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A C Rustan

Publications and source records attributed to A C Rustan.

At least 37 records · Page 2Linked to original sources
PubMed1994

Effect of chronic incubation of CaCo-2 cells with eicosapentaenoic acid (20:5, n-3) and oleic acid (18:1, n-9) on triacylglycerol production.

CaCo-2 monolayers, cultured for 1 week after reaching confluence, were incubated with micellar solutions of fatty acids for up to 7 days. These conditioned cells were incubated acutely (5 h) with eicosapentaenoic acid and oleic acid, and the levels of cell-associated and secreted triacylglycerol were determined. With acute addition of oleic acid, both cell-associated and secreted triacylglycerol were decreased in cells chronically exposed to eicosapentaenoic acid. This effect was observed after as little as 2 days of chronic incubation with eicosapentaenoic acid. A further decrease was found when these cells were incubated acutely with eicosapentaenoic acid, regardless of which radioisotopes were used to label precursors in the incubation media. The secretion of both labelled and total triacylglycerol and apolipoprotein B was reduced approximately 50% in cells incubated chronically with eicosapentaenoic acid. The amounts of triacylglycerol and apolipoprotein B within the cells were not decreased by chronic exposure to eicosapentaenoic acid. Our data indicate that CaCo-2 cells chronically incubated with eicosapentaenoic acid secrete significantly less triacylglycerol than cells incubated chronically with oleic acid. When eicosapentaenoic acid was also included acutely, triacylglycerol secretion was reduced even more. We conclude that chronic exposure of eicosapentaenoic acid to this intestinal cell type reduces the rate of chylomicron secretion and may help explain the decreased postprandial lipaemia observed in humans taking fish oil supplements.

Adenocarcinoma↗
PubMed1994

Fatty acid uptake and metabolism in CaCo-2 cells: eicosapentaenoic acid (20:5(n-3)) and oleic acid (18:1(n-9)) presented in association with micelles or albumin.

In vivo, long-chain fatty acids are incorporated into bile salt micelles, which solubilize the hydrophobic fatty acids before they are transported across the unstirred water layer to the intestinal brush border membrane. In the present study we have used CaCo-2 cells, cultured on filter membranes as a model of human enterocytes, and compared presentation of fatty acids bound to albumin with a micellar form. Absorption of eicosapentaenoic acid and oleic acid from micellar solutions was 4-times faster than from fatty acid-albumin complexes after 5 h incubation, and resulted in a corresponding increase in triacylglycerol synthesis and secretion. Mass determination of newly synthesized, cell-associated triacylglycerol after 5 h incubation, indicated a 5-fold increase in cells exposed to a micellar solution versus albumin-complexed fatty acids. A 2-fold larger fraction of the absorbed fatty acids was incorporated into triacylglycerol than into phospholipids when the fatty acids were presented as micelles. Analysis by resistive pulse technique showed that secreted lipoproteins of density less than 1.006 g/ml were in the same size-range as chylomicrons derived from human plasma. In spite of an increased amount of secreted triacylglycerol from cells supplemented with micellar fatty acids, there was no increase in the mean size of these particles. Synthesis and secretion of triacylglycerol in cells exposed to eicosapentaenoic acid and oleic acid were similar regardless of whether the fatty acids were presented to the cells associated with albumin or micelles, although the total amount of triacylglycerol synthesized and secreted was highest with micelles. When incubating CaCo-2 monolayers with eicosapentaenoic acid or oleic acid bound to albumin, a similar amount of radioactivity was released as CO2 and acid soluble products into the medium with less than 3% of the lipids being oxidized after 5 h of incubation. The oxidation rate of fatty acids in cells incubated with micelles was increased 40 to 100%. In conclusion, micellar fatty acids are absorbed, metabolized and influence secretion of lipoprotein particles to a higher extent than albumin-bound fatty acids in CaCo-2 cells, and there is no major difference between eicosapentaenoic acid and oleic acid.

Albumins↗
PubMed1994

Stimulation of fatty acid oxidation by a 3-thia fatty acid reduces triacylglycerol secretion in cultured rat hepatocytes.

