dolichols and Liver-Neoplasms

Interest in ubiquinone (UQ) has increased during recent years, mainly because of its antioxidant function and its use as a dietary supplement. However, our knowledge of the biosynthesis, catabolism, and regulation of this lipid in mammalian tissues is quite limited. UQ exhibits a high rate of turnover in all tissues indicating that cells possess efficient metabolic pathways for handling this compound and controlling its tissue levels. Besides reviewing the generally accepted metabolic pathway, alternative synthetic mechanisms are described. The lack of data concerning catabolism and regulation of this compound is emphasized. Reasons for the rather limited uptake of dietary UQ are discussed and alternative mechanisms for its beneficial effects on organ function are suggested. Since appropriate tissue uptake of dietary UQ probably only occurs in deficient states, the definition of partial UQ deficiency and its consequences is urgently needed. The possibility of raising tissue UQ levels by drug treatment or natural metabolites is raised as a choice of preference for the future.

Topics: Aging; Animals; Antioxidants; Diet; Dolichols; Half-Life; Humans; Lipid Metabolism; Liver; Liver Neoplasms; Mevalonic Acid; Oxidative Stress; Ubiquinone

Labeling of human colon carcinoma cells with [3H]dol, followed by extensive delipidation and removal of dol-P oligosaccharides, showed that dol are bound to cellular proteins with sizes of 5, 10, 27, 75 and > 140 kDa. HPLC purification of proteolytic products of [3H]dol- and [35S]cys-labeled proteins revealed a hydrophobic peak containing both dol and cysteine. The dol/cys-labeled products were clearly separated from GG-cys, and exhibited a hydrophobicity between that of dol-P and dol. In another set of experiments delipidated proteins were treated with methyl iodide, which cleaves thioether bonds. After HPLC purification of released dol-like lipids, these were subjected to mass spectrometry. This demonstrated molecular ions with the same mass as that of dol. Taken together our data provide evidence for the existence of proteins covalently modified with dol.

Topics: Autoradiography; Chromatography, High Pressure Liquid; Colonic Neoplasms; Cysteine; Dolichols; Hepatoblastoma; Humans; Liver Neoplasms; Melanoma; Mevalonic Acid; Neoplasm Proteins; Peptide Fragments; Protein Prenylation; Sulfur Radioisotopes; Tritium; Tumor Cells, Cultured

Certain enzymes of the mevalonate pathway have been investigated in persistent liver nodules induced in the rat by 2-acetylaminofluorene. In these nodules the dolichol level was increased 5-fold, the ubiquinone-9 content elevated 6-fold and the amount of cholesterol unchanged. Microsomal beta-hydroxy-beta-methylglutaryl-coenzyme A reductase activity was greatly increased compared to control liver tissue, which was also the case for the cytosolic farnesyl pyrophosphate synthase. A significant elevation of all-transgeranylgeranyl pyrophosphate synthase activity in the cytosol was also observed. The branch-point enzyme of microsomal dolichol synthesis, i.e. cis-prenyltransferase, was decreased in the nodules; whereas the activity of squalene synthase, the terminal regulating enzyme of cholesterol synthesis, remained unchanged. The dolichol species in nodular tissue were redistributed towards the longer chain length species. One factor regulating the chain length of the polyisoprene products formed in vitro was shown to be the ratio of the concentrations of isopentenyl pyrophosphate:farnesyl pyrophosphate employed. Other regulatory factors in the terminal steps of this biosynthetic pathway appear to determine the amounts and nature of the final isoprenoid compounds formed in vivo. In contrast to the microsomal trans-prenyltransferase activity, which was unchanged, the activity of nonaprenyl-4-hydroxybenzoate transferase, an enzyme participating in ubiquinone synthesis, was greatly elevated. The alterations observed in the activities of enzymes in the mevalonate pathway can at least partially explain the increased levels of dolichol and ubiquinone and the unchanged level of cholesterol found in liver nodules. It is reasonable to propose that this modified mevalonate metabolism will render nodular cells resistant to certain toxic factors and prone to cell proliferation.

