Lipoprotein lipid and protein synthesis in experimental nephrosis and plasmapheresis. I: Studies in rat in vivo

The incorporation of L-4,5-[³H]leucine into the ultracentrifugally separated apolipoproteins of very low, low, and high density lipoproteins (VLDL, LDL, HDL) and into serum albumin was found three-to four-fold higher in nephrotic than in normal rats one hour after intravenous injection. Incorporation of leucine into the circulating lipids was negligible. Increases of similar magnitude were obtained in the incorporation of simultaneously injected 1,5[¹⁴C] citrate into the lipids of VLDL, LDL, and HDL of nephrotic rats. Of the citrate carbons incorporated into serum and liver lipids, the proportion in cholesterol was higher in nephrotic rats when compared to normal rats. The incorporation of both precursors into total proteins and lipids of the liver vs. the incorporation into the lipoproteins was relatively lower in nephrotic than in control rats, indicating a preferential channeling into secretable products. The occurrence of enhanced new lipid synthesis in nephrosis was corroborated by the finding of markedly enhanced synthesis of lipoprotein-borne fatty acids and cholesterol from³H₂O. These results point out that while leucine is not an efficient in vivo precursor of lipoprotein lipids in nephrosis, de novo lipogenesis proceeds from other precursors. Similar trend of changes, though of smaller magnitude, was elicited in rats after double plasmapheresis, 18 hr apart, when measured 3 hr after the second plasma withdrawal. This indicates that the loss of circulating proteins either by direct removal or through kidney lesion stimulates the compensatory hepatic response involving excessive lipoprotein synthesis. Time-course studies showed that peak incorporation of leucine and citrate into the protein and lipid components of lipoproteins, respectively, as well as into serum albumin, occurred coincidentally 3 hr after the second plasmapheresis, suggesting an interdependence of the enhanced protein and lipid synthesis.     

Powder synthesis from metal-organic precursors

Thermal conversion of organic polymers such as polymethylsilane [(CH₃)₂Si]_n, polymethylphenylsilane, and polysilazane or inorganic polymer such as silicimide [Si(NH)₂]_n, and thermal and hydralylic decomposition of metal alkoxides, M(OR)_n, have been employed to obtain submicron size 30–500 Å powders, continuous fibers and thin films of refractory carbides, nitrides and oxides. The high surface activity associated with these powders make possible relatively low temperature processing of the powders compact to near theoretical density and uniform fine grain size bodies. Transmission electron microscopy is used to show nucleation, growth crystallite morphology of the powders synthesized, and microstructural features observed.     

Effect of marine oil and rapessee oil on composition of fatty acids in lipoprotein triacylglycerols from rat blood plasma and liver perfusate

The fatty acid patterns of triacylglycerols (TG) from very low density lipoprotein (VLDL) in blood plasma and liver-perfusate from rats fed partially hydrogenated marine oil or rapeseed oil were determined. In the plasma from rats fed rapeseed oil for three days and three weeks, there was a small but significant decrease in the percentage of 22∶1 fatty acid from 17.2 to 11.2% with length of feeding. In liver-perfusate, the comparable decrease with dietary rapeseed oil was from 18.5 to 5.2%, and with dietary marine oil from 13.4 to 8.0%. In contrast to the liver-perfusate, the remaining liver had only a very low 22∶1 composition (ca 2%) independent of feeding period or diet. The results indicated that the liver exported the very long chain fatty acids and that an adaptation took place after three days feeding with rapeseed oil or marine oil. This adaptation in the liver could possibly explain why TG accumulation in hearts, which appears after three days' feeding with rapeseed oil or marine oil, disappears after an extended feeding period.     

Lipoprotein lipid and protein synthesis in experimental nephrosis and plasmapheresis: II. Perfused rat liver