The present work shows that when mitochondrial beta-oxidation is stimulated by the hypolipemic, non-beta-oxidizable fatty acid analogue tetradecylthioacetic acid, there is a decrease in the secretion of triacylglycerol in cultured rat hepatocytes. In order to study the effects of tetradecylthioacetic acid in cells with different fatty acid oxidation rates, cells were grown without or with L-carnitine supplement or with addition of the beta-oxidation inhibitor L-aminocarnitine. In cells grown without and with L-carnitine in the medium, the oxidation of [1-14C]oleic acid was stimulated by tetradecylthioacetic acid, whereas it was not significantly changed by palmitic acid. In cells grown with L-aminocarnitine, oxidation of [1-14C]oleic acid was almost abolished both in the absence and in presence of tetradecylthioacetic acid. The effect of tetradecylthioacetic acid and palmitic acid on incorporation of [1-14C]oleic acid into triacylglycerol was similar under all conditions. In the presence of L-carnitine, secretion of oleic acid-labeled triacylglycerol was reduced significantly more by tetradecylthioacetic acid than by palmitic acid. The effects of tetradecylthioacetic acid and palmitic acid on secretion of oleic acid-labeled triacylglycerol were reversed in cells grown with L-aminocarnitine, where palmitic acid was the stronger inhibitor. These results were substantiated by determination of mass of triacylglycerol secreted. It is concluded that tetradecylthioacetic acid reduces secretion of triacylglycerol from rat hepatocytes mainly by acutely stimulating fatty acid oxidation.

Animals↗
PubMed1993

The hypotriglyceridemic effect of eicosapentaenoic acid in rats is reflected in increased mitochondrial fatty acid oxidation followed by diminished lipogenesis.

The effect of eicosapentaenoic acid (EPA) on fatty acid oxidation and on key enzymes of triglyceride metabolism and lipogenesis was investigated in the liver of rats. Repeated administration of EPA to normolipidemic rats resulted in a time-dependent decrease in plasma triglycerides, phospholipids and cholesterol. The triglyceride-lowering effect was observed after one day of feeding whereas lowering of plasma cholesterol and phospholipids was observed after five days of treatment. The triglyceride content of liver was reduced after two-day treatment. At that time, increased mitochondrial fatty acid oxidation occurred whereas mitochondrial and microsomal glycerophosphate acyltransferase was inhibited. The phosphatidate phosphohydrolase activity was unchanged. Adenosine triphosphate:citrate lyase, acetyl-CoA carboxylase, fatty acid synthetase and glucose-6-phosphate dehydrogenase were inhibited during the 15 d of EPA treatment whereas peroxisomal beta-oxidation was increased. At one day of feeding, however, when the hypotriglyceridemic effect was established, the lipogenic enzyme activities were reduced to the same extent in palmitic acid-treated animals as in EPA-treated rats. In cultured rat hepatocytes, the oxidation of [14C]palmitic acid to carbon dioxide and acid-soluble products was stimulated in the presence of EPA. These results suggest that the instant hypolipidemia in rats given EPA could be explained at least in part by a sudden increase in mitochondrial fatty acid oxidation, thereby reducing the availability of fatty acids for lipid synthesis in the liver for export, e.g., in the form of very low density lipoproteins, even before EPA induced peroxisomal fatty acid oxidation, reduced triglyceride biosynthesis and diminished lipogenesis.

Animals↗
PubMed1993

Dietary supplementation of very long-chain n-3 fatty acids decreases whole body lipid utilization in the rat.

