P.H.E. Groot

Postprandial lipoprotein metabolism in normolipidemic men with and without coronary artery disease.

A delayed clearance of postprandial lipoproteins from the plasma may play a role in the etiology of premature coronary atherosclerosis. To address this hypothesis, we studied chylomicron (remnant) metabolism in two groups of 20 selected normolipidemic men aged 35-65 years, a group of coronary artery disease (CAD) patients, and a matched control group with documented minimal coronary atherosclerosis. Subjects received an oral fat load supplemented with cholesterol and retinyl palmitate. Plasma samples obtained during the next 24-hour period were analyzed for total as well as d less than 1.019 g/ml and d greater than 1.019 g/ml triacylglycerol, cholesterol, and retinyl ester concentrations. Although both groups of patients responded identically in terms of the appearance of gut-derived lipids in the plasma, CAD patients showed a marked delay in the clearance of retinyl esters as well as in the normalization of plasma triacylglycerol concentrations. Postheparin plasma hepatic lipase activity was significantly lower in the CAD group. Apolipoprotein E phenotype measurements did not reveal marked differences in frequency between both groups. The frequency distribution was not unusual in comparison with the normal Dutch population. The magnitude of the postprandial responses of triacylglycerol and retinyl esters was correlated positively with the fasting levels of plasma triacylglycerol and negatively with high density lipoprotein subfraction 2 cholesterol concentrations. These data indicate that the clearance of postprandial lipoproteins in normolipidemic CAD patients as selected in the present study is delayed as compared with that of controls without coronary atherosclerosis and suggest that postprandial lipoproteins may play a role in the etiology of their disease.

Fatty Acid Activation: Specificity, Localization, and Function

Publisher Summary This chapter discusses fatty acid activation. Most cells can metabolize fatty acids. The most important pathways are β-oxidation to acetyl-CoA; esterification with glycerol phosphate, glycerol derivatives, sphingosine, glycol, and long-chain alcohols; chain elongation; desaturation; and α and ω oxidation. Fatty acid molecules are completely ionized at physiological pH values. The carboxylate ion is chemically unreactive toward nucleophilic substitution. Esters, thioesters, and acid anhydrides, in which both the carboxylate resonance and the negative charge on the carboxyl group disappear, are more reactive. Both microsomal and mitochondrial long-chain fatty acid activation is stimulated by the presence of high concentrations of several salts. Both activities are also strongly stimulated by the addition of the particle-free supernatant from several rat tissues. Purified mammalian acyl-CoA synthetases are very specific in their nucleotide requirement.

Effects of amount and type of dietary fat on serum lipids, lipoproteins and apolipoproteins in man: A controlled 8-week trial☆

We have studied whether a low-fat diet is as effective in lowering some risk factors for atherosclerosis as a diet rich in polyunsaturated fat (PUFA). During a 2.5 week control period, 60 volunteers were given a moderate-fat diet (MOD) providing 30% of the daily energy intake (energy %) in the form of fat, one-third of which was PUFA. For the next 5 weeks subjects were divided into 4 groups and received diets providing varying amounts of total fat and of PUFA: for group LO, 20 energy % PUFA; group HIPUF, 40 energy % fat and 19 energy % PUFA; and group HISAT, 40 energy % fat and 3 energy % PUFA. The diets contained the same amounts of cholesterol, phytosterols, oligosaccharides and other nutrients, known to affect serum lipid levels. All food was prepared daily and weighed out for each individual appropriate to his energy needs. Nutrient intakes were checked by 7-day records and by chemical analysis of double portions. On diet LO, total serum cholesterol concentration increased by 0.25 mmol/l while HDL cholesterol concentration did not change significantly. The HDL cholesterol/apoprotein-A1 ratio fell, and VLDL and LDL triglyceride centrations were elevated. On the HIPUF diet, total serum cholesterol concentration was not significantly lower, but HDL cholesterol concentration increased by 0.10 mmol/l. On the HISAT diet, total serum cholesterol concentration went up by 0.38 mmol/l; 0.12 mmol/l of this was due to HDL. LDL cholesterol/serum apoprotein-B ratios were unaffected by any of the diets. It was concluded that after 5 weeks, the influence of a low-fat, high-carbohydrate diet on the concentrations of serum lipoproteins was less favourable than that of moderate- or high-fat diets rich in polyunsaturated fatty acids.