Livers from rats with experimental hypoproteinemia induced by aminonucleoside-nephrosis or plasmapheresis were perfused with a [¹⁴C]-labeled amino acid mixture at physiological concentration. Compared to control rats, a significantly increased incorporation of the amino acid label was found in the apolipoproteins of the ultracentrifugally separated very low and high density lipoproteins (VLDL, HDL), and into albumin secreted into the perfusate. However, no increase in the amino acid-derived label was detected in VLDL- or HDL-borne lipids in nephrosis or plasmapheresis. Perfusion with U-[¹⁴C] leucine as a lipogenesis precursor at >10 times higher than physiological concentration resulted in 5-fold increase in the label incorporation into perfusate proteins in nephrosis but only in a slightly significant increase in perfusate lipids. In contrast, the incorporation of a preformed fatty acid, 9,10-[³H] oleate into VLDL and HDL lipids increased 3- to 4-fold in nephrosis. Both with leucine and oleate as precursors, the increments in the label appearing in perfusate proteins or lipids, respectively, were markedly greater than the increases in hepatic tissue proteins or lipids. The results indicate that amino acids are preferentially directed by the liver into the synthesis of circulating apolipoproteins and albumin in hypoproteinemia and do not seem to constitute an important precursor of the lipoprotein lipids. The increased production of apolipoproteins is associated with an increased incorporation of preformed fatty acids into lipoprotein lipids in addition to the previously reported stimulation of hepatic de novo lipid synthesis from precursors other than amino acids.     

Plasma and lipoprotein fatty acid composition in glycogen storage disease type I

Nocturnal intragastric feeding has been shown to be an effective means to improve clinical and biochemical features in glycogen storage disease type I (GSD-I). In this study, we investigated the fatty acid patterns in a whole plasma and in circulating lipoproteins in patients on this therapy. The results demonstrated massive concentration of total fatty acids coupled with higher levels of triglycerides, free cholesterol, cholesterol ester and phospholipids. This hyperlipidemia involved all fatty acids without distinction of carbon or bond numbers. However, the increase was more pronounced for saturated than polyunsaturated fatty acids, as was demonstrated by the ratios of both oleic acid to linoleic acid (1.91±0.40 vs 0.80±0.09 in controls) and of ω3+ω6 to ω9 fatty acid families (0.92±0.11 vs 1.66±0.08 in controls). The fatty acid patterns in very low (VLDL), low (LDL) and high (HDL) density lipoprotein showed substantial differences in composition, reflecting an association between an abnormal lipoprotein pattern and essential fatty acid deficiency. Furthermore, GSD-I patients exhibited a significant increase in VLDL (17±2 vs 47±7 mg/dl) and LDL cholesterol (124±7 vs 206±24 mg/dl), coupled with a decrease in HDL cholesterol (49±4 vs 28±3 mg/dl). These data documenting high LDL cholesterol and low HDL cholesterol associated with an increased concentration and proportion of saturated fatty acids suggest that GSD-I patients on nocturnal intragastric feeding are at high risk for atherosclerosis and its complications.     

Metabolic fate of sphingomyelin of high-density lipoprotein in rat plasma

The metabolic fate of high density lipoprotein (HDL) sphingomyelin in plasma was studied in rats over a 24-hr period after injection of HDL containing sphingomyelin which was¹⁴C-labeled in the stearic (18∶0) or lignoceric acid (24∶0) moiety and³H-labeled in the choline methyl groups. Decay of label in plasma followed three phases. The first two phases were similar for both isotopes and both types of sphingomyelin (t_1/2≃10 and 110 min). However, during the third phase (from 10 hr after injection),³H label disappeared more slowly than¹⁴C label from 18∶0 sphingomyelin, whereas the³H/¹⁴C ratio remained relatively constant when 24∶0 sphingomyelin was used. Intact, doubly-labeled 18∶0 sphingomyelin disappeared from HDL rapidly (t_1/2=38 min) by tissue uptake and by transfer to very low density lipoprotein (VLDL). VLDL contained up to 12% of the sphingomyelin 1 hr after injection. This is the first demonstration of a transferin vivo of sphingomyelin from HDL to VLDL. A similarly rapid transfer was also observedin vitro. Some nontritated, [¹⁴C]18∶0 or [¹⁴C]24∶0 sphingomyelin was redistributed more slowly into HDL. Doubly-labeled phosphatidylcholine appeared in VLDL and HDL within 1 hr after injection and reached 1.8 and 2.1% of the injected¹⁴C and³H in VLDL at 1 hr, and 4.8 and 6.9% in HDL at 3 hr, respectively.     

