Long-term maintenance of functional rat hepatocytes in primary culture by additions of pyruvate and various hormones

Mature adult rat hepatocytes were cultured as monolayers in serum-free Williams medium E containing 10(-7) M each of insulin (Ins), dexamethasone (Dex) and triiodothyronine (T3) and 30 mM pyruvate. The hepatocytes remained morphologically intact for at least 14 days, during which period they maintained normal liver functions such as the expressions of cytochrome P-450 mRNA and glucokinase and secretion of albumin. They also retained the ability to resume proliferation. Cells cultured with pyruvate had a much higher ATP level than those without pyruvate, suggesting that pyruvate can sustain functional hepatocytes for a long period in culture in the presence of Ins, Dex and T3, probably by producing enough energy for their maintenance.