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Incorporation of 14C-1-palmitate into rabbit fetal lipids in vivo.

14C-1-palmitate was injected into rabbit fetuses in utero. Fetal liver, blood, carcass, and placenta collected after 15, 60, 180, and 360 minutes were analyzed for radioactivity in lipid classes. Incorporation differed with tissue and lipid. Highest total early specific activity was detected in plasma but later liver displayed the most extensive incorporation. Phospholipid incorporation was more rapid in all tissues than that of other glycerides. Plasma triglycerides did not display any radioactivity for 3 hours. Results point to multicentric origin of plasma lipids. Ready utilization of the palmitate indicates that maternal fatty acid transfer plays an important role in fetal lipid hemeostasis.

Animals↗

Effect of maternal isoxsuprine on fetal rabbit lung biochemical maturation.

Isoxsuprine was given intramuscularly for 24 hours to pregnant rabbits at 26 to 27 days' gestation. There was no effect on fetal weight within one day of the last injection. Fetuses of isoxsuprine-treated mothers had higher lung lecithin (6.67 micron per gram +/- 2.81) and higher tracheal wash lecithin/sphingomyelin (L/S) ratios (5.85 +/- 4.5) than did untreated fetuses--lung lecithin (5.35 micron per gram 1.23) and tracheal wash L/S (4.08 +/- 2.3). In contrast, phosphorylcholine glyceride transferase activity of the fetal lung was less in isoxsuprine-treated fetal lung tissue.

Animals↗

Coomassie brilliant blue staining of lipids on thin-layer plates.

Coomassie brilliant blue staining of lipids on silica gel thin-layer chromatography plates is described. This stain proved to be useful for the wide-range detection of simple and complex lipids on thin-layer plates. It can stain several classes of lipids, including cholesterol, cholesterol esters, glycerides, phospholipids, ceramides, and neutral and acidic glycosphingolipids. It stains the spots evenly without a corrosive reagent, and is simple to use and suitable for storage. The visual detection limits of this stain for lipids were 0.05 to 0.5 microgram.

Chromatography, Thin Layer↗

Properties of lipid bilayer membranes made from lipids containing phytanic acid.

Besides the preparation of phytanic acid (3,7,11,15-tetramethylhexadecylic acid) according to the Dumas-Stass reaction, the synthesis of four different lipids containing phytanic acid residues is described. Diphytanoyl phosphatidylcholine was synthesized beginning from glycerylphosphorylcholine, whereas the other lipids, diphytanoyl phosphatidylethanolamine, diphytanoyl phosphatidylserine and monophytanoyl glyceride were prepared by total synthesis. Some properties of lipid bilayer membranes made from the lipids containing phytanic acid were investigated. The specific capacity of these membranes was measured. Its value of approximately 400 nF cm-2 was found to be similar to the value of membranes from lipids with unbranched fatty acid residues. Charge pulse experiments were performed using dipicrylamine as a molecular probe of membrane structure. The results were discussed on the basis of a higher viscosity of the membranes from lipids containing phytanic acid residues compared with unbranched fatty acid residues.

Amines↗

The composition and fluidity of normal and leukaemic or lymphomatous lymphocyte plasma membranes in mouse and man.

The lymphocyte surface membranes from normal and leukaemic or lymphomatous cells from man and mouse were isolated, characterized, and analyzed both biochemically and by diphenyl hexatriene fluorescence polarization. The cholesterol/phospholipid molar ratio for all the pure lymphocyte plasma membranes was 0.45--0.50, and the fluorescence polarization results showed that values much higher than this were not credible. The lipid composition of all the plasma membranes was remarkably similar, except for the concentration of free fatty acids and glycerides. The latter two were particularily high in the mouse lymphoma membrane and these, rather than a low cholesterol concentration, were responsible for the increased fluidity of the cells. The most prominent protein in most of the plasma membrane preparations was actin. This is found only by some authors, and its presence probably depends on the method of lymphocyte disruption.

Adult↗

Active labeling of phosphatidylcholines by [1-14C]docosahexaenoate in isolated photoreceptor membranes.