Carbothermal synthesis of vanadium and chromium carbides from solution-Derived precursors

Vanadium and chromium tartrate precursors prepared from aqueous solutions have been used as preceramic materials for carbothermal reactions with and without simultaneous nitridation. Their thermal behaviour has been investigated by TG/DTA, X-ray diffraction and measurement of surface areas. Under pyrolysis up to about 600°C, reactive composites consisting of intimately mixed carbon and amorphous M₂O₃ oxides are formed by salt decomposition. The subsequent processes of crystallization of oxides and carbothermal reduction render both the composites and the reduction products porous. The carbothermal reactions leading either to V₈C₇, a V(C,N,O) solid solution, and Cr₃C₂, respectively, proceed at moderate temperatures between 800 and 1100°C. The final products result as assemblages of fine carbide or carbonitride particles, however, with extensive heat treatment the particles grow and the surface areas diminish. Air oxidation of the final products has been studied by simultaneous TG/DTA.     

Leucine and isoleucine as in vitro precursors for lipid synthesis by rat aorta

The in vitro conversion of¹⁴C-labeled leucine, isoleucine, and pyruvate to specific lipids was compared in rat aorta, diaphragm, and fat pad. Total lipid specific radioactivity from all precursors was greatest in aorta. The ratio of label incorporated into polar lipids vs. neutral lipids by aorta was generally several-fold that incorporated by muscle and fat pad. The labeling of sterols in the aorta from¹⁴C-leucine and pyruvate was equivalent. It is concluded that leucine may be a substantial precursor to polar lipids and to sterols in rat aorta. This paper was presented in part at the 1974 Biochemistry and Biophysics Meeting of the Fed. Am. Soc. Exp. Biol.     

Influence of carbon on the synthesis of AlN powder from combustion synthesis precursors

AlN powders were synthesized by carbothermal reduction of combustion synthesis precursors. Water-soluble organics and carbon black were used as carbon sources. The effects of carbon on the synthesis of AlN powders were studied. Results showed that AlN powders were synthesized directly from γ-Al₂O₃ without γ-Al₂O₃ to α-Al₂O₃ phase transition when water-soluble organics were used as carbon sources, and the nitridation of the precursors could be completed at 1400 °C. However, AlN powders were synthesized from the nitridation of α-Al₂O₃ when carbon black was used as carbon source, and the reaction temperature for a complete conversion increased to 1500 °C. The particles of AlN powders synthesized with water-soluble organics was smaller than the particles of AlN powders synthesized with carbon black and their particle size distribution was sharper. The specific surface area of synthesized AlN powders increased with the increase of carbon content in the precursors.     

Metabolic studies in isolated rat liver cells: I. Lipid synthesis

Rat liver cells, isolated by chelate perfusion and extrusion through a tissue press, incorporated labeled acetate into cellular lipids and into lipids released into the suspending medium. Optimal rates of incorporation required supplementation of tris-KCl medium with Mg⁺⁺, Mn⁺⁺, succinate, citrate, nicotinamide, Coenzyme A, NADP and glucose-6-phosphate. The rate of acetate incorporation was markedly altered by changes in incubation media; tris-KCl was the most effective buffer. All the major classes of cellular lipids were labeled. ATP, BSA, inorganic phosphate, Ca⁺⁺, 2,4-dinitrophenol and sodium clofibrate were potent inhibitors of acetate incorporation. When added to the incubation mixture, several hormones altered the rate of acetate incorporation into lipids.     

Conjoined crystals. I. composition and physical properties

Conjoined Crystals is the name given to a new food emulsifier whose special properties are associated with its content of glycerol monoester in a stabilized α-crystalline form (6). It consists of the mixture of crystals formed by cooling a melt containing approximately equal molecular proportion of certain long chain fatty acid monoesters of glycerol and of 1,2-propanediol. A practical composition contains a blend of glycerol monostearate and 1,2-propanediol monostearate. Conjoined Crystals disperse readily in water and retain this ability for periods of over a year, a property which appears to be closely associated with their effectiveness in baking and in other applications. This water dispersibility contrasts with the behavior of the usual modification of glycerol monostearate which is difficult to disperse in water. Infrared analysis indicated that the major part of the glycerol monoester in the mixed crystals is in the α-crystalline modification. Studies by X-ray diffraction supported this conclusion. Thus, we have consistently associated ready water dispersibility and the α-form with the enhanced emulsifying ability. Communication No. 293.     