Serum Lipoproteins of Healthy Persons fed a Low-Fat Diet or a Polyunsaturated Fat Diet for Three Months A Comparison of two Cholesterol-Lowering Diets

Abstract In a controlled 16-week trial we have compared the effects on some risk factors for atherosclerosis of 2 cholesterol-lowering diets, one low in total fat and low in polyunsaturated fat (LO) and one moderate in total fat but high in polyunsaturated fat (MOD) as recommended by various advisory councils. All 35 volunteers were given diet MOD during a 2.5-week control period. This diet provided 31% of the daily energy intake (energy%) as total fat; one-third of the fatty acids were polyunsaturated (PUFA). VLDL plus LDL cholesterol decreased by 0.66 mmol/1 during the control period while the HDL cholesterol (HDL-C) level was unchanged. Total serum triglycerides decreased by 0.33 mmol/l. After the control period the subjects were randomized into 2 groups, one of which continued on diet MOD, while the other group received the low-fat diet LO, providing 2.1 energy% as total fat and 4 energy% as PUFA. All food was prepared daily and weighed out for each individual in amounts appropriate to the individuals energy needs. Nutrient intakes were checked by 7-day records and by chemical analysis of double portions. The diets contained the same amounts of cholesterol, phytosterols, oligosaccharides and other nutrients known to affect serum lipid levels. Total serum cholesterol increased by 0.21 ± 0.41 mmol/l on control diet MOD during the test period of 13 weeks. 0.09 ± 0.11 mmol/l of this was due to HDL-C. Total serum triglycerides remained constant during the test period in group MOD. On diet LO total serum cholesterol remained stable during the test period, but HDL-C was lowered by −0.06 ± 0.20 mmol/1. The difference in HDL-C between the diet groups was mainly located in HDL2. Total serum triglycerides increased by +0.32 ± 0.31 mmol/l in group LO; the increase in VLDL triglycerides was +0.22 ± 0.18 mmol/l. The depression of HDL on diet LO was also reflected in the ratio of HDL to total cholesterol which decreased by −0.02 ± 0.05 on diet LO, and was unchanged on diet MOD, and in the ratio of apo A1 to apo B (6% decrease on diet LO and constant on diet MOD). Combination of these data with those from a preceding study [Brussaard et al., Atherosclerosis, 36 (1980) 515] leads to the following conclusions: 1. (1) Both diets lower total serum cholesterol levels when compared with the habitual diets of affluent communities. 2. (2) The high-carbohydrate, low-fat diet causes lower HDL and higher fasting VLDL triglyceride levels than the recommended moderate-fat-high-PUFA diet.

Liver lipase and high-density lipoprotein lipoprotein changes after incubation of human serum with rat liver lipase

Human sera were incubated with rat liver lipase after inactivation of lecithin:cholesterol acyltransferase, and the changes in serum lipoprotein composition were measured. In the presence of liver lipase serum triacylglycerol and phosphatidylcholine were hydrolyzed. The main changes in the concentrations of these lipids were found in the high-density lipoprotein fraction. Subfractionation of high-density lipoprotein by rate-zonal ultracentrifugation showed a prominent decrease in all constituents of high-density lipoprotein2, a smaller decrease in the light high-density lipoprotein3 and an increase in the heavy high-density lipoprotein3. These data support a concept in which liver lipase is involved in high-density lipoprotein2 phospholipid and triacylglycerol catabolism and suggest that as a result of this action high-density lipoprotein2 is converted into high-density lipoprotein3.

