Effect of oxygen concentration on the expression of glutathione S-transferase activity in periportal and perivenous rat hepatocyte cultures

Abstract

Cultures of perivenous (PV) and periportal (PP) hepatocytes could provide suitable in vitro models for studying the zone-specific hepatotoxic potential of xenobiotics. However, it is not known whether cultured PP and PV hepatocytes keep their phenotypes when the microcirculation of the liver changes. This question has been studied by culturing rat hepatocytes at 13 and 4% (v/v) O 2, respectively, mimicking the acinar oxygen gradient. PP and PV adult rat hepatocytes were isolated by digitonin-collagenase in situ perfusion and cultured on plastic Falcon® and gas-permeable Petriperm® dishes in Williams' E medium and kept at 13 and 4% (v/v) O 2, respectively. Cultures at 20% (v/v) O 2 on plastic dishes served as a control. Two types of cultures were studied, namely conventional cultures either unsupplemented or supplemented with 30 m m pyruvate. The activities of glutamine synthetase (GS) and glutathione S-transferase (GST) were measured in freshly isolated PP and PV hepatocytes and all cultures. The heterogeneous expression of GS (PV>PP), observed in freshly isolated hepatocytes, was kept for at least 4 days in culture. Total, Mu and Alpha class GST activities were predominantly expressed in PV freshly isolated cells. However, no beneficial effect could be observed in culture by exposing the cells to their specific in vivo oxygen concentration. The best maintenance of GST PV predominance in culture was observed in Petriperm dishes at 20% (v/v) O 2, as well in pyruvate-supplemented as unsupplemented cultures. PV GST predominance was thus kept in particular when the highest oxygen concentration was used and made available to the cells through the gas-permeable membranes. The results on GS PV predominance support these findings.