Topics: 2-Acetylaminofluorene; Alkyl and Aryl Transferases; Animals; Dimethylallyltranstransferase; Dolichols; Enzyme Induction; Farnesyltranstransferase; Hydroxymethylglutaryl CoA Reductases; Liver; Liver Neoplasms; Male; Mevalonic Acid; Precancerous Conditions; Rats; Rats, Wistar; Transferases

HepG2 cells were cultured in the presence of different concentrations of cyclosporin A (CsA) or Nva2-cyclosporin (Nva2-Cs) for up to 20 days. At a low concentration (2 micrograms/ml) of CsA or Nva2-Cs, the [3H]thymidine incorporation into DNA and the rate of incorporation of [3H]leucine into total protein decreased by 20-25%. Concentrations of 10 micrograms/ml resulted in a 70% reduction of the [3H]thymidine incorporation in comparison with controls. Low concentrations of CsA resulted in mitochondria in the condensed state together with autophagosomes, large vacuoles, and elevated numbers of coated vesicles, as shown by electron microscopy. Low concentrations of Nva2-Cs resulted in swollen mitochondria, increased autophagocytosis, and increased numbers of intermediate filaments and microtubules. Higher doses of these substances (5 micrograms/ml) caused disarrangement of mitochondrial cristae, vesiculation of the endoplasmic reticulum, an elevated number of free polysomes, and accelerated autophagocytosis. Labeling of phospholipids and triglycerides with [3H]glycerol and of cholesterol and dolichol with [3H]acetate was decreased after exposure of HepG2 cells to CsA, or, in particular, Nva2-Cs. Phospholipids secreted from the cells into the medium exhibited an increased level of labeling, but the specific radioactivity of the neutral lipids in the medium was significantly decreased. Treatment of HepG2 cells with either CsA or Nva2-Cs doubled the mitochondrial cytochrome oxidase and carnitine acetyl-transferase, as well as microsomal NADPH-cytochrome c reductase activities. Such treatment also increased the cyanide-insensitive beta-oxidation of fatty acids in peroxisomes, as well as cytoplasmic DT-diaphorase and glutathione transferase activities. Prolonged treatment of the cells with CsA did not result in any cumulative effect. HepG2 cells appear to be suitable for studying the effects of cyclosporins on cellular structure and metabolism and in this system the two drugs studied here exhibited similar effects.

Topics: Carcinoma, Hepatocellular; Carnitine O-Acetyltransferase; Catalase; Cholesterol; Cyclosporine; Cyclosporins; Dihydrolipoamide Dehydrogenase; Dolichols; Electron Transport Complex IV; Fatty Acid Desaturases; Glutathione Transferase; Humans; In Vitro Techniques; Lipid Metabolism; Liver Neoplasms; Microscopy, Electron; NADH Dehydrogenase; Palmitoyl Coenzyme A; Proteins; Time Factors; Tumor Cells, Cultured

Cholesterol, ubiquinone and dolichol biosynthesis from mevalonic acid was measured in non-malignant and malignant cultured human lymphocytes, freshly isolated human mononuclear leucocytes and in cultured human hepatoma cells. The relative flux of mevalonate into ubiquinone, dilichol and cholesterol was not significantly different between malignant and non-malignant cells, although the extent of labelling of each product was an order of magnitude greater in the malignant cultured cells. The most prominent dolichol isolated from total cellular lipid and synthesized in short-term labelling of cultured leukaemic cells had a chain length one isoprene unit shorter than that observed in normal human cells. Cultured human hepatoma cells and mononuclear leucocytes isolated from the peripheral blood of individuals with lymphoblastic and myelogenic leukaemia similarly synthesized shorter-chain dolichols. The dolichols made in cultured non-tumorigenic cells, freshly isolated mononuclear leucocytes from a normal individual or a patient with non-haematological malignancy had normal chain length.

Topics: Carcinoma, Hepatocellular; Cell Line, Transformed; Chromatography, High Pressure Liquid; Dolichols; Humans; Leukemia; Leukocytes, Mononuclear; Liver Neoplasms; Lymphocytes; Mevalonic Acid; Tumor Cells, Cultured

