APOPTOSIS INDUCED BY TUMOR-NECROSIS-FACTOR-ALPHA IN RAT HEPATOCYTE CELL-LINES EXPRESSING HEPATITIS-B VIRUS

Three well differentiated SV40-immortalized rat hepatocyte cell lines, CWSV1, CWSV2, and CWSV14, and Hepatitis B Virus (HBV)-producing cell lines derived from them were examined for sensitivity to tumor necrosi s factor (TNF)-alpha. CWSV1, CWSV2, and CWSV14 cells were co-transfect ed with a DNA construct containing a dimer Of the HBV genome and the n eo gene and selected in G418 to generate stable cell lilies. Character ization of these cell lines indicated that they contain integrated I I BV DNA, contain low molecular weight HBV DNA compatible with the prese nce of HBV replication intermediates, express HBV transcripts, and pro duce HBV proteins The viability of CWSV1, CWSV2, and CWSV14 cells was not significantly altered when they were treated with TNF-alpha at con centrations as high as 20,000 U/ml. libe HBV-expressing CWSV1 cell lin e, SV1di36, and the HBV-expressing CWSV14 cell line, SV14di208, were a lso not killed when treated with TNF-alpha. However, the HBV-expressin g CWSV2 cell line, SV2di366, was extensively killed when treated with TNF-alpha. at concentrations ranging from 200 to 20,000 U/ml. Analysis of several different HBV-producing CWSV2 cell lines indicated that TN F-alpha killing depended upon the level of HBV expression. The TNF-alp ha-induced cell killing in high HBV-producing CWSV2 cell lines was acc ompanied by the presence of an oligonucleosomal DNA ladder characteris tic of apoptosis.