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PubMed1977

[Histochemical, analytical and thin layer chromatographical studies on bovine seminal vesicle lipids (author's transl)].

The lipids in the epithelium of seminal vesicles of 11 adult bulls were studied by means of histochemical methods and thin layer chromatography. In the large basal lipid vacuoles dominate cholesterol, esters of cholesterol and tryglycerides. Phosphatides are also present in the large basal lipid vacuoles as well as in the smaller apical lipid droplets of the columnar cells. The existence of alkyl diglycerides, neutral plasmalogens and methyl esters of fatty acids, proved by 2-dimensional chromatography, can be taken as evidence that fatty acids, cholesterol, triglycerides and phosphatides are synthesized within the seminal vesicle epithelial cells. Free fatty acids as well as those incorporated in mono-, di-, triglycerides and polar lipids of the seminal vesicle epithelium are predominantly unsaturated, thus pointing to rapid mobilization and turnover of their esters. The cholesterol in the basal lipid vacuoles of bovine seminal vesicle is possibly eliminated from the epithelium via the subepithelial capillaries for there is no evidence of lipid secretion into the glandular lumen nor of a continued steroid synthesis from cholesterol within the seminal vesicle.

Animals↗
PubMed1985

Protein phosphorylation and platelet secretion.

Platelet secretion in response to physiologic stimuli appears to result from the complementary stimulation of two processes--granule centralization and granule membrane fusion. Granule centralization is produced by actin-myosin contraction which is initiated by a movement of calcium ions into the cytoplasm. The calcium binds to calmodulin to form a complex which activates myosin light chain kinase to phosphorylate myosin light chain (MLC). Once phosphorylated in this fashion, actin-myosin contraction occurs. Granule membrane fusion can be produced selectively by phorbol myristate acetate and oleoyl-acetyl diglyceride, both of which activate protein kinase C. Phosphorylation of a 47,000 dalton intracellular protein (47K) by protein kinase C may be critical to granule membrane fusion. The mechanism of action of 47K is presently unknown. The combined phosphorylation of MLC and 47K in response to most physiologic agonists which cause granule secretion, and the synergistic effects on granule secretion of agents which independently stimulate MLC and 47K phosphorylation, suggests secretion usually results from the interaction of granule centralization and granule membrane fusion.

Actin Cytoskeleton↗
PubMed1985

A radioisotopic method for the measurement of free fatty acid turnover and adrenoceptor response in small fragments of human adipose tissue.

New techniques for measuring rates of triglyceride hydrolysis and the re-esterification of resultant free fatty acids (FFA) in small fragments of human adipose tissue are described. Buffered medium containing [14C]-glucose and [3H]-palmitate of high specific activity is used to label newly synthesized acylglycerides in incubated tissue. 14C is incorporated only into the glyceride 'back bone' of acylglycerides, and the rate of incorporation may be used as a measure of the rate of synthesis of each glyceride molecular species. Due to mixing of exogenous [3H]-palmitate with endogenously released FFA, the actual specific activity of FFA esterified to [14C]-glycerides is always less than that of the [3H]-palmitate in the medium. The degree of dilution of [3H]-palmitate with unlabelled FFA is directly proportional to the rate of lipolysis. Thus, determination of the 14C/3H ratio in newly synthesized triglycerides permits the assessment of adrenoceptor dose-response characteristics (lipolysis) of unweighed fragments of adipose tissue. If rates of 14C-tri- and diglyceride accumulation are measured in tissue fragments whose cellularity and rate of glycerol release has been determined, the fraction of newly lipolyzed FFA which are re-esterified can be calculated. This 'cycle' of re-esterification controls the ratio of FFA: glycerol leaving the adipocyte. These techniques can be applied to serial subcutaneous adipose tissue samples from awake subjects in various clinical and experimental states.

Adipose Tissue↗
PubMed1985

Immunogenic properties of glycolipids of Nocardiopsis dassonvillei.

