Enzymes and cytological studies in rat hepatoma transplants, primary liver tumors, and in liver following azo dye feeding or partial hepatectomy.

Abstract

The study of the intracellular distribution of acid and alkaline phosphatases, ribonucleases, cathepsin, adenosine triphosphatase, and glutamic dehydrogenase enzymes and of the mitochondrial population in liver, liver tumors (both primary and transplantable tumors), regenerating liver, and liver of animals fed azo dyes has shown that: 1. The number of mitochondria per whole liver in rats fed DAB was lower than in the controls. 2. The mitochondrial decrease phenomenon in tumor and “precancerous livers” was apparently not due to the high mitotic activity of these tissues, since the average liver cell of regenerating liver contained the same number of mitochondria as the sham-controls (but lower amount than normal liver). These studies in regenerating liver have stressed the importance of comparing liver of partially hepatectomized rats with sham-controls. 3. The alkaline phosphatase component which is Mg-insensitive was shown to be the one affected in liver neoplasia. The association of high alkaline phosphatase activity (Mg-insensitive) of the nuclear fraction of liver tumors and regenerating liver with the high rate of cell division in these tissues appeared doubtful, since the nuclear fraction isolated from the intestinal mucosa contained no activity, whereas the same fraction prepared from brain cortex exhibited comparatively high activity. 4. Acid phosphatase activity was lower in azo dye-fed rats whether the azo dye was a carcinogen (DAB and 3′-Me-DAB) or not (2-Me-DAB). These results were taken as an indication that a lower acid phosphatase in “precancerous liver” was not characteristic of liver carcinogenesis. Acid phosphatase activity was very low in hepatoma transplants; in liver tumors acid phosphatase activity was the same as in liver. In regenerating liver the acid phosphatase was not changed as compared with sham-controls. 5. Adenosine triphosphatase activity was confirmed to be lower in liver tumors than in liver. Adenosine triphosphatase activity was preferentially localized in the nuclear and microsomal fractions of tumor, whereas, in liver, adenosine triphosphatase activity was mostly concentrated in the mitochondrial fraction. 6. Cathepsin activity was lower in Novikoff hepatoma than in liver and distributed nonpreferentially in all cell fractions isolated by differential centrifugation. The behavior of the cathepsin enzyme was thus different from the other lysosomal enzyme investigated (acid phosphatase). 7. Glutamic dehydrogenase enzyme could not be demonstrated in Novikoff hepatoma transplants or isolated fractions. This finding was discussed in relation to the possible function of this enzyme and examined in view of other enzymatic activities known to be “absent” in Novikoff hepatoma transplants. Glutamic dehydrogenase is the first enzyme known to be “lacking” in liver tumor mitochondria. 8. Analysis of the nitrogen distribution pattern in liver tumor fractions as compared with other tissues supported the concept that such a pattern is a biochemical characteristic of tumors. The similarity between the intracellular distribution of nitrogen and ribonuclease in liver tumors was discussed briefly.