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Growth kinetics of diethylnitrosamine-induced, enzyme-deficient "preneoplastic" liver cell populations in vivo and in vitro.

Summary Rats fed 5 mg diethylnitrosamine per kg per day die from hepatocellular carcinoma after 138.0 ± 10.7 days (n = 258). Thirty days after start of feeding, circumscribed areas of deficiencies of glucose 6-phosphatase and adenosine triphosphatase appear in the enzyme histochemical preparation of the liver. The number of enzyme-deficient areas increases approximately exponentially until the 70th day, after which it remains stationary. By grouping the enzyme-deficient areas of the liver sections in 54 classes according to size as measured by planimetry (increment per class, 0.0584 sq mm) as a function of time, it can be seen that only areas of Class 1 to 8 (0.0584 to 0.467 sq mm) are present until the 80th day. Thereafter the number of larger areas up to Class 54 (3.154 sq mm) increases discontinually. In autoradiograms prepared after seven injections of thymidine-3H at 6-hr intervals, at early stages the labeling index of hepatocytes of enzyme-deficient areas is not significantly different from the surrounding parenchyma. From the 90th day onward, however, it is greatly increased in individual enzyme-deficient areas, while others maintain a low labeling index. Hepatocytes from nodular enzyme-deficient portions of liver tissue of diethylnitrosamine-fed rats proliferate in vitro, in contrast to those of normal adult liver of control rats. The thymidine-3H labeling index of these cultured hepatocytes 30 min after addition of thymidine-3H differs by a factor of about 25 between individual liver explants of diethylnitrosamine-fed rats under standardized culture conditions. Foci of enhanced proliferation are seen within slowly proliferating hepatocytes. From the varying behavior of the enzyme-deficient cell populations as demonstrated in vivo and in vitro, it is concluded that they are not a homogeneous preneoplastic population but rather potentially preneoplastic cells in various stages of development. Rapidly proliferating subpopulations of the enzyme-deficient cell groups might be foci of origin of hepatocellular carcinomas.