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Rats received a choline-deficient diet containing 0.1% (w/w) DL-ethionine (CDE) for 4, 10, 14 or 22 weeks. A separate group was treated for 4 weeks with CDE and then received a normal diet for 4 weeks. The L and M2 isoenzymes of pyruvate kinase were immunocytochemically demonstrated in liver sections. L-PK expression was strongly reduced in the hepatocytes after 4 weeks of treatment and remained low until the end of the study. Withdrawal of CDE after 4 weeks followed by 4 weeks normal diet resulted in a nearly full recovery of L-PK expression as compared to untreated controls. At later stages (10-22 weeks of CDE- treatment) many pseudolobules, preneoplastic foci of altered hepatocytes (FAH) such as combined clear/acidophilic cell foci (CCF/ACF) and mixed/basophilic cell foci (MCF/BCF), and hepatocellular adenomas (HCA) were observed. Pseudolobules showed a slight reduction in L-PK-expression, and were negative for M2-PK. In all clear cell components of CCF/ACF excessively storing glycogen, L-PK-expression was increased compared to both the surrounding parenchyma and hepatocytes of controls. In acidophilic cell components with less pronounced glycogen storage L-PK expression was similar to that of pseudolobules showing a slightly reduced content of this enzyme protein. M2-PK was invariably negative in CCF/ACF. In most MCF glycogen-storing subpopulations expressed L-PK, whereas in all glycogen-poor basophilic populations L-PK protein was strongly reduced. M2-PK was not expressed in most of these MCF. However, in rare MCF the reduction in L-PK expression was combined with a significant expression of M2-PK. In HCA M2-PK underwent a further increase, although to a variable degree, while L-PK remained strongly reduced. Our results show that an isoenzyme shift from L-PK to M2-PK takes place at a late stage of the hepatocarcinogenic process, and that those MCF with a low L-PK expression and a reexpression of M2-PK most probably represent the direct precursor lesions of hepatocellular neoplasms.

carcinogenesis

Pyruvate kinase isoenzyme shift from L-type to M2-type is a late event in hepatocarcinogenesis induced in rats by a choline-deficient/DL- ethionine-supplemented diet