Isolated bovine rod outer segments and photoreceptor disks actively incorporated [1-14C]docosahexaenoate (22:6) into phospholipids when incubated in the presence of CoA, ATP, and Mg2+. About 80% of the esterified fatty acid was in phosphatidylcholine (PC). Microsomal and mitochondrial fractions incorporated as much 22:6 as rod outer segments, but it was distributed among various phospholipids and neutral glycerides. The isolated photoreceptor membrane thus contains an acyl-CoA synthetase which activates the fatty acid and a docosahexaenoyl-CoA-lysophosphatidylcholine acyltransferase activity. The specific radioactivity of PC was higher in rod outer segments than in the other subcellular fractions. About 2/3 of the label in photoreceptor membrane PC was in its dipolyunsaturated molecular species and 1/3 in hexaenes. Dipolyunsaturated PCs showed high turnover rates of 22:6 in all three subcellular membranes, especially in mitochondria. Retinal membranes in vitro seem to take up free [14C]22:6 from the medium by simple diffusion or partition into the membrane lipid. The ability of these membranes to activate and esterify [1-14C]22:6 indicates that docosahexaenoate-containing molecular species of retina lipids, including those of photoreceptor membranes, are subject to acylation-deacylation reactions in situ.

Animals↗

Structural organisation and lipid composition of the epicuticular accessory layer of infective larvae of Trichinella spiralis.

The epicuticle of infective larvae of Trichinella spiralis represents the interface between this intracellular nematode parasite and the cytosol of mammalian skeletal muscle. The macromolecular structures that make up the epicuticle were studied by freeze-fracture electron microscopy and compositional analysis. Three fracture planes were observed: one with a typical plasma membrane-type bilayer organisation which was overlaid by two extended layers of lipid in an inverted cylindrical configuration. This overall structure remained unchanged in response to variations in temperature between 20 degrees C and 45 degrees C. The lipid cylinders were on average 6.8 nm in diameter, with randomly-associated particles that were not dissociated by high-salt treatment, indicative of hydrophobically associated proteins. The majority of the lipids were non-polar, consisting of cholesterol, cholesterol esters, mono- and tri-glycerides, and free fatty acids. Three major classes of phospholipids were identified: phosphatidylethanolamine, phosphatidylglycerol and phosphatidylcholine. Total lipid extracts did not adopt an inverted cylindrical or micellar configuration on isolation, but formed flat sheets of lamellae as did the purified polar and non-polar fractions of the lipids. Isolated lipids did not undergo thermally-induced polymorphism between 20 degrees C and 60 degrees C and there was no pH dependency of the structures adopted. The fatty acid saturation levels of the phospholipids were compatible with the observation that they did not form polymorphic structures on isolation. We suggest that this unusual configuration is probably stabilised by the associated (glyco)proteins and may be required for selective permeation of nutrients from the host cell cytosol and/or for maintaining the high curvature of the parasite within the cell.

Animals↗

Lipoprotein lipase and uptake of chylomicron triacylglycerol and cholesterol by perfused rat mammary tissue.

The role of lipoprotein lipase in the uptake of chylomicron triacylglycerol and cholesterol from blood was studied in perfused inguinal-abdominal mammary tissue of rats lactating 10-15 days. Lipoprotein lipase activity in the tissue was reduced, from 0.47 to 0.10 units/g, by removing the anterior pituitary gland from lactating rats 2 days before the experiment. Perfused mammary tissue of normal lactating rats took up 12% of the chylomicron triacylglycerol infused, whereas the tissue of hypophysectomized lactating rats took up less than 1%. About two-thirds of the triacylglycerol taken up was retained as glyceride, and the rest was hydrolyzed and released to the perfusing fluid as fatty acids and glycerol. Autoradiographic studies of perfused tissues of normal lactating rats showed that both the acyl and glycerol moieties derived from chylomicron triacylglycerol were incorporated into milk lipid droplets. Perfused mammary tissue of normal lactating rats also took up 15% of the chylomicron cholesterol infused, whereas the tissue of hypophysectomized lactating rats took up less than 1%. The findings demonstrate that chylomicron cholesterol is taken up with triacylglycerol by lactating mammary tissue, and that uptake of both lipids is markedly suppressed when lipoprotein lipase activity is low, as in tissue of hypophysectomized rats. It is proposed that uptake of triacylglycerol from chylomicrons by mammary tissue requires the action of lipoprotein lipase, while uptake of cholesterol is dependent on reduction of the triacylglycerol core, resulting from action of the enzyme on the core and uptake of lipolytic products by the tissue.

