1-oleoyl-2-acetylglycerol and Cell-Transformation--Neoplastic

To discover the relationship between the activation of protein kinase C and the generation of reactive oxygen in the tumor promotion process, we investigated the effects of radical scavengers on diacylglycerol-promoted transformation in BALB/3T3 cells. Superoxide dismutase (SOD) showed inhibitory effects on both 1-oleoyl-2-acetylglycerol (OAG)-promoted and diolein-promoted transformation. Catalase (CT) suppressed the promoting effects of diolein by up to 70%. Mannitol (MT), a hydroxyl radical (.OH) scavenger, inhibited diacylglycerol-promoted transformation dose-dependently. These results suggest that activation of protein kinase C alone is insufficient and that generation of reactive oxygen accompanied by activation of the enzyme is essential to the promotion process in BALB/3T3 cells.

Topics: Animals; Catalase; Cell Transformation, Neoplastic; Cells, Cultured; Diglycerides; Free Radical Scavengers; Free Radicals; Mannitol; Methylcholanthrene; Mice; Oxygen; Superoxide Dismutase

Tumour-promoting phorbol esters and diacylglycerols have a variety of biochemical and biological effects in common in a number of cell culture systems; thus, it has been suggested that diacylglycerols are endogenous functional analogues of phorbol esters. In order to examine more directly the possible relevance of these functional analogues in tumour promotion, we have studied the promoting effect of 1-oleoyl-2-acetyl glycerol (OAG) in the two-stage transformation model with BALB/c 3T3 cells, which is designed to simulate in vivo two-stage carcinogenesis. When BALB/c 3T3 cells were treated once with a low dose of 3-methylcholanthrene (MCA), almost no increase in cell transformation occurred. However, repeated application of OAG after MCA treatment enhanced cell transformation. OAG enhanced transformation to a lesser extent than a potent promoter of in vivo and in vitro carcinogenesis, phorbol-12,13-didecanoate. OAG also continuously inhibited junctional intercellular communication when BALB/c 3T3 cells were treated daily at confluence. These results indicate that diacylglycerols mimic phorbol esters in promoting cell transformation and that the enhancement may be related to an inhibitory effect on intercellular communication.

Topics: Animals; Cell Communication; Cell Transformation, Neoplastic; Cells, Cultured; Diglycerides; Glycerides; Mice; Mice, Inbred BALB C; Time Factors

Alterations in patterns of protein phosphorylation after exposure to phorbol esters were compared in chicken embryo fibroblasts and KD cells (a human fibroblast line) by two-dimensional gel electrophoresis. A substantial increase in phosphorylation was observed of a major, markedly acidic protein of pI = 4.5 in two-dimensional gels of each cell type. However, the apparent molecular weights of these phosphoproteins differed substantially in the two species with a molecular weight of 67,000 in chicken fibroblasts and one of 80,000 in human KD cells. Both phosphoproteins, termed 67K and 80K respectively, contained phosphoserine and a small amount of phosphothreonine, but no detectable level of phosphotyrosine. Tryptic and chymotryptic phosphopeptide maps of 67K were nearly identical to those of 80K. These results indicate that they are related molecules, even though considerably different in apparent molecular weight, and that the induction of phosphorylation of these closely related, major, acidic phosphoproteins by phorbol esters is conserved from avian to human cells. In chicken embryo fibroblasts infected by a temperature-sensitive mutant of Rous sarcoma virus, phosphorylation of 67K was found to be elevated at 36 degrees C (transformed phenotype) compared to 41.5 degrees C (normal). Although the function of these closely related 67K and 80K phosphoproteins is unknown, the elevated level of phosphorylation could be involved in some aspects of transformation, and the increase is mimicked by treatment with phorbol esters.

Topics: Amino Acids; Animals; Avian Sarcoma Viruses; Cell Transformation, Neoplastic; Cells, Cultured; Chick Embryo; Diglycerides; Fibroblasts; Humans; Molecular Weight; Peptide Mapping; Phosphorylation; Protein Kinase C; Proteins; Tetradecanoylphorbol Acetate

Evidence has been accumulated that the phorbol ester receptor in intact cells is protein kinase C. However, it is not certain whether the various actions of 12-O-tetradecanoylphorbol-13-acetate (TPA) on cultured cells are all mediated by the activation of protein kinase C. Therefore, we examined the effects of inhibitors of protein kinase C, palmitoylcarnitine (PC) and phloretin (PH), on several actions of TPA on cells. PC at the concentration of 30 micrograms/ml completely prevented the inhibitory actions of TPA on differentiation of Friend leukemic cells (FLC) induced by hexamethylene bisacetamide (HMBA) and on metabolic cooperation of V79 cells. PC, however, did not prevent the TPA-induced promotion of the transformation of BALB/3T3 cells, even at the concentration of 40 micrograms/ml. On the other hand, PH at the concentration of 30 micrograms/ml did not inhibit the actions of TPA to inhibit differentiation of FLC and metabolic cooperation of V79 cells, but completely inhibited the transformation of BALB/3T3 cells and its promotion by TPA. These results indicate that protein kinase C possibly mediates some of the actions of TPA, such as the inhibition of differentiation and metabolic cooperation, but not that of the promotion of in vitro transformation.

Topics: Animals; Cell Communication; Cell Differentiation; Cell Line; Cell Transformation, Neoplastic; Diglycerides; Mice; Palmitoylcarnitine; Phloretin; Protein Kinase C; Superoxides; Tetradecanoylphorbol Acetate