Surgical samples of human hepatic tissue were analysed morphologically and biochemically and highly differentiated hepatomas were compared with two control groups: morphologically normal liver tissue surrounding the tumour, and tissue from normal livers. In tumour homogenates cholesterol levels were more than twice, ubiquinone levels about half and the concentration of free dolichol about 10% of the control value. The levels of dolichyl phosphate were basically similar, whereas the phospholipid level was slightly lower in the tumours. In microsomes isolated from hepatomas, the level of cholesterol was about 30% higher than the control value. HMG-CoA reductase activity in microsomes isolated from hepatomas was elevated almost 100% in comparison to control. In hepatomas, no major alterations in the compositions of dolichol or dolichyl phosphate could be observed. The relative amounts of alpha-saturated and alpha-unsaturated polyprenols were also basically unaltered in hepatomas. Liver samples were incubated with 3H-mevalonic acid and radioactivity was monitored in polyprenols. With control tissue, incorporation was considerably higher in alpha-unsaturated polyprenols than in their alpha-saturated counterparts. In the tumours the rates of incorporation into both polyprenol fractions were much lower, although still higher in the alpha-unsaturated fraction. Labelling of polyisoprenols containing 19 isoprene residues was higher than that of 20 residues. The pattern of labelling in the polyisoprenyl-P fraction was similar. In hepatomas the incorporation into cholesterol and ubiquinone-10 was about 100% higher and 50% lower respectively compared with control tissue. The results in this study of hepatomas indicate that the levels of various lipids may be influenced not only by the regulatory enzyme HMG-CoA reductase, but also by other enzymes catalysing reactions subsequent to this regulatory point. It is also suggested that levels of cholesterol, ubiquinone and dolichol may be regulated independently subsequent to the branch point at farnesylpyrophosphate.

Topics: Carcinoma, Hepatocellular; Cholesterol; Dolichol Phosphates; Dolichols; Humans; Liver Neoplasms; Terpenes; Ubiquinone

The levels of cholesterol, ubiquinone and dolichol and the polyprenol composition of dolichol in human hepatocellular carcinomas (hepatomas) with different degrees of differentiation were analyzed and compared with healthy liver tissue. Dolichols were also analyzed in liver metastases. The total level of cholesterol was increased, while the levels of dolichol and ubiquinone were decreased in all hepatomas, but no correlation between these levels and the degree of differentiation of the hepatomas could be observed. The level of dolichol decreased more in the hepatomas than in the liver metastases. The dolichol fraction from hepatomas with a low degree of differentiation contained higher relative amounts of short polyisoprenols (D17) and slightly lower relative amounts of D21 compared with healthy liver tissue, metastatic liver tumors or hepatomas with a high degree of differentiation. The significance of the lipid values found in the different groups is discussed.

Topics: Adult; Carcinoma, Hepatocellular; Cell Membrane; Cholesterol; Dolichols; Humans; Liver Neoplasms; Middle Aged; Terpenes; Ubiquinone

The lipid composition of human hepatocellular carcinomas was examined. The level of dolichol in the tumours was decreased compared to control tissue, whereas the concentration of dolichyl phosphate did not exhibit any major change. A decrease in the amount of dolichyl ester was also observed. The pattern of individual polyisoprenoids in the free dolichol pool was changed in several carcinomas with a relative increase in the shorter dolichols. The isoprenol composition in the dolichyl ester and phosphate fractions of tumours were basically similar to those of controls. alpha-Unsaturated polyisoprenols are present in control livers at a level of 3% of the total free polyisoprenoid fraction, while this value was increased in the tumours. Similar to dolichol, the amount of ubiquinone was also decreased. The content of cholesterol was increased, while the fatty acid pattern of the dolichyl esters showed minor alterations. These modifications in lipid content indicate different mechanisms for the regulation of dolichol and dolichyl phosphate concentrations. The high levels of sterols in contrast to the low polyisoprenol content suggests interference with the regulation of the mevalonate pathway, which is the common biosynthetic route for cholesterol, ubiquinone and dolichol.

Topics: Adult; Aged; Carcinoma, Hepatocellular; Cholesterol; Dolichol Phosphates; Dolichols; Fatty Acids; Humans; Liver Neoplasms; Middle Aged; Ubiquinone

Homogenates and microsomal fractions prepared from biopsies of highly differentiated human hepatocellular carcinomas were found to contain low levels of dolichol in comparison with control tissue. In contrast, the amount of dolichyl phosphate in tumor homogenates was unchanged and actually increased in the microsomal fraction. The pattern of individual polyisoprenoids, both in the free and the phosphorylated dolichol fractions of hepatomas, did not exhibit any major alterations compared to the control. The rates of incorporation of [3H]mevalonic acid into dolichol and dolichyl phosphate in hepatomas were low. The dolichol monophosphatase activities in microsomal fractions from hepatomas and controls did not show any major differences, whereas the activity of the CTP-dependent dolichol kinase was increased in tumor microsomes. Glycosylation of endogenous dolichyl phosphate and of total protein using certain nucleotide-activated sugars was found to be slightly elevated in microsomal fractions from the tumor itself when compared to the control. The reasons for the differences in the levels of polyisoprenoids in hepatomas and control tissue are discussed.