One-step synthesis of radioactive acyl-CoA and acylcarnitines using rat liver mitochondrial outer membrane as enzyme source

Rat liver mitochondrial outer membrane enriched preparations have proven to be a convenient enzyme source for synthesizing coenzyme A (CoA) and carnitine esters of radioactive fatty acids. These membranes are simple to isolate and they retain acyl-CoA ligase and carnitine palmitoyltransferase activities well upon storage. Enzyme purification is not required. A novel aspect of the present procedure is that the same enzymatic incubation step allows both the acyl-CoA and the acylcarnitine esters to be obtained simulataneously when carnitine is present, but produces acyl-CoA ester only when carnitine is not included. Under the conditions described, the conversion of [1-¹⁴C]octanoic acid to the respective esters was about 95%; the corresponding figure for [1-¹⁴C]palmitic acids was over 70%. The procedure seems suitable for synthesizing the labeled CoA and carnitine esters from a variety of radioactive fatty acids. A preliminary account of this work has been published (ref. 1).     

Bio-inspired synthesis: understanding and exploitation of the crystallization process from amorphous precursors

Many biominerals, such as mollusk nacre, sea urchin, bone and teeth, are found to form by an amorphous precursor pathway, and these biominerals have remarkable properties, which are better than their artificial material counterparts that are formed at high temperatures and high pressures. More than ever, synthesizing technologically relevant materials following nature's way with a specific size, shape, orientation, organization, and complex form has been a focus of ongoing interest due to the increasing need for low cost and environmentally friendly approaches to processing advanced materials. Herein, we present recent developments in the crystallization process from amorphous precursors by primarily drawing on results from our own laboratory, and discuss some unique characteristics from the transformation process that can be exploited for the design and synthesis of artificial functional materials.     

Combustion synthesis of heterogeneous calcium phosphate bioceramics from calcium oxide and phosphate precursors

The solid-state reaction of combustion synthesis has been demonstrated to be capable of rapidly producing materials for biomedical applications. Reactant powders of CaO and P₂O₅ were pressed into cylinders and reacted by heating a tungsten filament in either an argon, CO₂ or N₂ atmosphere. Reaction systems examined were: (1) 3CaO + P₂O₅ → Ca₃(PO₄)₂ (TCP) and (2) 4CaO + P₂O₅ → Ca4P₂O₇ + O₂ (TTCP). The product pellets were bright white with a thin glassy appearing outer layer observed with a very porous interior (range of 5–1000 μm pore diameter) regardless of reaction atmosphere. No evidence of sintering is apparent such as intact precursor particles or necking between particles. In the TCP system characteristic XRD peaks for crystalline β-, α-TCP, and HA are noted while in the TTCP system tetracalcium phosphate, α-TCP, and HA are present. DSC analysis indicates the ignition temperature 450°C when P₂O₅ decomposes to PO₃ to react with CaO. SHS is a viable method to manufacture heterogeneous calcium phosphates from CaO and P₂O₅ precursors. The formation of HA and TCP, regardless of stoichiometry or atmosphere is due to thermodynamic energies of the products while other phases are preserved due to limited kinetics in the system’s rapid heating and cooling.     

Phospholipid fatty acid composition in type I and type II rat muscle

The fatty acid composition of the membrane phospholipids phosphatidylcholine (PC) and phosphatidylethanolamine in insulin-sensitive Type I (soleus) and insulin-resistant Type II (EDL) muscle is not known. In the present studies, soleus and EDL muscles were removed from 250–300 g Sprague-Dawley rats, and the fatty acid composition of total and individual phospholipid (PL) species was quantitated. As expected, triglyceride content was increased twofold in soleus muscle. No quantitative differences in the individual PL species or cholesterol content were found between the two muscles. However, a striking difference in PL fatty acid composition was observed in the PC fraction. An increase in 16∶0 with decreases in 18∶0, 18∶1, 22∶5n-3, and 22∶6n-3 (P<0.001 for each) was observed in the PC fraction of EDL compared to that from soleus, consistent with reduced elongation of PC fatty acids. Inhibition of fatty acid oxidation with the carnitine palmitoyl transferase-1 inhibitor, etomoxir, did not alter the fatty acid pattern in either muscle. We conclude that an alteration in PL fatty acid composition consistent with reduced elongation of both saturated and unsaturated fatty acids is observed in Type II muscle. The restriction of these alterations to the PC fraction has important implications. Deceased (June 28, 1996).     