Animals↗

Soluble rat adipocyte phosphatidate phosphatase activity: characterization and effects of fasting and various lipids.

Phosphatidate phosphatase (phosphatidate phosphohydrolase, EC 3.1.3.4) was present at very high specific activity in the soluble fraction of isolated rat adipocytes. Using phosphatidate in aqueous dispersion 90% of its hydrolysis depended on the presence of Mg2+. Mg2+ appeared to almost saturate the enzyme at 20-40 mM with no indication of an optimum. The substrate concentration was optimum at 1.2 mM and the pH at 6.8. Initial rates were linear for only 4-5 min at optimum conditions. Increasing inhibition occurred at high phosphatidate concentrations. At optimum conditions acid or alkaline phosphatase activity was not measurable. The Mg2+-dependent activity was enhanced by 3-sn-phophatidylcholine and inhibited by albumin, 3-sn-phosphatidyletanolamine, 3-sn-phosphatidylinositol, diacylglycerol, oleoyl-CoA, and oleate. Oleoyl-CoA was the most potent "effector". Fasting for 24, 48 and 72 h decreased the activity both relative to protein and to DNA. The activity thus decreased to about one-third of that of the fed rat during 72 h of fasting. The effects of Mg2+, various lipids, and fasting may indicate that some form of control of glyceride synthesis can be exerted through the soluble phosphatidate phosphatase.

Adipose Tissue↗

The acyl-CoA desaturases of microsomes from rat liver and the Morris 7777 hepatoma.

We have investigated the role of the microsomal oxidative desaturase in defining the aberrant phosphoglyceride fatty acid composition of hepatomas. The microsomal delta 9-stearoyl-CoA, delta 6-oleoyl(linolenoyl)-CoA, and delta 5-eicosatrienoyl-CA desaturase activities were studied in control and host liver and in the poorly differentiated Morris 7777 hepatoma. The delta 9-stearoyl-CoA desaturase of the hepatoma was significantly decreased (42%) relative to control liver, yet the hepatoma specific activity was twice that of host liver. Additionally, the specific activity of the delta 9-stearoyl-CoA desaturase of the tumor was found to decrease with increasing tumor weight. Also this desaturase was inactivated by freezing and thawing. The delta 6-oleoyl(linolenoyl)-CoA and delta 5-eicosatrienoyl-CoA desaturases of the hepatoma were 39% and 4% of control, respectively. The electron transport components involved in the desaturase system were reduced, although this did not appear to be rate-limiting. In addition, two competing metabolic reactions which could lower the observed desaturase activities, hydrolysis of the thioester and incorporation of substrate acyl-CoA molecules into glycerides, did not appear to be responsible for the lowered desaturase activities of the tumor. Thus, it appears that reduced levels of the desaturases themselves may be responsible for the observed activities. These results indicate that the capacity of the hepatoma to biosynthesize polyunsaturated fatty acids is greatly reduced and this is consistent with the decreased polyene content observed in many neoplasms.

Animals↗

Cell triacylglycerol accumulation from very low density lipoproteins isolated from normal and hypertriglyceridemic human sera.

Human fibroblast cells in culture increased their intracellular triacylglycerol levels when exposed to very low density lipoproteins (VLDL) isolated from human plasma. This response was dependent on the amount of VLDL added. VLDL from normal, type IV or type V sera gave similar results. Lipoprotein lipase enhanced this intracellular triacylglycerol accumulation. It was concluded that human fibroblast cells in culture have at least two mechanisms for triacylglycerol uptake from VLDL: (1) uptake from intact lipoprotein either by surface transfer of lipoprotein lipid or internalization of the entire lipoprotein particle, and (2) re-esterification of lower glyceride and fatty acids released by lipoprotein lipase degradation of VLDL.

