ABSTRACT
Fatty acid synthetase (FAS) is overexpressed in various tumor tissues, and its inhibition and/or malonyl-CoA accumulation have been correlated to apoptosis of tumor cells. It is widely recognized that both omega-3 and omega-6 polyunsaturated fatty acids (PUFAs) depress FAS expression in liver, although epidemiological and experimental reports attribute antitumor properties only to omega-3 PUFA. Therefore, we investigated whether lipogenic gene expression in tumor cells is differently regulated by omega-6 and omega-3 PUFAs. Morris hepatoma 3924A cells were implanted subcutaneously in the hind legs of ACI/T rats preconditioned with high-lipid diets enriched with linoleic acid or alpha-linolenic acid. Both-high lipid diets depressed the expression of FAS and acetyl-CoA carboxylase in tumor tissue, this effect correlating with a decrease in the mRNA level of their common sterol regulatory element binding protein-1 transcription factor. Hepatoma cells grown in rats on either diet did not accumulate malonyl-CoA. Apoptosis of hepatoma cells was induced by the alpha-linolenic acid-enriched diet but not by the linoleic acid-enriched diet. Therefore, in this experimental model, apoptosis is apparently independent of the inhibition of fatty acid synthesis and of malonyl-CoA cytotoxicity. Conversely, it was observed that apoptosis induced by the alpha-linolenic acid-enriched diet correlated with a decrease in arachidonate content in hepatoma cells and decreased cyclooxygenase-2 expression.
Subject(s)
Apoptosis/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Isoenzymes/genetics , Liver Neoplasms, Experimental/pathology , Prostaglandin-Endoperoxide Synthases/genetics , alpha-Linolenic Acid/administration & dosage , Acetyl-CoA Carboxylase/genetics , Acetyl-CoA Carboxylase/metabolism , Animals , Apoptosis/genetics , CCAAT-Enhancer-Binding Proteins/genetics , Cyclooxygenase 2 , Diet , Dietary Fats, Unsaturated , Down-Regulation/genetics , Fatty Acid Synthases/genetics , Fatty Acid Synthases/metabolism , Fatty Acids, Omega-6/genetics , Fatty Acids, Omega-6/metabolism , Fatty Acids, Unsaturated/genetics , Fatty Acids, Unsaturated/metabolism , Isoenzymes/biosynthesis , Liver Neoplasms, Experimental/drug therapy , Liver Neoplasms, Experimental/genetics , Liver Neoplasms, Experimental/metabolism , Prostaglandin-Endoperoxide Synthases/biosynthesis , Rats , alpha-Linolenic Acid/metabolismABSTRACT
The hindlimb-suspended rat was used as animal model to investigate the effects induced by immobilization of the skeletal muscle in the expression of the genes encoding hepatic lipogenic enzymes. Following a 14-day period of immobilization, rats were injected intraperitoneally with radioactive acetate, and the labeling of hepatic lipids and cholesterol was evaluated 15 min after the isotope injection. The incorporation of labeled acetate in lipids and cholesterol was almost three times higher in the liver of immobilized rats than in control animals as a consequence of the enhanced transcription of the genes encoding acetyl-CoA synthase, acetyl-CoA carboxylase, fatty acid synthase, and 3-hydroxy-3-methylglutaryl-CoA reductase. The high expression of the key enzymes for fatty acid and cholesterol synthesis induced by immobilization was not paralleled by an increase of the hepatic sterol-regulatory element binding protein (SREBP)-1 and SREBP-2 mRNA content. However, the expression of the mature form of SREBP-1 and SREBP-2 was higher in the nuclear fraction of immobilized rat liver than in controls due to a significant increase of the cleavage of the native proteins. Immobilization also affected the expression of proteins involved in lipid degradation. In fact, the hepatic content of peroxisome proliferator-activated receptor-alpha (PPARalpha) mRNA and of PPARalpha target genes encoding carnitine palmitoyl transferase-1 and acyl-CoA oxidase were significantly increased upon immobilization.
Subject(s)
Immobilization/physiology , Lipids/biosynthesis , Liver/enzymology , Acetates/metabolism , Animals , CCAAT-Enhancer-Binding Proteins/analysis , Cholesterol/biosynthesis , DNA-Binding Proteins/analysis , Enzymes/genetics , Lipids/analysis , Liver/chemistry , Male , Models, Animal , RNA, Messenger/analysis , Rats , Rats, Wistar , Sterol Regulatory Element Binding Protein 1 , Transcription Factors/analysisABSTRACT
A method for quantitating glycerophosphorylcholine by flow injection analysis is reported in the present paper. Glycerophosphorylcholine phosphodiesterase and choline oxidase, immobilized on controlled porosity glass beads, are packed in a small reactor inserted in a flow injection manifold. When samples containing glycerophosphorylcholine are injected, glycerophosphorylcholine is hydrolyzed into choline and sn-glycerol-3-phosphate. The free choline produced in this reaction is oxidized to betain and hydrogen peroxide. Hydrogen peroxide is detected amperometrically. Quantitation of glycerophosphorylcholine in samples containing choline and phosphorylcholine is obtained inserting ahead of the reactor a small column packed with a mixed bed ion exchange resin. The time needed for each determination does not exceed one minute. The present method, applied to quantitate glycerophosphorylcholine in samples of seminal plasma, gave results comparable with those obtained using the standard enzymatic-spectrophotometric procedure. An alternative procedure, making use of co-immobilized glycerophosphorylcholine phosphodiesterase and glycerol-3-phosphate oxidase for quantitating glycerophosphorylcholine, glycerophosphorylethanolamine and glycerophosphorylserine is also described.
