)-5-iodocytosine than for 9β-arabinofuranosyl AMP, 1,2′-deoxy-2′-fluoro-β-D-arabinofuranosyl-5-ethyluracil or 1,2′-deoxy-2′-fluoro-β-D-arabinofuranosyl thymine. Viral replication, however, resumed following discontinuation of treatment.
More studies are needed to further confirm the relevance of this tissue culture system for the screening of new potential anti-hepatitis B virus agents.