Fibroblasts↗

De novo fatty acid synthesis in the perfused rat lung. Incorporation of palmitate into phospholipids.

1. The incorporation of exogenously derived [14C]palmitate and endogenously synthesized [3H]palmitate (from 3H2O) was measured in the isolated perfused lung. 2. Over 40% of the fatty acid esterified into lung disaturated phosphatidylcholine was derived from de novo synthesis. 3. A major portion of the palmitate synthesized de novo was incorporated in the 2 position of disaturated phosphatidylcholine. 4. Streptozotocin-induced diabetes and the compound 5-(tetradecyloxy)-2-furoic acid markedly inhibited de novo fatty acid synthesis while the incorporation of exogenously supplied palmitate increased into disaturated phosphatidylcholine, primarily in the 2 position. 5. Treatment with insulin resulted in an increase in [14C]glucose incorporation into lung phospholipid, with the largest increase appearing in the glyceride-glycerol fraction of the phosphatidylcholine species. 6. Insulin neither stimulated de novo fatty acid synthesis nor increased exogenous palmitate incorporation. 7. These data show: (1) that de novo fatty acid synthesis in the perfused rat lung is involved in the remodeling reactions in the synthesis of phosphatidylcholine, and (2) that diabetes affects the relative contribution of de novo synthesized and exogenously supplied palmitate available for the esterification of lung phospholipid.

Animals↗

Effect of salts on membrane binding and activity of adipocyte phosphatidate phosphohydrolase.

Isosmotic replacement of sucrose in a low ionic strength homogenizing buffer (0.25 M sucrose/1 mM EDTA/1 mM Tris-HCl, pH 7.4) with KCl increased the microsomal and decreased the soluble phosphatidate phosphohydrolase (EC 3.1.3.4) activity of isolated rat fat cells. At 54 mM KCl the microsomal specific activity was increased 6-fold and the soluble activity was decreased to less than one-third. Binding of enzyme was promoted by KCl and NaCl when the once isolated soluble and microsomal fractions were recombined and incubated at 37 degrees C. Half-maximal binding occurred at about 17 mM salt and maximal binding at about 50 mM. The pH optimum of binding was 7.8 in 15 mM Hepes. MgCl2, CaCl2 and spermine prevented desorption of microsomal enzyme at mu molar levels and maximal effects were observed at concentrations below the 1 mM level. At maximum, however, the prevention of desorption was less by these salts than it was by KCl. MgCl2 and spermine also interfered with the effect of KCl. Moderate salt-induced loading of microsomes with the phosphohydrolase (specific activity increased 3.5-fold) increased their ability to incorporate 14C into triacylglycerol from sn-[U-14C]glycerol 3-phosphate while a high loading (specific activity increased 6-fold) had no effect or even suppressed it. The results are discussed in relation to a role of translocation of phosphatidate phosphohydrolase in glyceride biosynthesis and its control.

Adipose Tissue↗

Comparative metabolism of free and esterified fatty acids by the perfused rat heart and rat cardiac myocytes.

Studies have been conducted on the uptake and metabolism of unesterified oleic acid and lipoprotein triacylglycerol by the perfused rat heart, and of oleic acid, free glycerol and lipoprotein triacylglycerol by rat cardiac myocytes. The perfused heart efficiently extracted and metabolized unesterified fatty acid and the fatty acid released during lipolysis of the recirculating triacylglycerol. The released glyceride glycerol, however, was largely accumulated in the perfusion media. Cardiac myocytes also extracted and rapidly metabolized unesterified fatty acid. As with the intact heart, free glycerol was poorly utilized by cardiac myocytes. Although the cells appeared to extract a small amount of available extracellular triacylglycerol presented as very low density lipoprotein, this was shown to be unmetabolized, suggesting adsorption rather than surface lipolysis and uptake of the released fatty acid. The data suggest that myocytes are unable to metabolize triacylglycerol fatty acids without prior lipolysis by extracellular (capillary endothelial) lipoprotein lipase.