Two specific glycolipids identified as monomannosyl diglyceride (G1) and monoacylated glucose (G2), isolated from Nocardiopsis dassonvillei strains, were found to be biologically active. They elicited antibody response in rabbits when administered with some carrier components: lecithine, methylated bovine serum albumin (MBSA) and Freund incomplete adjuvant (FIA). Antibodies to the glycolipids were detected in antisera to G1 and in the sera against crude cell antigen of N. dassonvillei (N). The ability of the lipids to induceskin delayed hypersensitivity (SDH) as well as adjuvant and mitogenic effect were also studied. Generally glycolipid G1 seemed to be more active in antibody response whereas glycolipid G2 was a better stimulator of cellular response. Glycolipid G2 appeared to be distinct adjuvant and mitogenic factor.

Actinomycetales↗
PubMed1967

Increase in cell lipid and cytoplasmic particles in mammalian cells cultured at reduced pH.

The hydrogen ion concentration of the medium has been shown to exert a regulatory effect on the lipid content of cultured mammalian cells. Reduction of the pH of the medium from 7.4 to 6.9 causes a significant increase in cell lipid, relative to cell protein, within 2-3 days. Triglycerides are increased twofold and account for 75% of the additional lipid. Polar lipids, on the other hand, remain nearly constant in concentration. Concurrent with the increase in lipid, particles with an average diameter of 1 micro appear in the cytoplasm. Because the density of these particles is low, ultracentrifugation of the cell homogenate separates the particles completely from the other subcellular structures. The amount of lipid in the particle fraction is approximately equal to the increase in total cell lipid. As shown by silicic acid column chromatography, the particle lipid contains about 75% triglycerides, 15% diglycerides plus an unknown substance, and smaller amounts of material in the monoglyceride and sterol ester-hydrocarbon fractions. The quantitative results indicate that the lipid accumulated at low pH is assembled into discrete cytoplasmic particles.

Animals↗
PubMed1969

Synthesis of lipids from acetate by human preputial and abdominal skin in vitro.

Lipogenesis in vitro from acetate-1-(14)C was studied in human preputial skin and abdominal skin. Radioactive lipids were separated by column chromatography on Florisil and by thin-layer chromatography on silica gel. Radioactivity was incorporated chiefly into the triglyceride, sterol, and polar lipid fractions, while lesser amounts of (14)C were found in the hydrocarbon, wax, diglyceride, monoglyceride, and fatty acid fractions; labeling of steryl esters was minimal. On thin-layer chromatography, the radioactive polar lipids had mobilities similar to lysolecithin, phosphatidyl choline, phosphatidyl ethanolamine, and phosphatidic acid. The radioactive fatty acids of the different lipid fractions were separated by gas-liquid chromatography. The major (14)C-labeled acids were 16:0 and 18:0. Radioactivity was also detected in acids 14:0, 15:0, 16:1, 18:1, 18:2, 20:0, 20:1, 22:0, 24:0, 24:1, and 26:0. No radioactivity could be detected in arachidonic acid, although this fatty acid comprises 9% of the chromatographed fatty acids. The pattern of incorporated (14)C was different from the percentage mass composition of the fatty acids. Skin is therefore active in the biosynthesis of a wider variety of lipids than previously demonstrated.

Abdomen↗
PubMed1971

Studies on the hormone-sensitive lipase of adipose tissue.

Sucrose gradient centrifugation has been used to examine the triglyceride lipases present in extracts of rat epididymal adipose tissue. The aqueous infranatant recovered between the pellet and fat cake of tissue homogenates which had been centrifuged at 40,000 g was shown to contain two types of triglyceride lipase activity. One of these appears in the 15s region and has been identified as the active form of the "hormone-sensitive lipase" believed to be responsible for initiating the hydrolysis of tissue triglyceride stores in response to lipolytic stimuli. The activity of this enzyme was selectively increased in extracts prepared from tissue exposed to epinephrine and decreased in extracts of insulin-treated tissue. The increased lipolytic activity of extracts of tissue from fasted or fasted-refed rats was also found largely in this region. When the tissue was incubated with orthophosphate-(32)P, radioactivity was incorporated into a protein migrating at 15s. A second peak of triglyceride lipase activity appeared in the 6s region coincident with the location of the monoglyceride and diglyceride lipase activities. The amount of 6s triglyceride lipase activity did not correlate with changes in the lipolytic activity of the tissue from which the extracts were prepared, and its physiological function remains to be elucidated. The lipoprotein lipase and the short-chain triglyceride lipase ("tributyrinase") each moved more slowly in the gradient than the 6s triglyceride lipase. Both the 6s and 15s enzymes were shown to be present in washed adipocytes isolated from the tissue by collagenase digestion.