Rats were fed lard or n-3 fatty acid-supplemented diets ad libitum to study whole body oxidation of lipid and carbohydrate. One group of male rats was fed 21% fat (by weight) containing 19.5% lard and sufficient amounts of essential fatty acids (1.5%). Another group of rats had 6.5% of the lard replaced by ethyl esters of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). The rats were fed these diets for 6-8 weeks. Body weight gain was similar for the two groups and absorption of fat was complete in animals fed both types of fatty acids. Indirect calorimetric measurements, after 3-5 weeks on these diets, by continuous registration of O2 consumption and CO2 formation showed no difference in mean energy expenditure during the experimental period. However, the mean respiratory quotient (RQ) was significantly increased for animals fed the n-3 fatty acid-supplemented diet. This was noted both under fasting conditions and after receiving a test meal of similar fatty acid composition for both feeding groups. Thus, mean substrate utilization demonstrated reduced oxidation of fat and increased oxidation of carbohydrate, during fasting as well as fed periods for the n-3 fatty acid group as compared to the lard group. After an additional 2-3 weeks, blood plasma, liver, and muscle samples were collected, and adipocytes and hepatocytes were isolated. Reduced postprandial plasma concentrations of triacylglycerol, phospholipids, unesterified fatty acids, and glycerol were promoted by the n-3 fatty acid diet as compared to lard. Plasma concentration of glucose was slightly increased, and liver and muscle content of glycogen were decreased in the n-3 fatty acid-fed rats. Experiments with isolated adipocytes showed decreased basal lipolysis after feeding n-3 fatty acids for 6-8 weeks for suspended epididymal adipocytes, whereas stimulated lipolysis by isoproterenol (0.1 microM) was higher in both epididymal and mesenteric adipocytes from rats fed n-3 fatty acids as compared to animals fed lard. In addition, epididymal adipocytes from rats fed n-3 fatty acids were significantly smaller than cells from animals fed lard. Hepatic peroxisomal fatty acid oxidation was significantly higher for n-3 fatty acid-supplemented animals, but total fatty acid oxidation was similar in both dietary groups. The hepatic content of triacylglycerol and phospholipids was similar for both diets. These results demonstrate that n-3 fatty acid replacement of a high-fat diet containing mostly saturates and monoenes for several weeks promotes reduced use of fat as energy source. This may be explained by decreased plasma concentration of unesterified fatty acids.(ABSTRACT TRUNCATED AT 400 WORDS)

Adipose Tissue↗
PubMed1992

Serum lipids, hepatic glycerolipid metabolism and peroxisomal fatty acid oxidation in rats fed omega-3 and omega-6 fatty acids.

Rats were fed, for 3 weeks, high-fat (20% w/w) diets containing sunflower-seed oil, linseed oil or fish oil. Chow-fed rats were used as a low-fat reference. The high-fat diets markedly reduced non-fasting-rat serum triacylglycerol as compared with the low-fat reference, and the highest reduction (85%) was observed with the fish-oil group, which was significantly lower than that of the other high-fat diets. The serum concentration of phospholipids was significantly reduced (30%) only in the fish-oil-fed animals, whereas serum non-esterified fatty acids were reduced 40-50% by both the fish-oil- and linseed-oil-fed groups. The liver content of triacylglycerol showed a 1.7-fold increase with the fish-oil diet and 2-2.5-fold with the other dietary groups when compared with rats fed a low-fat diet, whereas the hepatic content of phospholipids was unchanged. Peroxisomal fatty acid oxidation (acyl-CoA oxidase) was 2-fold increased for the rats fed fish oil; however this was not significantly higher when comparison was made with rats fed the linseed-oil diet. There was no difference in phosphatidate hydrolysis (microsomal and cytosolic fractions) among animals fed the various diets. Acyl-CoA:diacylglycerol acyltransferase activity was increased by all high-fat diets, but the fish-oil-diet-fed group showed a significantly lower enzyme activity than did rats fed the other high-fat diets. A linear correlation between acyl-CoA:diacylglycerol acyltransferase activity and liver triacylglycerol was observed, and the microsomal enzyme activity was decreased 40-50% by incubation in the presence of eicosapentaenoyl-CoA. CoA derivatives of arachidonic, linolenic and linoleic acid had no inhibitory effect when compared with the control. These results indicate that dietary fish oil may have greater triacylglycerol-lowering effect than other polyunsaturated diets, owing to decreased triacylglycerol synthesis caused by inhibition of acyl-CoA:diacylglycerol acyltransferase. In addition, increased peroxisomal fatty acid oxidation and decreased availability of non-esterified fatty acids could also contribute by decreasing the amounts of fatty acids as substrates for triacylglycerol synthesis and secretion.

