Biochemical microanalysis of pyruvate kinase activity in preneoplastic and neoplastic liver lesions induced in rats by N-nitrosomorpholine

Abstract

Fundamental aberrations in carbohydrate metabolism have been previously demonstrated in focal hepatic lesions emerging early during hepatocarcinogenesis induced in rat liver by limited oral administration (stop model) of N-nitrosomorpholine. Using this experimental approach, we have now investigated quantitatively the activity of pyruvate kinase (PK), a key enzyme of glycolysis and gluconeogenesis, in individual preneoplastic and neoplastic hepatic lesions, particularly in glycogen storage foci, mixed cell foci, basophilic cell foci, hepatocellular adenomas and carcinomas and in a specific type of preneoplastic hepatic lesion designated as an enzymatically hyperactive focus (EHF). The focal lesions were dissected from freeze-dried tissue sections with a laser microdissection device. This permits the excision of very small foci and the measurement of enzyme activities in serial sections of the same focus with different substrate concentrations, thus enabling possible changes in the isoenzyme pattern to be detected. On average, PK activity was increased in glycogen storage foci. Mixed cell foci showed a nearly normal or slightly decreased enzyme activity. However, a pronounced reduction in PK activity was observed in low glycogen basophilic foci and in basophilic hepatic tumors. An exceptionally high PK activity was found in one glycogenotic adenoma. An increased activity was also observed in EHF. The results suggest that a reduction in PK activity is a relatively late event during the sequence of cellular changes leading from glycogenotic foci to hepatocellular carcinomas. A drastic decrease in the enzyme activity occurs only when low glycogen basophilic cell populations develop from the glycogenotic foci in later stages of hepatocarcinogenesis.