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Insulin-like stimulation of glucose transport in isolated adipocytes by fatty acids.

Glucose transport rate as assessed with the 3-0-Methylglucose method was significantly increased in adipocytes preincubated with aliphatic carboxylic acids. The magnitude of stimulation apparently depended on the chain length of the carboxylic acid, and was highest with palmitic acid (130%). The stimulation was additive to the effect of insulin, and reflected a decrease of km rather than an increase in vmax of the transport rate. The results suggest that fatty acids may modulate the activity of the glucose transporter, providing an insulin-dependent supply of adipose tissue with glycerol-phosphate during lipolysis for reesterification of excess fatty acids. Further, it is suggested that fatty acids mediate the stimulatory effect of catecholamines on glucose transport as observed in isolated fat cells.

3-O-Methylglucose↗

Effect of fatty acids on herbicide transport across Caco-2 cell monolayers.

Oral ingestion of pesticides can be a major exposure route. These compounds are frequently consumed in the presence of triacylglycerides, which are then hydrolyzed to free fatty acids. The purpose of this work was to examine the effect of two common fatty acids, palmitic (PA) and oleic (OA) acids, and the biological emulsifier sodium taurocholate (TC) on the absorption of three herbicides (trifluralin, alachlor and atrazine) by Caco-2 cell monolayers. Trifluralin's absorption was enhanced (p < 0.05) in the presence of OA whereas the greatest absorption of atrazine and alachlor occurred with PA and the control media, respectively. Trifluralin had significantly lower absorption through the monolayer than either alachlor or atrazine (p < 0.001). A mass balance study demonstrated that trifluralin accumulated within the cell monolayer (13.85% of the donor after 3 h of exposure), but alachlor and atrazine (1.27% and 0.85%, respectively) did not. This response was linear with time (21.89% trifluralin after 6 h of exposure), and demonstrated the potential for continued release of trifluralin after source removal. These experiments demonstrated that fatty acids and an emulsifier can influence absorption of herbicides across small intestinal epithelium.

Acetamides↗

Arachidonic acid modulates [14C]stearic acid incorporation into phosphatidylinositol, in human neuroblastoma cells.

In the human neuroblastoma cell line SK-N-BE(2), arachidonic acid (AA), supplied in the medium at micromolar concentrations, markedly enhanced [14C]stearic acid (SA) (but not [14C]palmitic acid or [14C]oleic acid) incorporation into phosphatidylinositol (PtdIns). AA failed to stimulate [14C]SA incorporation into PtdIns precursors, namely phosphatidic acid and cytidinediphosphodiacylglycerol: furthermore, enhanced [14C]SA incorporation, brought about by exogenously administered AA, was not restricted to PtdIns tetraenoic species. When cells were pulsed for 1 h with [14C]SA (either in the presence or absence of AA) and then reincubated in AA- and [14C]SA-free medium, a marked loss of radioactivity from PtdIns was observed, that however was restricted to molecular species other than tetraenoic. These results are discussed in the light of possible mechanisms through which PtdIns achieves the 1-stearoyl-2-arachidonoyl configuration.

Arachidonic Acid↗

A simple method for the preparation of phosphatidylcholine labelled at 2-acyl position.

Phosphatidylcholine with a labelled acyl group at 2 position was prepared enzymatically from lysophosphatidylcholine and labelled fatty acids. Under optimal conditions, over 90% of [5,6,8,9,11,12,14,15-3H] arachidonic acid, [9,10-3H] palmitic acids, [1-14C] linoleic acid or [9,10-3H] oleic acid was incorporated into the phosphatidylcholine, and the incorporation was quantitative up to 500 nmol of [1-14C] linoleic acid. Analysis of the labelled phosphatidylcholine formed indicated that radioactivity was incorporated exclusively at 2-acyl position.

Animals↗

Acetyl-coenzyme-A carboxylase in cultured hepatocytes. Effects of exogenous fatty acids on the content, synthesis and degradation of the enzyme.