Effect of dietary palm oil and its fractions on rat plasma and high density lipoprotein lipids

Male Sprague Dawley rats were fed semipurified diets containing 20% fat for 15 weeks. The dietary fats were corn oil, soybean oil, palm oil, palm olein and palm stearin. No differences in the body and organ weights of rats fed the various diets were evident. Plasma cholesterol levels of rats fed soybean oil were significantly lower than those of rats fed corn oil, palm oil, palm olein or palm stearin. Significant differences between the plasma cholesterol content of rats fed corn oil and rats fed the three palm oils were not evident. HDL cholesterol was raised in rats fed the three palm oil diets compared to the rats fed either corn oil or soybean oil. The cholesterol-phospholipid molar ratio of rat platelets was not influenced by the dietary fat type. The formation of 6-keto-PGF_1α was significantly enhanced in palm oil-fed rats compared to all other dietary treatments. Fatty acid compositional changes in the plasma cholesterol esters and plasma triglycerides were diet regulated with significant differences between rats fed the polyunsaturated corn and soybean oil compared to the three palm oils.     

Stearic acid modifies very low density lipoprotein lipid composition and particle size differently from shorter-chain saturated fatty acids in cultured rat hepatocytes

Stearic acid as compared to myristate, palmitate, or oleate is poorly incorporated into triacylglycerol, a major lipid component of very low density lipoprotein (VLDL). The present study investigated the effects of these fatty acids on VLDL metabolism in cultured rat hepatocytes. All fatty acids stimulated [2-³H] glycerol incorporation into VLDL lipids and secretion of [³H]-labeled VLDL by hepatocytes. However, the rate of [³H]-labeled VLDL secretion in the presence of nonlabeled stearate (12.8±0.7 pmol/mg protein/4h) was 46, 59, and 22% of that observed for those treated with myristate, palmitate, and oleate, respectively. [1-¹⁴C]Stearate as a substrate was also less effective than other labeled fatty acids to be incorporated into VLDL lipids. Of total VLDL lipids synthesized from [1-¹⁴C] stearate, triacylglycerol accounted for 78% as compared to 88–97% of that derived from palmitate, myristate, and oleate. The amounts of apoB100 and apoB48 were the same in hepatocytes treated with or without exogenous fatty acids. Similarly, the rate of apoB synthesis from [³⁵S] methionine was not affected by exogenous fatty acids. The treatment of cells with various saturated fatty acids increased the particle size of VLDL to different extents. The largest particles of VLDL, with a mean diameter of 79.3±11.9 nm, were seen in the cells treated with stearate, followed by those treated with palmitate and myristate (45.5±9.8 and 38.6±6.8 nm, diameter, respectively). Clearly, hepatocytes treated with stearate secrete less VLDL and produce larger VLDL particles than those treated with shorter-chain saturated fatty acids.     

The effect of antioxidant deficiency on tissue lipid composition in the rat. I. Gastrocnemius and quadriceps muscle

The gastrocnemius and quadriceps muscle phospholipids of the antioxidant-deficient rat fed a source of both linoleate and linolenate showed a progressive net increase in arachidonate, a progressive net decrease in all other polyunsaturated fatty acids, and there was a concomitant accumulation of fluorescent pigment of the lipofuscin or ceroid type in the tissue. An increased incorporation of intraperitoneally injected, isotopically labeled acetate into not only arachidonate but also the other higher polyunsaturated fatty acids, was observed. The net loss of the higher polyunsaturated fatty acids from the membrane lipids (presumably via lipid peroxidation) apparently was partially compensated by a homeostatic mechanism which involved conversion of the available precursors, linoleate and linolenate, to the higher polyunsaturated fatty acids. The rates of decrease of the polyunsaturated fatty acids in the muscle phospholipids and accumulation of fluorescent pigment in the tissue were correlated with the rate of production of creatinuria.     

Effect of hypothyroidism on the lipid composition of rat plasma and erythrocyte membranes

The effect of hydrothyroidism on plasma and erythrocyte membrane lipid components has been investigated. This pathological state is accompanied by a) a cholesterol increase of about 60% in plasma, and at the same time a 22% reduction in erythroycte membranes; b) 44% and 30% phospholipid level decreases in both plasma and red cell membranes, respectively; and c) almost unaffected phospholipid and fatty acid compositions of both plasma and erythrocyte membranes. All changes were corrected by treatment of the hypothyroid rats with triiodothyronine for two days. These findings suggest that in hypothyroid rats a reduced transfer of cholesterol from plasma to erythrocyte membrane probably takes place. This could explain, at least in part, the increased hematic cholesterol level observed in hypothyroid animals. In red cell membranes, the simultaneous decrease in cholesterol and phospholipid levels does not alter the cholesterol/phospholipid molar ratio, thus avoiding their abnormal function.