Selective degradation of the high density lipoprotein-2 subfraction by heparin-releasable liver lipase.

The lipase released from the liver after intravenous injection of heparin (liver lipase, EC 3 .l .1.3) has a broad substrate specificity [l-3]. Hydrolysis of ester bonds in triacylglycerol, diacylglycerols, 1 or 2 monoacylglycerols, pahnitoyl-coenzyme A and phosphatidylcholine (the 1 acyl ester bond) has been demonstrated, using artificial lipid emulsions. Although a role in serum lipoprotein metabolism has generally been considered, the physiological function of this enzyme has not been well established. Evidence has been presented, based on in vivo inhibition of liver lipase by intravenous injection of a specific antiserum in rats, that liver lipase is involved in HDL and possibly LDL-phospholipid catabolism [4-61. Based on these experiments a role of liver lipase in the conversion of HDL-2 into HDL3 was proposed. This hypothesis was developed further by in vitro incubations of liver lipase with rat HDL-2 [7] and by in vivo turnover studies of HDLphospholipids and cholesterol esters [8]. Here, the hypothesis was further investigated. As HDL2 and HDL3 are poorly defmed in rat, we turned to human serum and measured the phospholipase A activity of purified human heparin-releasable liver lipase with isolated HDL-2 and HDL-3. It was found that the rate of phosphatidylcholine hydrolysis with HDL2 as substrate exceeds the rate with HDL3 by a factor of -10. These data provide further evidence for a role of liver lipase in the degradation of HDL-2.

The activation and oxidation of octanoate and palmitate by rat skeletal muscle mitochondria

Abstract 1. 1. Rat skeletal muscle mitochondria can oxidize octanoate just as well as palmitate. The oxidation of both fatty acids is strongly carnitine dependent, which indicates that the fatty acid CoA ester formation (fatty acid activation) is localized on the outer membrane. 2. 2. The palmitoyl-CoA synthetase and octanoyl-CoA synthetase activities were measured in these mitochondria. Palmitoyl-CoA synthetase was inhibited by octanoate while the octanoyl -CoA synthetase was inhibited by palmitate. Both inhibitions seem to be of the competitive type. These results suggest that both fatty acids are activated by the same enzyme. 3. 3. Octanoyl-CoA synthetase in rat skeletal muscle mitochondria was strongly (3 to 4 times) stimulated by the addition of skeletal muscle cytosol or by the addition of a high concentration of salt.

Low density lipoprotein receptor of macrophages facilitates atherosclerotic lesion formation in C57Bl/6 mice.

Macrophage-derived foam cells play an important role in the initiation and progression of atherosclerosis. To examine the role of the macrophage low density lipoprotein receptor (LDLr) in atherosclerotic lesion formation, bone marrow from LDLr knockout [LDLr(-/-)] mice was transplanted into irradiated wild-type C57Bl/6 [LDLr(+/+)] mice. After 3 months on an atherogenic diet, C57Bl/6 mice, reconstituted with LDLr(-/-) bone marrow, showed a mean lesion area of 34.7 x 10(3)+/-22.4 x 10(3) microm(2) compared with 100. 8 x 10(3)+/-33.0 x 10(3) microm(2) (P<0.001) in control C57Bl/6 mice that were transplanted with LDLr(+/+) bone marrow. There were no significant differences in total serum cholesterol, triglyceride levels, and lipoprotein profiles between the 2 groups. Histochemical analysis of macrophage LDLr expression in the atherosclerotic lesions indicated that C57Bl/6 mice, reconstituted with LDLr(+/+) bone marrow, showed extensive staining of the foam cells in the atherosclerotic lesions, whereas mice reconstituted with LDLr(-/-) bone marrow showed only a few LDLr-positive foam cells. In vitro, peritoneal macrophages isolated from wild-type C57Bl/6 mice were, respectively, 4.7- and 10.7-fold more effective in cell association and degradation of atherogenic (125)I-beta-very low density lipoprotein than were LDLr(-/-) peritoneal macrophages, establishing that the LDLr on macrophages is important for the interaction of macrophages with beta-very low density lipoprotein. It is concluded that the LDLr on macrophages can facilitate the development of atherosclerosis, possibly by mediating the uptake of atherogenic lipoproteins.