Acyltransferases↗
PubMed1992

Effect of dietary supplementation with n-3 polyunsaturated fatty acids on physical properties and metabolism of low density lipoprotein in humans.

The effects of marine n-3 polyunsaturated fatty acids were investigated in relation to the chemical and physical properties of low density lipoprotein (LDL) and how these changes affected LDL metabolism in humans. The subjects received supplements of six capsules daily, each capsule containing 1 g of either highly concentrated ethyl esters of n-3 fatty acids (85% eicosapentaenoic acid and docosahexaenoic acid) (n = 12) or corn oil (56% linoleic and 26% oleic acid) (n = 11). After 4 months of oil supplementation, the following changes were observed in the lipid moiety of the n-3-enriched LDL particles compared with LDL from the corn oil group: LDL cholesteryl ester, as well as the amount of total lipids of LDL, was significantly lower (0.97 +/- 0.12 versus 1.19 +/- 0.23 mg/mg protein and 1.88 +/- 0.40 versus 2.45 +/- 0.31 mg/mg, respectively; mean +/- SD, n = 6, p less than 0.05); the amount of eicosapentaenoic and docosahexaenoic acids and the unsaturation index increased (104.0 versus 29.4 micrograms/mg protein and 6.64 versus 5.49, respectively); and differential scanning calorimetry showed that LDL cholesteryl ester melting temperature was lowered by 2 degrees C (27.6 +/- 0.8 degrees versus 29.5 +/- 0.2 degrees C). The only effect observed on the protein moiety was an increase in the ratio of apolipoprotein (apo) B to cholesterol (0.66 +/- 0.17 versus 0.82 +/- 0.14 mg/mg cholesterol; p less than 0.05). Circular dichroism spectra of LDL indicated an alpha-helix content of 46 +/- 5% in apo B from both groups. No difference was observed by 13C nuclear magnetic resonance spectroscopy in the ratio of "active" to "normal" lysine residues of apo B. No detectable differences in the size of n-3 fatty acid-enriched LDL particles versus control LDL could be measured by either electron microscopy of negatively stained LDL (24.5 +/- 2.0 versus 25.0 +/- 1.5 nm) or dynamic light scattering (24.9 +/- 0.9 versus 24.9 +/- 0.4 nm). LDL from the fish oil and corn oil groups showed similar susceptibility to Cu(2+)-catalyzed lipid peroxidation, as indicated by the amount of lipid peroxides formed during the oxidation time, and degradation of oxidatively modified LDL in J774 macrophages as a function of Cu2+ oxidation time. No effect of n-3 fatty acids was observed on LDL metabolism. Specific uptake and degradation of n-3 fatty acid-enriched LDL were similar to those for control LDL in HepG2 cells as well as in human skin fibroblasts, and they showed the same ability to stimulate cholesteryl ester synthesis.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗
PubMed1992

Influence of eicosapentaenoic acid (20:5, n-3) on secretion of lipoproteins in CaCo-2 cells.

CaCo-2 cells, grown on filter membranes, were used to study the effects of fatty acids on cellular metabolism of triacylglycerol and phospholipids. The rate of triacylglycerol secretion was enhanced more than 2-fold, from 1 to 2 weeks after reaching confluency, in the presence of 0.6 mM fatty acids. Triacylglycerol secretion and oxidation of oleic acid increased 2- and 9-fold, respectively, with this culture system, as compared to cells grown on conventional plastic dishes. Eicosapentaenoic acid (20:5 n-3), when compared to oleic acid, did not reduce formation of triacylglycerol or enhance phospholipid synthesis in CaCo-2 cells during short term (less than 24 h) experiments, when the cells resided on membranes, regardless of what type of radioisotopes were used as precursors in the incubation media. However, the n-3 fatty acid was preferentially incorporated into phosphatidylinositol, lysophosphatidylcholine, and sphingomyelin, as compared to oleic acid. The disappearance from the apical medium and cellular uptake of labeled eicosapentaenoic and oleic acid were similar during incubations up to 24 h, and the metabolism of these fatty acids to acid-soluble materials and CO2 was equal. Light scattering analysis indicated that secreted lipoproteins of density less than 1.006 g/ml were in the same size-range as chylomicrons derived from human plasma. Assessment of secreted apolipoprotein B showed that by incubating CaCo-2 cells with oleic acid, apolipoprotein B levels increased approximately 1.4-fold when compared to cells incubated with eicosapentaenoic acid, whereas the amount of triacylglycerol and size-range of particles were similar for the two fatty acids. Our data indicate that CaCo-2 cells grown on filter membranes exhibit enterocyte-like characteristics with the ability to synthesize and secrete chylomicrons. Eicosapentaenoic acid and oleic acid are absorbed, metabolized, and influence secretion of lipoprotein particles in a similar way, except for some differences in incorporation of the fatty acids into certain phospholipid classes and a reduced secretion of apolipoprotein B. The culture conditions, including time after confluency and cellular support, are critical for the rate of secretion in the presence of eicosapentaenoic acid and oleic acid.

