1. Methotrexate was administered immediately after partial (70%) hepatectomy, resulting in complete inhibition of dihydrofolate reductase in 24 h-regenerating liver.
2. At 48 h and 72 h after partial hepatectomy, thymidylate synthase activity was increased, whereas thymidine kinase was inhibited, by the injection of methotrexate. The DNA and RNA contents and the liver weight were also reduced in methotrexate-treated rats.
3. The immunoblotting assay showed that methotrexate stimulated the synthesis of thymidylate synthase protein in 48 h-regenerating liver. At the same time, thymidylate synthase activity was directly inhibited by methotrexate. The mechanisms of inhibition of these enzymes by methotrexate appeared to be different.