Organ and intracellular localization of short-chain acyl-CoA synthetases in rat and guinea-pig

1. Homogenates of rat epididymal fat pad, heart, kidney, lactating mammary gland, liver, skeletal muscle and small intestinal mucosa have been partitioned into a particulate and supernatant fraction. With reliable marker enzymes for the mitochondrial matrix and the cytosol: propionyl-CoA carboxylase and pyruvate kinase, the distributions of the acyl-CoA synthetase activities measured at 1 and 10 mM C2, C3 and C4 over mitochondria and cytosol have been calculated. From these values an estimate was made of the K0.5 of the fatty acids. 2. A distinct fatty acid-activating enzyme was assumed to be present in one of the compartments when that fatty acid was activated with a K0.5 less than or equal to 1.5 mM in an amount of greater than 13% of the total cellular activity. Adipose tissue, gut, liver and mammary gland, all organs of a high lipogenetic capacity, contained a cytosolic acetyl-CoA synthetase. At 1 mM acetate 60, 31, 77 and 83% of the total cellular activities in these organs were cytosolic in nature, with activities of 0.021, 0.32, 0.37 and 1.16 mumol C2 activated per min per g wet weight, respectively. 3. Mitochondrial acetyl-CoA and butyryl-CoA synthetases were found in adipose tissue, gut, heart, kidney, mammary gland and muscle. They were absent in liver. Adipose tissue and liver contained a mitochondrial propionyl-CoA synthetase with activities at 1 mM C3 of 0.014 and 1.50 mumol C3 activated per min per g wet weight, respectively. 4. At 1 mM, C2 was activated with decreasing rates by kidney, heart, mammary gland and gut (7.6-1.0 mumol C2 activated per min per g wet weight). C3 (1 mM) activation was about equal (1.6-1.9 mumol C3 activated per min per g wet weight) in liver, kidney and heart. C4 (1 mM) was activated with decreasing rates by heart, liver, kidney and gut (4.0-0.5 mumol C4 activated per min per g wet weight) in the order given. 5. The influence of the isolation method and the diet on fatty acid activation in small intestinal mucosal scrapings have been studied. To demonstrate the existence of cytosolic acetyl-CoA synthetase in fed animals a pre-treatment of everted intestine by low amplitude vibration has been found essential. Also C16 activation was highly (95%) decreased in a non-pre-vibrated preparation. 24 h starvation lowered cytosolic C2 and total C16 activation by 90 and 80%, respectively. Refeeding of starved rats with a balanced fat-free diet, and not with sucrose only, gave the same cytosolic C2 and total C16 activation as normally fed rats. 6. In guienea-pig heart, kidney, liver and muscle about the same partitions have been found as in the respective rat organs. The acetate activation in liver was factor 6 lower. Acetate and butyrate activation in guinea-pig muscle was much higher (6 and 37 times, respectively).

Acyl-CoA synthetases in guinea-pig liver mitochondria: Purification and characterization of a distinct propionyl-CoA synthetase

Guinea-pig liver mitochondria contain three soluble ATP-dependent acyl-CoA synthetases: (a) a medium-chain acyl-CoA synthetase, (b) a salicylate activating enzyme, and (c) a propionyl-CoA synthetase. A complete separation of these enzymes has been accomplished and the resulting preparation of propionyl-CoA synthetase (Spec. act. 4 units/mg protein) accepts acetate, propionate and butyrate as substrates with a high preference for propionate.