Putrescine-dependent invasive capacity of rat ascites hepatoma cells.

The effects of inhibitors of polyamine synthesis on the invasive capacity of rat ascites hepatoma (LC-AH) cells were examined by in vitro assay of penetration of the LC-AH cells through a monolayer of calf pulmonary arterial endothelial (CPAE) cells. Pretreatment of LC-AH cells with alpha-difluoromethylornithine (DFMO), an inhibitor of ornithine decarboxylase, before seeding them onto a CPAE cell monolayer and culturing them for 24 h in the absence of DFMO decreased the number of penetrating tumor cells time and dose dependently (about 35% of the maximal inhibition) without affecting their viability or proliferative activity. DFMO treatment caused a marked decrease in the intracellular level of putrescine but not of spermidine or spermine. The DFMO-induced decreases in invasive capacity and putrescine level were almost completely reversed by the addition of putrescine to the medium during pretreatment with DFMO or invasion assay but were not affected by exogenous spermidine or spermine. No change in the invasive capacity was observed when the CPAE cells were treated with DFMO and the LC-AH cells with methylglyoxal-bis(guanylhydrazone), an inhibitor of S-adenosylmethionine decarboxylase, which depressed the spermidine and spermine levels but increased the putrescine level in the LC-AH cells. These results suggest that intracellular putrescine modulates the in vitro invasive capacity of LC-AH cells.     

Chromatin-associated glycoproteins of normal rat liver and Novikoff hepatoma ascites cells.

Glycoproteins were demonstrated in the 0.6 M NaCl extract of chromatin of normal liver and Novikoff hepatoma cells by periodic acid-Schiff staining of sodium dodecyl sulfate-polyacrylamide gels. In chromatin extracts of Novikoff hepatoma cells, six major periodic acid-Schiff-positive bands coincident with Coomassie Blue-stained protein bands were found with molecular weights of 30,000, 54,000, 64,000, 75,000, 104,000, and 127,000. A corresponding extract from normal rat liver chromatin revealed the presence of four major periodic acid-Schiff-positive bands that migrated with protein bands at molecular weights of 16,000, 30,000, 54,000, and 75,000. The glycoproteins of these extracts contained either mannose or alpha-D-glucose, fucose, and N-acetylglucosamine as shown by the specificity of lectins in affinoelectrophoresis. One of the glycoproteins detected by affinoelectrophoresis was very basic.     

Nucleolar phosphoproteins of normal rat liver and Novikoff hepatoma ascites cells.

The nucleolar acid-soluble proteins from normal rat liver and Novikoff hepatoma ascites cells were labeled in vivo for 2 hr after the injection of [32P]orthophosphate and in vitro with [gamma-32P]adenosine triphosphate in systems containing 0.25 M sucrose, 5 mM MgCl2, 12.5 mM NaCl, and 0.05 M Tris-HCl buffer, pH 7.3, AT 37 DEGREES. Two-dimensional polyacrylamide gel electrophoresis and autoradiography showed that approximately 40 and 20 protein spots were labeled in vivo and in vitro, respectively. There were some marked differences in labeling in vivo between normal rat liver and Novikoff hepatoma acid-soluble nucleolar proteins. By 32P analysis of gel slices, the proportion of the total 32P incorporated into protein Spot A1-4 was greater in normal liver, and the proportion of 32P incorporated into some high-molecular-weight protein spots, such C23-24 and C26-27, was greater in the Novikoff hepatoma ascites cells. With the in vitro incubation system, the 32P uptake per mg protein was about twice as high in Novikoff hepatoma nucleolar proteins as in normal rat liver nucleolar proteins but, generally, the same proteins were labeled in both tissues.