Lymphocytes from hepatic inflammatory infiltrate kill rat hepatocytes in primary culture

In the last few years it has become possible in the liver to isolate lymphocytes from inflammatory infiltrates and to culture them in vitro. Most of the lymphocyte clones obtained are CD 8 + cytotoxic cells, but interactions between these lymphocytes and hepatocytes in primary culture have not been analysed previously. In this study, cloned human T lymphocytes from liver biopsies and from the peripheral blood of patients with chronic hepatitis B or primary biliary cirrhosis, after phenotypical and functional characterization into CD 8+ or CD 4+ cytotoxic lymphocytes, were activated in an antigen-independent fashion by adding either anti CD 3 or anti CD 2/R-3 monoclonal antibodies to the cell suspension. The activated cells were then coincubated with rat hepatocytes in primary culture. The killing capacity of the activated lymphocytes was monitored by light and electron microscopy and by measurement of lactic dehydrogenase (LDH)-release into the culture medium. It was found that cytotoxic CD 8 +, but not CD 4 + helper lymphocytes very effectively killed hepatocytes. The killing effect was dependent on the time of cocultivation and on effector-target (E/T) ratio. Total breakdown of the hepatocyte monolayer was achieved after 10–20 h coculture and at an E/T ratio of 10 to 1. As LDH-release in the culture medium reached about 80% of the total LDH-content, most of the hepatocytes were lysed by activated lymphocytes. Cytotoxic activity of clones obtained from different biopsies was comparable with that of clones from peripheral blood. Hepatocytes in primary culture seem to be very sensitive to the killing capacity of activated cytotoxic lymphocytes. Supported by DFG grants Ra 362/5-2 and SFB 311 A7 (G.R.) and A5 (H.P.D.)