Apolipoproteins B↗
PubMed1991

Effect of dietary n-3 and n-6 fatty acids on fatty acid desaturation in rat liver.

To study the effect of high-fat diets with varying contents of n-3 and n-6 fatty acids on the metabolism of essential fatty acids, the rat liver microsomal fatty acid desaturases were measured. The rats were fed for 3 weeks with diets high in linseed oil (18:3(n-3)), sunflower seed oil (18:2(n-6)) or fish oil (20:5(n-3) and 22:6(n-3)) (20%, w/w) using pellet fed rats as a reference. The delta 6-desaturase using 18:2(n-6) or 18:3(n-3) as substrates was stimulated 1.5-2.5-fold by linseed or sunflower seed oil, compared to the pellet reference. The delta 5-desaturase was stimulated 3.5-fold with linseed oil and 2.5-fold with sunflower seed oil, while the delta 9-desaturase was inhibited by all the high-fat diets. The delta 6-, 5- and 9-desaturase activities were in all cases considerably reduced with fish oil as compared to linseed and sunflower seed oil diets. With pellet fed rats the rates were highest for delta 9-desaturation and in decreasing order lower for delta 5-desaturation, delta 6-desaturation with 18:3 (n-3) as substrate and finally delta 6-Desaturation with 18:2(n-6) as substrate. The content of 20:4(n-6) in liver phospholipids increased with the diets rich in 18:2(n-6), and was reduced for the fish oil diet enriched in 20:5 and 22:6(n-3) fatty acids. The amount of 20:5(n-3) in phospholipids was as high with linseed oil diet as with the fish oil diet, while the 22:6(n-3) content was only increased with the fish oil diet.

Animals↗
PubMed1990

Effect of 3- and 4-thia-substituted fatty acids on glycerolipid metabolism and mitochondrial beta-oxidation in rat liver.

Treatment of normolipidemic rats by alkylthiopropionic acid (CETTD), resulted in a dose- and time-dependent increase in total dihydroxyacetone phosphate acyltransferase (DHAPAT) activity, in extent comparable to that of 3-thiadicarboxylic acid (BCMTD) and alkylthioacetic acid (CMTTD). Thus, in CETTD- and CMTTD-treated rats, the specific DHAPAT activity increased in the microsomal, peroxisomal and mitochondrial fractions. In contrast, repeated administration of the peroxisome proliferator, BCMTD, decreased the specific DHAPAT activity both in the peroxisomal fraction and in purified peroxisomes. A three-fold increase in specific activity was, however, revealed in the mitochondrial fraction. Whether the variation of the DHAPAT activity in the mitochondrial and microsomal fractions among the feeding groups can be explained by increased number of enlarged and small peroxisomes sedimenting in the fractions, are to be considered. Subcellular fractionation studies confirmed previous findings that rat liver glycerophosphate acyltransferase (GPAT) was located both in mitochondria and the microsomal fraction. BCMTD was considerably more potent than CMTTD in stimulating the microsomal and mitochondrial GPAT activities. Administration of CETTD marginally affected the isoenzymes of GPAT. Diacylglycerol acyltransferase (DGAT) activity was increased by 35% in BCMTD and CMTTD treated rats, but by administration of CETTD the enzyme activity was decreased by more than 80%. The acyl-CoA cholesterol acyltransferase (ACAT) activity was marginally affected in animals treated with BCMTD, CMTTD and CETTD. Thus, the results indicate that the initial steps in the synthesis of triacylglycerols and ether glycerolipids as well as the last step in triacylglycerol synthesis could not be identified as mediating the fat accumulation or the lowering of triacylglycerol content in liver of CETTD, or BCMTD and CMTTD treated rats. On the other hand, CMTTD increased the palmitoyl-CoA oxidation in mitochondria, and CETTD considerably inhibited the activity. Therefore, it is conceivable that the development of fatty liver with CETTD is mostly due to inhibition of mitochondrial beta-oxidation.

