Biochemical functions of adult rat hepatocytes in primary culture.

Liver parenchymal cells from adult rats were isolated by treatment with collagenase and cultured as monolayers in Williams medium E with 10% fetal or calf serum. The additions of dexamethasone and insulin to the medium were essential for maintaining liver functions of the cells. These cells synthesized and secreted various serum proteins into the medium. Gluconeogenesis and glycogenolysis were enhanced by glucagon, and lipogenesis was stimulated by insulin. Many enzymes were also induced by various hormones. These activities were very low in freshly isolated cells, but were restored when the cells were cultured for a few days. Markers of plasma membranes, such as 5'-nucleotidase and insulin receptors, were reduced to half the normal levels on freshly isolated cells, but they were restored to the normal levels during culture of the cells without added hormones. Analysis of the profile of amino acids in the medium showed that freshly isolated cells were in a catabolic state of protein turnover and released branched chain amino acids into the medium, but that cultured cells consumed amino acids, not only for protein synthesis, but also for other metabolic processes, such as gluconeogenesis. These findings show that freshly isolated cells have impaired functions and are unsuitable for use in studies of liver metabolism, but that after culture for a few days the cells regain the activity of normal liver and hance become useful for studies of liver functions. Studies with these cells are simpler and give clear results than studies in vivo.