Interaction of hypoxia and carbon tetrachloride toxicity in hepatocyte monolayers

Abstract

The toxicity of carbon tetrachloride (CCl 4) in monolayer cultures of primary hepatocytes was investigated at oxygen concentrations that prevail in the liver under conditions that range from normoxia to hypoxia: 0.5, 1, 2, and 20% O 2. CCl 4 was administered in the vapor phase at concentrations that produce aqueous concentrations at 37°C of 0.4, 2.0, and 4.0 m M. Damage was assayed by leakage of aspartate transaminase and the inclusion of Trypan Blue immediately after the 2-hr incubation and after an additional 6-hr incubation in 20% O 2. Only in the case of 0.5% O 2 and 4 m M CCl 4 were the monolayers damaged (18%) immediately after the 2-hr exposure; all other exposed cells were undamaged at that time point and the dose response of cell death as a function of CCl 4 and oxygen concentration was not evident until the 6-hr time point. The monolayers exposed to 4 m M CCl 4 and 1, 2, or 20% O 2 exhibited little immediate damage but were all 100% dead 6 hr later. The monolayers exposed to 2 m M CCl 4 and 0.5, 1, 2, or 20% O 2 were 53, 48, 40, and 22 ± 2% dead after 6 hr, respectively. These results suggest that effects of CCl 4 exposure, for example alterations in the function or synthesis of essential proteins, require several hours to affect cell viability.