Animals↗

Effect of phosphatidylcholine and free fatty acids on the activity of pancreatic lipase-colipase.

Mixed micelles of bile salt and phospholipids inhibit the lipase-colipase-catalysed hydrolysis of triacylglycerols. Free fatty acids can reverse this inhibition and reactivate lipase-colipase. This reactivation is either due to the formation of a high-affinity complex between lipase and colipase induced by free fatty acids and/or to a change of the quality of the interface. Lauric acid, oleic acid and linoleic acid are the most potent reactivators, while short-chain free fatty acids have no effect and long-chain, saturated free fatty acids inhibit the lipase-colipase activity further. The physiological relevance of these results is evident as the glyceride emulsion reaching the duodenum already contains free fatty acids due to the activity of lingual lipase in the stomach.

Animals↗

Lipolysis products promote the formation of complexes of very-low-density and low-density lipoproteins.

In the course of lipolysis, surface lipid products may accumulate on very-low-density lipoproteins (VLDL). To investigate potential lipoprotein interactions mediated by such products, radiolabeled low-density lipoproteins (LDL) were incubated with VLDL and bovine milk lipoprotein lipase in the presence of limited free fatty acid acceptor. With partial VLDL degradation, association of radiolabeled LDL with VLDL remnants or larger aggregates of VLDL density was demonstrated by gradient gel electrophoresis, agarose chromatography, and density gradient ultracentrifugation. VLDL-LDL complex formation was also observed in incubations with lipid extracts from lipolyzed VLDL or with purified palmitic acid in the absence of lipolysis. Complex formation was inhibited by addition of increasing amounts of albumin as free fatty acid acceptor, but could be detected at molar ratios of free fatty acids/albumin that occur in vivo. Composition analysis of LDL reisolated following incubation with VLDL and lipase under conditions favoring partial complex formation revealed enrichment in glycerides and depletion of cholesterol. We conclude that lipolysis products can promote the formation of stable complexes of LDL and VLDL, and that physical interactions of this nature may play a role in the transfer of lipids and apolipoproteins between lipoprotein particles.

Albumins↗

Decreased serum lipids, serum insulin and triacylglycerol synthesis in adipose tissue of JCR:LA-corpulent rats treated with benfluorex.

Rats of the JCR:LA-corpulent strain were treated with benfluorex daily at a dose of 25 mg/kg body weight. This strain of rat, if homozygous for the cp gene (cp/cp), is hyperphagous, obese, hypertriglyceridemic, insulin resistant and in the case of male rats, atherosclerosis prone. The benfluorex treatment produced a sharp reduction in food intake which remained suppressed despite recovery toward normal after 2 weeks of treatment. This was accompanied by sustained decreases in body weight and adipose tissue mass. The ability of adipose tissue from female rats to take up glucose and convert it to lactate, glyceride-glycerol and fatty acids was decreased. This decrease was largely due to decreased adipose tissue mass. The serum concentrations of glucose, lactate, triacylglycerol, cholesterol, phospholipids and insulin were decreased in both sexes. The treatment also improved glucose tolerance and decreased corticosterone concentrations in male rats only. While reduction of food consumption contributes to the effects seen, benfluorex clearly had significant direct metabolic effects. The effects are consistent with an improved insulin sensitivity leading to a decrease in circulating triacylglycerol. The changes produced by benfluorex are all in directions that should inhibit atherogenesis in this animal model for the human obesity/hypertriglyceridemia/insulin resistant syndrome.

Adipose Tissue↗

Oscillation of electrical potential in a porous-membrane doped with triolein induced by an Na+/K+ concentration gradient.

The electrical potential across a fine pore membrane doped with trioleoyl glyceride (triolein) and separating aqueous solutions of 0.5M NaCl and 0.5M KCl, respectively, was studied. It was found that this system showed rhythmic and sustained oscillations of electrical potential between the two aqueous solutions. These oscillations were attributed to the change of permeability of Na+ and K+ ions across the membrane, which originated from the phase-transition of triolein molecules within the fine pores.

Mathematics↗