Acyltransferases↗
PubMed1990

Regulation of fatty acid oxidation and triglyceride and phospholipid metabolism by hypolipidemic sulfur-substituted fatty acid analogues.

The mechanisms behind the hypotriglyceridemic effect of 1,10-bis(carboxymethylthio)decane (3-thiadicarboxylic acid) and tetradecylthioacetic acid and the development of fatty liver caused by 3-tetradecylthiopropionic acid (Aarsland et al. 1989. J. Lipid Res. 30: 1711-1718.) were studied in the rat. Repeated administration of S-substituted non-beta-oxidizable fatty acid analogues to normolipidemic rats resulted in a time-dependent decrease in plasma triglycerides, phospholipids, and free fatty acids. This was accompanied by an acute reduction in the liver content of triglycerides and an increase in the hepatic concentration of phospholipids. Mitochondrial fatty acid oxidation was stimulated, whereas lipogenesis was inhibited. The activity of phosphatidate phosphohydrolase decreased while the activity of CTP:phosphocholine cytidylyltransferase increased. These results suggest that the observed triglyceride-lowering effect was due to increased mitochondrial fatty acid oxidation accompanied by a reduction in the availability of the substrate i.e., free fatty acid, along with an enzymatic inhibition (phosphatidate phosphohydrolase). Administration of 3-tetradecylthiopropionic acid led to a drastic increase in the hepatic triglyceride content. Levels of plasma triglyceride phospholipid and free fatty acid also increased. Phosphatidate phosphohydrolase activity was stimulated whereas CTP:phosphocholine cytidylyltransferase was inhibited. Mitochondrial fatty acid oxidation was decreased. These data indicate that the development of fatty liver as an effect of 3-tetradecylpropionic acid is probably due to accelerated triglyceride biosynthesis, which is mediated by an increase in the availability of fatty acid along with stimulation of phosphatidate phosphohydrolase. The results of the present study speak strongly in favor of the hypothesis that phosphatidate phosphohydrolase is a major rate-limiting enzyme in triglyceride biosynthesis. Furthermore, they point out that the biosynthesis of triglycerides and phospholipids might be coordinately regulated. Such regulation is possibly mediated via phosphatidate phosphohydrolase and CTP:phosphocholine cytidylyltransferase. Whether the increase in hepatic phospholipids via increased CDP-pathway accounts for an increase of lipid components for proliferation of peroxisomes (3-thiadicarboxylic acid and tetradecylacetic acid) should be considered.

Animals↗
PubMed1989

Eicosapentaenoic acid inhibits hepatic production of very low density lipoprotein.

Synthesis and secretion of triacylglycerol and cholesterol ester by cultured hepatocytes were highest in the presence of oleic, palmitic, stearic and linoleic acid, and lowest with eicosapentaenoic (EPA) and docosahexaenoic acid. Cellular formation and secretion of triacylglycerol and cholesterol ester were reduced by 50-80% in the presence of EPA in comparison with oleic acid. Reduced formation of triacylglycerol and cholesterol ester was also observed when EPA was given together with oleic acid. EPA caused higher incorporation of (3H)water into phospholipid and lower incorporation into triacylglycerol and cholesterol ester as compared with oleic acid. Rat liver microsomes incorporated EPA-CoA into triacylglycerol and cholesterol ester to a lesser extent than oleoyl-CoA. Decreased formation of triacylglycerol and cholesterol ester was also observed when EPA-CoA was given together with oleoyl-CoA, whereas palmitoyl-CoA, stearoyl-CoA, linolenoyl-CoA and arachidonoyl-CoA had no inhibitory effects. In conclusion, inhibition of acyl-CoA:1,2-diacylglycerol acyltransferase and acyl-CoA:cholesterol acyltransferase by EPA may be important for decreased hepatic synthesis and secretion of triacylglycerol and cholesterol ester.