Studies were made on the content, synthesis and degradation of acetyl-coenzyme-A carboxylase in JTC-25 - P3 cells, hepatocytes which can be maintained in a protein-free and lipid-free chemically defined medium. The addition of corn oil or fatty acid to the medium resulted in a decrease in the activity level of the enzyme without impairing the viability of cells. All the fatty acids tested exhibited this effect, although linoleic acid and oleic acid were more effective than palmitic acid, stearic acid and arachidonic acid. Immunochemical titration and Ouchterlony double-diffusion analysis indicated that the decrease in the activity level of the enzyme observed in cells incubated in medium supplemented with fatty acid can be ascribed to a reduction of the quantity of the enzyme. Isotopic leucine incorporation studies with the use of immunochemical techniques demonstrated that this reduction of the enzyme content is due to a decrease in the rate of synthesis of the enzyme. The rate of degradation of the enzyme was essentially unaffected, the half-life being 25 and 28 h, respectively, in cells incubated in the presence and absence of fatty acid. It was shown that most of the isotopic fatty acid added to the medium was incorporated into cellular phospholipids, while a very small portion of it was recovered in triglyceride and nonesterified fatty acid.

Acetyl-CoA Carboxylase↗

Biological and physicochemical characteristics of four serotypes of Salmonella enteritidis.

Four serotypes of Salmonella enteritidis, Anatum ATCC 9270, Newbrunswick ATCC 1608, Oranienburg 200 E, and Pullorum RM, were studied to determine biological, chemical, or physical differences which might explain variations in Salmonella virulence as previously reported by McCullough and Eisele (J. Infect. Dis. 88:278-289, 1951; 89:259-265, 1951). These investigators found that serotype Pullorum was significantly less virulent than serotypes Newport, Derby, Barielly, Meleagridis and Anatum when fed to healthy humans. Results of our own experiments showed that serotype Pullorum RM had a generation time approximately twice that of serotype Anatum 9270. The volume of serotype Pullorum was approximately one-half the volume of the other serotypes used (Anatum 9270, Newbrunswick 1608, Oranienburg 200 E, Cubana 12007, and Meleagridis DR). The number of cells required to yield 1 g dry weight was substantially higher for serotype Pullorum RM than for serotypes Anatum 9270, Newbrunswick 1608, and Oranienburg 200 E. The yield of endotoxin per gram dry weight for serotype Pullorum RM averaged 22 mg/g, whereas yields of endotoxin for serotypes Anatum 9270, Newbrunswick 1608, and Oranienburg 200 E averaged 32 to 35 mg/g. The relative abundance of the four major fatty acids (measured by gas chromatography) also showed distinct differences among the serotypes. Pullorum RM contained less lauric and 3-hydroxymyristic acids and more myristic and palmitic acids than the other three serotypes. The identity of 3-hydroxymyristate was confirmed by mass spectroscopy. Serotype Pullorum RM required 10 times more lipopolysaccharides (endotoxin) to obtain a 50% lethal dose in mice than the other three serotypes. When the lipid part was separated from the polysaccharide and solubilized with bovine serum, the 50% lethal dose of serotype Pullorum RM was equal to that of the other three.

Endotoxins↗

Heptakis(2,6-O-dimethyl)beta-cyclodextrin: a novel growth stimulant for Bordetella pertussis phase I.

The effect of cyclodextrins on the growth of Bordetella pertussis Tohama phase I in synthetic medium was evaluated. The addition of cyclodextrins, especially heptakis(2,6-O-dimethyl)beta-cyclodextrin (Me beta CD), to a complete synthetic medium such as Stainer-Scholte medium gave the same number of individual colonies and growth rates as those on Bordet-Gengou medium. Furthermore, with the addition of Me beta CD, growth inhibition by fatty acids such as oleic or palmitic acid was overcome and normal cell growth was observed. This modified Stainer-Scholte medium, designated as cyclodextrin solid medium (CSM), supported excellent growth of 20 lyophilized clinical isolates. Serotypes of the organisms after 10 passages on this CSM plate were not changed. These results suggest that Me beta CD is a significant growth stimulant and CSM is one of the most suitable synthetic media for culture of B. pertussis phase I.