Adipose Tissue↗
PubMed1974

Reversible protein kinase activation of hormone-sensitive lipase from chicken adipose tissue.

Hormone-sensitive lipase partially purified from adipose tissue of laying hens was markedly activated by cyclic AMP-dependent protein kinase. Activation was approximately 4-fold (ranging up to as great as 10-fold) compared with the much lower degree of activation obtained with analogous preparations from rat and human adipose tissues (59 and 86%, respectively). The partially purified preparations contained adequate endogenous protein kinase activity to effect complete activation with addition of cyclic AMP, ATP, and Mg(2+). Activation was blocked by protein kinase inhibitor (from rabbit skeletal muscle) but could be restored fully by addition of excess exogenous protein kinase (from bovine skeletal muscle). The fully activated lipase was slowly deactivated by dialysis at 4 degrees C and then rapidly and almost fully reactivated by addition of cyclic AMP and ATP-Mg(2+). Reactivation was blocked by protein kinase inhibitor. This deactivation-reactivation cycle was rapid at 23 degrees C with dialysis against charcoal and could be demonstrated repeatedly using a single preparation. The reversible deactivation of protein kinase-activated enzyme is presumed to reflect the action of a lipase phosphatase. Lipase prepared from tissue previously exposed to glucagon yielded a much smaller degree of activation than lipase prepared from tissue not exposed to the lipolytic hormone, indicating that the physiological hormone-induced activation is probably similar to or identical with the protein kinase activation demonstrated in the cell-free preparations. Under the conditions of assay used, the partially purified lipase fraction contained diglyceride, monoglyceride, and lipoprotein lipase activities. However, treatment with cyclic AMP-dependent protein kinase had virtually no effect on these lipase activities.

Adenosine Triphosphate↗
PubMed1979

Interactions of C-reactive protein and complement with liposomes. II. Influence of membrane composition.

We found previoulsy that interaction of C-reactive protein (CRP) with liposomal model membranes resulted in complement(C)-dependent membrane damage. In the present study, we investigated the influence of membrane composition on the interactions of CRP and C with liposomes. Adsorption experiments showed that binding of CRP was greatest to strongly positive liposomes. A lesser, but still substantial, extent of CRP binding also was observed with negative liposomes, but negligible amounts of CRP bound to neutral or weakly positive liposomes. CRP-mediated consumption of hemolytic C, and C-dependent glucose release from liposomes both were strongly influenced by liposomal charge, positive being superior to negative. Glucose release and, to a lesser extent, consumption of hemolytic C were inversely related to phospholipid fatty acyl chain length. Phospholipid fatty acyl unsaturation and liposomal cholesterol concentration both had strong influences on C consumption and glucose release. The data suggest that CRP-mediated C consumption and membrane damage require an optimum membrane fluidity. Complement damage in the presence of CRP was enhanced by certain sphingolipids and also by digalactosyl diglyceride, but not by sphingomyelin. Our results thus demonstrate that CRP-mediated C consumption and C-dependent membrane damage both are influenced by the liposomal membrane composition.

Adsorption↗
PubMed1972

Uptake and metabolism of circulating chylomicron triglyceride by rabbit aorta.

To determine if chylomicron triglycerides are taken up and metabolized by the arterial wall, rabbit abdominal aortas were perfused in situ for various times up to 2 hr with blood-buffer containing isotopically labeled substrates. Labeled chylomicrons were obtained by feeding [(3)H]palmitic acid or [(3)H]glyceryl trioleate to rats and rabbits with cannulated thoracic ducts. After aortic perfusion with these chylomicrons, more than 85% of aortic lipid ester radioactivity was in triglyceride; when labeled glycerol or palmitic acid was perfused, most aortic ester lipid radioactivity was in diglycerides and phospholipids. This indicated that, during perfusion with chylomicrons, intact triglyceride molecules were taken up by aorta. The rate of triglyceride fatty acid uptake by the inner avascular segment approached maximal values at low concentrations of perfusate triglyceride fatty acids (2 mm), whereas uptake in the outer capillary perfused segment increased with increasing triglyceride fatty acid concentration (0.4-25 mm). By double-radioisotope techniques it was shown that aortic free fatty acid was derived from both perfusate free fatty acids and from hydrolysis of lipoprotein glycerides within the aortic wall. Uptake of chylomicron triglyceride by perfused aorta was independent of triglyceride hydrolysis, which was quantitatively small.