Acyl Coenzyme A↗
PubMed1988

Eicosapentaenoic acid inhibits cholesterol esterification in cultured parenchymal cells and isolated microsomes from rat liver.

The effects of eicosapentaenoic acid on synthesis and secretion of cholesterol and cholesterol ester by cultured rat hepatocytes were studied. In the presence of eicosapentaenoic acid cellular cholesterol esterification was decreased by 50-75% compared to oleic acid as measured by radioactive precursors and mass. Secretion of cholesterol ester was reduced by 50-60% in the presence of eicosapentaenoic acid as evaluated by radiolabeled fatty acids, mevalonolactone, and mass measurement. Oleic, palmitic, and stearic acid increased, whereas eicosapentaenoic and docosahexaenoic acid decreased synthesis and secretion of cholesterol ester as compared to a fatty acid-free control. Cellular and secreted free cholesterol were unaffected by eicosapentaenoic acid in comparison with oleic acid. The reduced cholesterol esterification was observed within 1 h and lasted for at least 20 h. Eicosapentaenoic acid caused lower cholesterol esterification than oleic acid in the concentration range 0.2-1.0 mM fatty acid and reduced the stimulatory effect of oleic acid on cholesterol ester formation. Cholesterol esterification and release of cholesterol ester were markedly increased by 25-hydroxycholesterol in the presence of eicosapentaenoic acid as well as oleic acid. Experiments with liver microsomes revealed that radioactive eicosapentaenoic acid and eicosapentaenoyl-CoA were poorer substrates (7-30%) for cholesterol esterification than oleic acid and oleoyl-CoA. Reduced formation of cholesterol ester was also observed when eicosapentaenoyl-CoA was given together with labeled oleoyl-CoA, whereas palmitoyl-CoA, stearoyl-CoA, linolenoyl-CoA, and arachidonoyl-CoA had no inhibitory effect. In conclusion, eicosapentaenoic acid reduced cellular cholesterol esterification by inhibiting the activity of acyl-CoA:cholesterol acyltransferase. The lowered cholesterol esterification caused by eicosapentaenoic acid secondly decreased secretion of very low density lipoprotein cholesterol ester.

Animals↗
PubMed1988

Eicosapentaenoic acid reduces hepatic synthesis and secretion of triacylglycerol by decreasing the activity of acyl-coenzyme A:1,2-diacylglycerol acyltransferase.

The mechanism for the reduced hepatic production of triacylglycerol in the presence of eicosapentaenoic acid was explored in short-term experiments using cultured parenchymal cells and microsomes from rat liver. Oleic, palmitic, stearic, and linoleic acids were the most potent stimulators of triacyl[3H]glycerol synthesis and secretion by hepatocytes, whereas erucic, alpha-linolenic, gamma-linolenic, arachidonic, docosahexaenoic, and eicosapentaenoic acids (in decreasing order) were less stimulatory. There was a linear correlation (r = 0.85, P less than 0.01) between synthesis and secretion of triacyl[3H]glycerol for the fatty acids examined. The extreme and opposite effects of eicosapentaenoic and oleic acids on triacylglycerol metabolism were studied in more detail. With increasing number of free fatty acid molecules bound per molecule of albumin, the rate of synthesis and secretion of triacyl[3H]glycerol increased, most markedly for oleic acid. Cellular uptake of the two fatty acids was similar, but more free eicosapentaenoic acid accumulated intracellularly. Eicosapentaenoic acid caused higher incorporation of [3H]water into phospholipid and lower incorporation into triacylglycerol and cholesteryl ester as compared to oleic acid. No difference was observed between the fatty acids on incorporation into cellular free fatty acids, monoacylglycerol and diacylglycerol. The amount of some 16- and 18-carbon fatty acids in triacylglycerol was significantly higher in the presence of oleic acid compared with eicosapentaenoic acid. Rat liver microsomes in the presence of added 1,2-dioleoyl-glycerol incorporated eicosapentaenoic acid and eicosapentaenoyl-CoA into triacylglycerol to a lesser extent than oleic acid and its CoA derivative. Decreased formation of triacylglycerol was also observed when eicosapentaenoyl-CoA was given together with oleoyl-CoA, whereas palmitoyl-CoA, stearoyl-CoA, linoleoyl-CoA, linolenoyl-CoA, and arachi-donoyl-CoA had no inhibitory effect. In conclusion, inhibition of acyl-CoA:1,2-diacylglycerol O-acyltransferase (EC 2.3.1.20) by eicosapentaenoic acid may be important for reduced synthesis and secretion of triacylglycerol from the liver.

