Induction of hepatocyte stimulating activity by T3 and appearance of the activity despite inhibition of DNA synthesis by adriamycin

Abstract

A hepatocyte stimulating activity (HSA) has been extracted from rats that had received an injection of a pharmacological dose of T3 20 hours earlier. The injection of HSA from T3-treated rats into different recipient rats that had previously had 40% of their liver removed resulted in a significant increase in hepatic DNA synthesis. The injection of saline or HSA from normal rat liver had little or no effect on hepatic DNA synthesis in recipient rats. HSA from the T3-treated rats also stimulated DNA synthesis in Novikoff hepatoma cells and primary hepatocytes in culture, and in isolated normal rat liver nuclei in a nuclear incorporating system. In further experiments in which the increased DNA synthesis that follows partial hepatectomy was blocked by adriamycin, HSA appeared in these non-regenerating livers. This latter observation had indicated that the development of HSA is not merely an accompaniment of DNA synthesis.

Fig. 1

DNA synthesis at different times after HSA injection. Animals were injected with 25 mg protein containing HSA at zero time. At 10, 14, 22 and 34 hours, animals were injected with 10 μCi [³H] thymidine and killed 2 hours later. Incorporation of [³H] thymidine into hepatocytes was determined as described in “Methods”. Each point on the graph represents the average of two determinations. ●——●, injection of HSA prepared from 70% hepatectomized rats. ○——○, injection of saline.

Fig. 2

Stimulation of DNA synthesis in nuclei isolated from rats injected with HSA. Animals were injected with HSA, T, or saline and nuclei were isolated 20 hours later. [³H] TTP incorporation was determined as described in “Methods”. 2A Injections into normal rats: ●——●, injection of T₃ 200 μg/100 g; x——x, HSA from weanling rats; ○——○, saline. 2B Injections were made into animals that had 40% of their liver removed 4 hours before the injection. ●——●, HSA from T₃,-injected rat; x——x, HSA from weanling rats; ○——○, saline.

Fig. 3

Stimulation of DNA synthesis in isolated nuclei by liver extracts. The assay procedure has been described in “Materials and Methods”. Control value is the average of 21 determinations and the values with extracts added are from 3 determinations each. ■ = Control = Sham operated adult rat extract = 70% partially hepatectomized rat HSA = T₃ injected rat HSA □ = Weanling HSA