Animals↗
PubMed1972

Hydrolysis of glyceryl tri(1- 14 C)octanoate and glyceryl tri(1- 14 C)oleate monolayers by postheparin lipolytic activity.

The hydrolysis of monomolecular films of glyceryl tri[1-(14)C]octanoate and glyceryl tri[1-(14)C]oleate has been demonstrated by measurement of the decrease in surface radio-activity that occurs in the presence of postheparin plasma. The hydrolysis displayed first order kinetics and was proportional to enzyme concentration over a 10-fold range. No hydrolysis was observed in the absence of enzyme, and only slight activity (1%) was found in plasma taken from subjects before heparin administration. The hydrolysis was stimulated to a variable extent by Ca(2+). The first product of hydrolysis of the monolayer was identified as 1,2-diglyceride, which was subsequently converted to 2-monoglyceride. Inhibition of triglyceride hydrolysis was observed when postheparin plasma was preincubated in 2 m NaCl, 10(-4) m protamine, 10 mm Na(4)P(2)O(7), and 0.1 m NaF. Monolayer techniques avoid some but not all of the problems associated with emulsified lipid substrates and appear to be applicable for study of post-heparin lipolytic activities.

Carbon Isotopes↗
PubMed1974

Incorporation of (2-3H)glycerol into rat brain 1,2-diacyl-sn-glycero-3-phosphorylcholine and 1,2-diacyl-sn-glycerol molecular species in vivo.

Rat brain 1,2-diacyl-sn-glycerols (diglycerides) and 1,2-diacyl-sn-glycerols obtained from 1,2-diacyl-sn-glycero-3-phosphorylcholine after treatment with phospholipase C differ markedly in carbon number distribution. 70% of the 1,2-diacyl-sn-glycerols had a total of 38 fatty acid carbon atoms, and there was no detectable change in the 1,2-diacyl-sn-glycerol mass pattern between 7 and 23 days of age. In contrast, 1,2-diacyl-sn-glycero-3-phosphorylcholine contained at most 10% of this molecular species in the brains of rats of comparable age. A small increase in the C(36) species of 1,2-diacyl-sn-glycero-3-phosphorylcholine, which is associated with myelination, was noted between 10 and 17 days. The incorporation of intracranially injected [2-(3)H]glycerol into 1,2-diacyl-sn-glycero-3-phosphoryl-choline species with polyunsaturated fatty acids containing 20 or 22 carbon atoms was greater than into the species containing only saturated and/or monoenoic fatty acids between 30 min and 24 hr. The 1,2-diacyl-sn-glycerol fractions containing polyunsaturated fatty acids had the lowest specific activity at 30 min. The specific activity of the particular 1,2-diacyl-sn-glycerol fraction containing the stearate-arachidonate pair is the lowest for 4 hr after intracranial injection of the isotope. Thus, molecular species of 1,2-diacyl-sn-glycerol and 1,2-diacyl-sn-glycero-3-phosphorylcholine differed considerably in their labeling patterns, and a direct precursor-product relationship could not be demonstrated during the time period studied.

Animals↗
PubMed1968

Cytidine triphosphate: phosphatidic acid cytidyltransferase in Escherichia coli.

An enzyme has been found in particulate fractions of Escherichia coli that catalyzes the incorporation of cytidine triphosphate (CTP) into lipid in the presence of exogenous phosphatidic acid and Mg(++). The product has been identified enzymatically and by chromatography as cytidine diphosphate diglyceride. The reaction is optimal at a pH of 6.5 and Mg(++) concentration of 5-10 mm. The apparent K(m) for CTP is 7 x 10(-4)M and for phosphatidic acid, 2 x 10(-3)M. The reaction rate falls off rapidly with time and ceases entirely after 1 hr as the result of inactivation of the system by Mg(++).

Cytosine Nucleotides↗
PubMed1972

Intracellular sites of lipid synthesis and the biogenesis of mitochondria.