Antigens, Bacterial↗

[Analysis of physicochemical property and composition of fatty acid of almond oil].

Almond oil was extracted from almond and saponified with saturated KOH-methanol. The mixed fatty acids methylated by methanol-H2SO4(4:1, V/V). Methyl esters were extracted by ether and detected by gas chromatography. The major fatty acids in Taiyuan almond oil were found to be about 68% oleic acid (C18:1), 25% linoleic acid(C18:2), 4.6%-4.8% palmitic acid(C16:0) and a little of palmitoleic acid(C16:1), stearic acid(C18:0). A trace of arachidic acid(C20:0) was also found.

Chromatography, Gas↗

Lipid biosynthesis in Paramphistomum microbothrium (Trematoda).

In vitro studies with Paramphistomum microbothrium indicated that the trematode is capable of synthesizing its complex lipids using exogenous substrates. U-14C glucose and 2-14C acetate were predominantly incorporated into phospholipids while 1-14C oleic acid and U-14C palmitic acid appeared more in the neutral lipids. A large proportion of the labelled acetic acid incorporated into neutral lipids appeared in the triglycerides. P. microbothrium is capable of synthesizing its cholesterol de novo.

Acetates↗

Effect of leucine and alpha-ketoisocaproic acid on NAD biosynthesis from tryptophan or nicotinic acid in the isolated rat liver cells.

The effects of leucine and alpha-ketoisocaproic acid on the NAD biosynthesis from (carboxyl-14C) nicotinic acid or (benzene ring-U-14C) tryptophan were investigated in the isolated rat liver cells. The quantities of NAD formed from tryptophan and nicotinic acid increased linearly during 60 min-incubation. Leucine, which was essentially not metabolized by the isolated liver cells, had no inhibitory effect on the NAD synthesis. On the other hand, alpha-ketoisocaproic acid, alpha-keto acid analogue of leucine, which was rapidly metabolized to ketone bodies, diminished a quantity and a specific radioactivity of NAD synthesized from both tryptophan and nicotinic acid to a similar extent. Palmitic acid, which was oxidized to ketone bodies by the isolated rat liver cells, also diminished an amount and a specific radioactivity of NAD synthesized from nicotinic acid. Ketone bodies exogenously added to the incubation medium exerted a similar inhibitory effect on NAD synthesis. Results demonstrate that substances which can be oxidized to ketone bodies are inhibitory to NAD biosynthesis from tryptophan and nicotinic acid in the isolated rat liver cells.

Animals↗

[A simple method for the synthesis of phosphatidylcholine with the fatty acid substituted in the 2-position].

The preparation of PC with the 14C-fatty acids palmitic, stearic, oleic and linoleic acid at OH-group in position 2 is described. The starting material is egg yolk lecithin. By the attack of snake venom phospholiphase lyso-PC is produced which is reacylated by the appropriate fatty acid anhydride. In comparison with the methods published up to now this preparation has the advantage of higher yields and greater simplicity. By means of in vivo synthesis it is impossible to get PC species with only one fatty acid in a defined position. Working with radioactive fatty acids the specific radioactivity can be adapted to the requirements. The procedure can be made on a semi-technical scale.

Carbon Radioisotopes↗

Plasma palmitate turnover in subjects with thermal injury.