Acyl Coenzyme A↗
PubMed1987

Calcium-antagonists inhibit secretion of very-low-density lipoprotein from cultured rat hepatocytes.

The effects of different calcium-antagonists on secretion of very-low-density lipoprotein (VLDL) from cultured rat hepatocytes were examined. Verapamil (an inhibitor of voltage-dependent calcium channels) and EGTA (a calcium chelator) decreased VLDL-triacylglycerol secretion in a concentration-dependent manner, with maximum inhibition (about 90%) at 0.2 mM-verapamil and 5 mM-EGTA. Inorganic calcium-antagonists such as lanthanum, nickel, cobalt and manganese decreased secretion of VLDL-triacylglycerol by 55-95%, whereas the calcium-agonist barium did not affect secretion. Inhibition of VLDL-triacylglycerol secretion appeared within 30 min, without inhibition of triacylglycerol synthesis. Pulse-chase experiments revealed that verapamil and cobalt inhibited the secretory pathway itself. Cobalt showed a concentration-dependent inhibition of VLDL-triacylglycerol secretion, with maximal effect at 8 mM. Although inhibition by cobalt was not completely reversible, Trypan Blue exclusion and lactate dehydrogenase leakage indicated that the hepatocytes were not injured by cobalt or any of the other calcium-antagonists tested. Inhibition of protein synthesis by cycloheximide did not affect triacylglycerol secretion (up to 2 h), and the observed effects were therefore probably not due to impaired production of apolipoproteins. Taken together, these results suggest that calcium is important for secretion of VLDL particles.

Animals↗
PubMed1987

Inhibition of very-low-density lipoprotein secretion by chloroquine, verapamil and monensin takes place in the Golgi complex.

The effects of chloroquine, verapamil and monensin on secretion of very-low-density lipoproteins (VLDLs) were studied in cultured rat hepatocytes. Maximum inhibition of VLDL-triacylglycerol secretion by 50-90% of control was reached at 200 microM chloroquine, 200 microM verapamil and 5 microM monensin, whereas no effect on cellular triacylglycerol synthesis was observed. The inhibition could be seen within 15 min and was reversible after washout of the drugs. Chloroquine and verapamil inhibited both cellular protein synthesis and protein secretion, whereas monensin reduced protein secretion without any effect on protein synthesis. Control experiments with cycloheximide revealed that intact protein synthesis was not necessary for secretion of VLDL-triacylglycerol during 2 h. Electron micrographs of cells treated with chloroquine, verapamil or monensin showed swollen Golgi cisternae containing VLDL-like particles. By morphometry, a more than 2-fold increase in volume fractions and size indices of Golgi complexes and secondary lysosomes was observed, except that monensin had no significant effect on these parameters of secondary lysosomes. These results suggest that the inhibition of VLDL secretion by chloroquine, verapamil and monensin which takes place in the Golgi complex might be due to disruption of trans-membrane proton gradients. An increase in pH of acidic Golgi vesicles may cause swelling and disturb sorting and membrane flow through this organelle.

Animals↗