Experimental data are presented on the intracellular localization in rat liver of three enzymes which are involved in the biosynthesis of phosphatidylethanolamine and diphosphatidylglycerol. These enzymes are phosphatidylserine decarboxylase, CDP-diglyceride-l-alpha-glycerophosphate phosphatidyl transferase, and phosphatidylethanolamine-l-serine phosphatidyl transferase. It was found that the first two enzymes are primarily mitochondrial while the latter enzyme is primarily microsomal. The intracellular sites for the biosynthesis of phosphatidylcholine, phosphatidylethanolamine, and diphosphatidylglycerol are discussed, and the implications of their sites of biosynthesis on the assembly processes involved in the biogenesis of mitochondria are considered.

Amino Alcohols↗
PubMed1979

[Fatty acid composition of glycerolipids from potato leaves].

A method for rapid isolation of glyco- and phospholipids from potato leaves by a two-fold separation in a thin layer of silica gel is described. Using gas-liquid chromatography, the fatty acid compositions of monogalactosyldiglyceride, digalactosyldiglyceride, sulfolipid, phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl glycerol, phosphatidyl inositol, diphosphatidyl glycerol, phosphatidic acid and non-identified lipid from potato leaves were determined. The monogalactosyl diglyceride was found to contain up to 25% of 7,10,13-hexadecatrienic acid. Trans-3-hexadecenic acid as well as phosphatidyl glycerol is a constituent component of phosphatidic acid, diphosphatidyl glycerol and the non-identified lipid.

Chromatography, Gas↗
PubMed1979

[Blood serum lipoprotein spectrum in myocardial infarct].

The lipoprotein spectrum was studied in polyacrylamide gel and by thin-layer chromatography on silica gel in 236 patients with myocardial infarction in different periods of the disease, in 45 patients with unstable angina pectoris, in 75 healthy persons and in 8 dogs with experimental myocardial infarction. Besides an increase in the concentration of cholesterol and triglycerides and a decrease in the level of diglycerides in blood serum, there is a disturbance in the ratio of phospholipid fractions, mainly in myocardial infarction. The level of pre-beta-lipoproteins was increased in the acute period of myocardial infarction, the level of beta-lipoproteins in the subacute period. Complications and concomitant inflammatory diseases were attended by a higher level of beta-lipoproteins. A connection was noted between the lipoprotein spectrum, predominantly of the pre-beta-fraction, and the ABO phenotype and the season.

ABO Blood-Group System↗
PubMed1969

Improved quantitation of plasma lipids by direct gas-liquid chromatography.

A preliminary digestion of total plasma lipid extracts with phospholipase C, which converts the lysolecithins, lecithins, and sphingomyelins into monoglycerides, diglycerides, and ceramides, respectively, has been shown to facilitate subsequent determination of the plasma lipids by gas-liquid chromatography. A further improvement in the chromatographic elution pattern results from acetylation or trimethylsilylation of the liberated alcohol moieties prior to injection into the chromatograph. If tridecanoin is used as internal standard, quantitative estimates can be rapidly obtained for plasma lysolecithins, free cholesterol, lecithins, sphingomyelins, cholesteryl esters, and triglycerides, as well as for free fatty acids. Other plasma lipids do not occur in sufficiently high concentrations to interfere with the analysis. The determination requires 0.1-0.5 ml of plasma and about 6 hr of processing, but many samples can be processed at a time.

Acetates↗
PubMed1970

Cuticular lipids of adults and puparia of the Australian sheep blowfly Lucilia cuprina (Wied.).

The presence of a strong contact component in the sex and ovipositing behavior of the sheep blowfly Lucilia cuprina Wied. prompted an investigation into the chemical composition of the cuticular wax of the adult male and female flies as well as that of the blowfly puparia. Thin-layer chromatography indicated that the lipids in all the waxes examined comprise hydrocarbons, nonglyceryl esters, triglycerides, free fatty acids, and hydroxy compounds, probably diglycerides and monoglycerides. Phospholipids were not detected. Straight-and branched-chain saturated compounds, the latter often pre-dominating, are present in the hydrocarbon, free fatty acid, and ester fractions. Unsaturated molecules were absent. The hydrocarbons resemble those of the cricket to some extent, but the absence of unsaturated compounds is in striking contrast to both the cricket and the cockroach. Pheromones may be present in the low molecular weight fatty acids obtained on brief extraction of the insects.

Animals↗