Using a continuous infusion of [1-13C] palmitic acid tracer, plasma palmitate turnover was measured 14 times in nine bandaged, thermally injured adults. Plasma glucose (102 +/- 4 mg/dl), insulin (21 +/- 4 microU/ml), and glucagon (296 +/- 34 pg/ml) levels were significantly elevated compared with values in uninjured controls. Circulating plasma epinephrine (67 +/- 11 pg/ml) and norepinephrine (219 +/- 57 pg/ml) levels were more than twofold their respective control values of 261 +/- 4 pg/ml and 211 +/- 7 pg/ml but less than the previously defined plasma threshold levels for lipolytic effects of these catecholamines as circulating hormones. Plasma palmitate and free fatty acid concentrations, 113 +/- 8 and 452 +/- 38 microM, respectively, were not different from control values but palmitate flux (2.66 +/- 0.28 mumol kg-1 min-1) and free fatty acid turnover calculated therefrom (10.53 +/- 1.13 mumol kg-1 min-1) were significantly elevated compared to the control rates. While palmitate turnover significantly correlated with plasma palmitate concentration and with per cent body surface area burned, there was no relationship between palmitate flux and circulating epinephrine or norepinephrine levels. These data raise new questions about the relative catabolic roles of catecholamines in bandaged, thermally-injured patients.

Adolescent↗

Gestational variation of fatty acid composition of human amniotic fluid lipids.

The amount of total lipids of the amniotic fluid of normal human subjects increases progressively with gestational age and is found to be maximum in labor at term. The lecithin/sphingomeylin ratio, which invariably increased with gestational age, is 2:1 or greater beyong 34 weeks of pregnancy and becomes maximum at term labor. No definite correlation exists between the palmitic acid content of total lipids and gestational age; but the palmitic acid content of the lecithin fraction increases with gestational age and this increase is markedly accelerated near the 34th week of pregnancy. Also it is observed that the arachidonic acid of cholesterol esters increases with gestational age. Therefore, it is suggested that both the palmitic acid level of lecithin and the arachidonic acid level of cholesterol esters might serve as more reliable indices that current methods being employed for the assessment of fetal lung maturity.

Amniotic Fluid↗

Studies in Tetrahymena membranes substrates for desaturation of fatty acyl chains in Tetrahymena pyriformis microsomes.

[1-14-C]Palmitoyl-Co A was incubated with Tetrahymena microsomes containing the complete enzyme system for desaturation during various time periods. The level of [1-14C]palmitoleoyl-CoA increased to a maximum during the 1--3 min incubation time, while [1-14C]palmitoleic acid in the phospholipid reached a maximum level during 6--7 min incubation time. The radioactivity of [1-14C]palmitoleic acid in free fatty acid and the triglyceride fraction was not significantly observed upon 3 min incubation. Incubation of [1-14C]palmitoyl-CoA with microsomes in the absence of NADH produced [1-14C]palmitoyl lipid without desaturation. Radioactive palmitic acids in the microsomal lipids were not converted to palmitoleic acids after addition of NADH by the complete enzyme system. When microsomes prepared from cells labeled with [1-14C]palmitic acid or [1-14C]stearic acid were incubated alone in the presence of O2 and NADH, no significant increase in [1-14C]palmitoleic acid in the phospholipid was observed, wherease an increase in [1-14C]linoleic acid and gamma-[1-14C]linolenic acid did occur at the expense of [1-14C]oleic acid in the phospholipid. From these results it can be concluded that the enzyme involving desaturation of palmitic acid to palmitoleic acid requires palmitoyl-CoA as the substrate. However, the possibility of oleoyl and linoleoyl phospholipids being substrates in the desaturation of Tetrahymena microsomes was suggested.

Acyl Coenzyme A↗

Water-soluble fatty acid derivatives as acylating agents for reversible lipidization of polypeptides.

A novel method allowing the conjugation of a fatty acid to a peptide or protein in aqueous buffer is described in this paper. L-Cysteinyl 2-pyridyl disulfide (CPD) (III), which was obtained by reacting L-cysteine (I) with 2,2-dithiopyridine (II), was reacted with the N-hydroxysuccinimide ester of palmitic acid (IV) to yield a water-soluble derivative of palmitic acid, termed Pal-CPD (V). Pal-CPD (V) could be reacted with a sulfhydryl-containing peptide or protein in aqueous buffer to yield the palmitic acid-derivatized conjugate (VI). The palmitic acid-derivatized Bowman-Birk protease inhibitor (BBI), synthesized using this conjugation method, was demonstrated to have 140-fold higher uptake into Caco-2 cell monolayers compared to native-BBI. The biological activity of the conjugate, as assessed using an in vitro transformation assay, was retained.