Topics: Carcinoma, Hepatocellular; Cytidine Triphosphate; Cytosine Nucleotides; Dolichol Phosphates; Dolichols; Glycosylation; Humans; Liver Neoplasms; Mevalonic Acid; Microsomes, Liver; Middle Aged; Phosphoric Monoester Hydrolases; Phosphotransferases; Phosphotransferases (Alcohol Group Acceptor); Polyisoprenyl Phosphates

The lipid compositions of homogenates and microsomal fractions derived from surgical samples of highly differentiated human hepatoma, morphologically normal regions outside the tumours and from normal livers were analysed. A few enzyme activities were also assayed. Hepatoma microsomes demonstrated considerably lowered levels of cytochromes P-450 and b5. Hepatoma homogenates exhibited increased levels of cholesterol, normal amounts of dolichyl-P and slightly lowered levels of total phospholipid. The levels of dolichol, dolichol ester and ubiquinone in hepatoma homogenates were prominently decreased. In tumour microsomes the levels of cholesterol and dolichyl phosphate were increased considerably while the levels of phospholipid and dolichol were lowered. The phospholipid composition of tumour homogenates was roughly similar to that of control tissue. In tumour microsomes the relative amounts of phosphatidylserine and phosphatidylinositol were about 30% decreased, whereas the major phospholipids showed minor increases in amount. The rate and pattern of incorporation of [3H]glycerol into individual phospholipids in liver slices from control and hepatoma tissue did not differ to any larger extent. The fatty acid composition of tumour homogenates exhibited minor differences in comparison to the control with the greatest changes in the sphingomyelin fraction. In hepatoma microsomes the fatty acid compositions of the major phospholipids were altered moderately, with evident decreases in the relative amounts of the long-chain polyunsaturated fatty acids. In hepatoma homogenates the fatty acid composition of dolichol esters differed only slightly from the control pattern. These results indicate that the major disturbance in the lipid metabolism of highly differentiated hepatomas is localized to the mevalonate pathway, thus affecting mainly the levels of cholesterol, dolichol and ubiquinone.

Topics: Carcinoma, Hepatocellular; Cholesterol; Cytochrome b Group; Cytochrome P-450 Enzyme System; Cytochromes b5; Dolichols; Fatty Acids; Humans; Lipids; Liver; Liver Neoplasms; Microsomes, Liver; Middle Aged; Phospholipids; Ubiquinone

Topics: Carcinoma, Hepatocellular; Cholesterol; Diterpenes; Dolichols; Humans; Hydroxymethylglutaryl CoA Reductases; Liver; Liver Neoplasms; Microsomes; Microsomes, Liver; Ubiquinone

Topics: Carcinoma, Hepatocellular; Diterpenes; Dolichol Phosphates; Dolichols; Humans; Liver; Liver Neoplasms; Phosphorylation

Hyperplastic nodules and hepatocarcinomas were produced in rat liver by 2-acetylaminofluorene-containing diet. The homogenates and isolated microsomes were analyzed for the content of lipid intermediates and glycosylation reactions. The dolichol content of hyperplastic nodules increases four times in the homogenate and six times in the microsomes. In developed hepatocarcinoma, the amount of dolichol was doubled. Concerning the distribution pattern of the polyprenols, there is a change in the relative amounts of dolichols with 18 and 19 residues. In contrast to the free alcohol, dolichyl phosphate was greatly decreased in nodules, a finding which might be explained by a decreased dolichol kinase and an increased dolichol monophosphatase activity. The percentage of total phosphorylated dolichol was related to the glycosylating capacity. In microsomes, mitochondria, and homogenate from normal liver and in homogenate from hyperplastic liver nodules, the percentages of dolichyl phosphate were 23, 2, 16, and 4, respectively. At maximal glycosylation in vitro, only part of the total dolichyl phosphate was glycosylated. Dolichol-mediated protein glycosylation exhibited a general decrease in the microsomes from nodules and cancer tissue; it is suggested that the main cause of the decrease is a shortage of the available dolichyl phosphate which is rate limiting and which also contributes to the synthesis of the modified oligosaccharide chain.

Topics: 2-Acetylaminofluorene; Animals; Carbohydrate Metabolism; Dolichol Phosphates; Dolichols; Liver Neoplasms; Male; Phosphoric Monoester Hydrolases; Phosphotransferases; Phosphotransferases (Alcohol Group Acceptor); Polyisoprenyl Phosphates; Proteins; Rats; Rats, Inbred Strains

Topics: Animals; Cell Transformation, Neoplastic; Dolichols; Glucose; In Vitro Techniques; Lipid Metabolism; Liver Neoplasms; Microsomes, Liver; Proteins; Rats