Acylation↗

Phospholipid synthesis in isolated porcine gastric mucous cells.

Phosphatidylcholine (PC) and phosphatidylethanolamine (PE) are the major phospholipids of the gastric mucosal surface barrier and chiefly originate from mucous cells. Among these phospholipids PC with palmitic acid as its hydrophobic moieties is believed to protect the gastric mucosa by its hydrophobic properties. We investigated the phospholipid synthesis of isolated porcine gastric mucous cells in vitro and incubated them in the presence of radiolabelled precursors. Incorporation of 3H-choline into PC and of 14C-ethanolamine into PE was linear at 1, 10, and 100 mumol/l substrate concentration for at least 6 h. Half-maximal rate of precursor incorporation was achieved at 21 and 15 mumol/l of choline and ethanolamine, respectively. Ethanolamine inhibited PC synthesis and choline inhibited PE synthesis. A small amount of radioactivity originating from 14C-ethanolamine and from the methyl groups of 3H-methionine were incorporated into PC. Palmitic acid was incorporated into PC more than PE. Indomethacin did not influence the de novo synthesis of PC and PE via the Kennedy pathway, but inhibited the incorporation of 3H-methionine into PC. These results indicate that in gastric mucous cells PC and PE synthesis de novo depends on the concentrations of choline and ethanolamine. The palmitic acid content of PC depends on the availability of palmitic acid as a substrate: indomethacin-induced mucosal damage is not explained by modulation of phospholipid synthesis de novo.

Animals↗

Selective partitioning of dietary fatty acids into the VLDL TG pool in the early postprandial period.

Circulating triacylglycerol (TG) arises mainly from dietary fat. However, little is known about the entry of dietary fat into the major TG pool, very low-density lipoprotein (VLDL) TG. We used a novel method to study the specific incorporation of dietary fatty acids into postprandial VLDL TG in humans. Eight healthy volunteers (age 25.4 +/- 2.2 years, body mass index 22.1 +/- 2.3 kg/m2) were fed a mixed meal containing 30 g fish oil and 600 mg [1-13C]palmitic acid. Chylomicrons and VLDL were separated using immunoaffinity against apolipoprotein B-100. The fatty acid composition of lipoproteins was analyzed by gas chromatography/mass spectrometry. [1-13C]palmitic acid started to appear in VLDL TG 3 h after meal intake, and a similar delay was observed for eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Approximately 20% of dietary fatty acids entered the VLDL TG pool 6 h after meal intake. DHA was clearly overincorporated into this pool compared with [1-13C]palmitic acid and EPA. This seemed to depend on a marked elevation of this fatty acid in the nonesterified fatty acid pool. In summary, the contribution of dietary fatty acids to early postprandial VLDL TG is substantial. The role of DHA in VLDL TG production will require further investigation.

Adult↗

Plasma and cellular zinc levels and membrane lipid composition in streptozotocin diabetic rats.

1. Lipid and zinc analyses were conducted on liver mitochondrial and microsomal membranes as well as erythrocyte ghosts from streptozotocin (STZ) treated animals. 2. In STZ animals, analysis of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) fatty acids revealed an increase in palmitic acid and a corresponding decrease in stearic acid. 3. Polyunsaturated fatty acids were also affected, with an increase in 18:2, decrease in 20:4 and an increase in 22:6 in STZ animals compared to controls. 4. The change in fatty acid composition was observed in all three membrane fractions. 5. Plasma zinc levels in STZ animals were elevated while no difference was observed in membrane bound zinc. 6. Thus, while there appears to be both altered trace metal as well as membrane lipid metabolism in STZ treated animals, membrane bound zinc does not seem to be affected.

Animals↗