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Neural crest cells (NCCs) have been identified as an important target population relative to ethanol-induced teratogenicity in both mouse and avian models. Additionally, whole embryo culture mouse models have shown strain-related differences in sensitivity to ethanol-induced damage following acute exposure during early NCC development. That differential sensitivity of NCCs may contribute to these strain differences has been unexplored. For this purpose, cultured NCCs from an inbred mouse strain (C57BL/6J; C57) that is more sensitive to ethanol-induced teratogenicity than an outbred strain (ICR) were compared. This study showed that the incidence of cell death was significantly higher for the C57 NCCs than those from the ICR strain at all ethanol concentrations tested, and as early as 12 hours after initial exposure to 100 mM ethanol. The lateral mobility of the membrane lipids was faster and the membrane GM1 content was lower in C57 cells than ICR cells both under control conditions and at all doses and times tested. Ethanol exposure resulted in significant increases in the membrane lipid lateral mobility, and decreases in the membrane GM1 content that occurred in a dose and time-dependent manner in the NCCs from both strains. A significant correlation was found between the GM1 content and lateral mobility of the membrane lipids, the lateral mobility of membrane lipids and cell viability, as well as the GM1 content and cell viability in the NCCs from both strains. These results suggest that different strain sensitivities to ethanol-induced teratogencity may lie, at least in part, in the interstrain differential response of the NCC population and that the vulnerability of the NCCs to ethanol-induced death may be related to their endogenous membrane GM1 content.
Platelet-activating factor (PAF; sn-1-O-alkyl-2-acetyl-sn-glycero-3- phosphocholine) is thought to be an important mediator of embryo-endometrial interactions in early pregnancy, and an understanding of its role in the establishment of early human pregnancy can only follow an understanding of its mechanism of action. In a human endometrial epithelial cell line, HEC-1B, the presence of mRNA encoding the platelet-activating factor receptor was demonstrated by reverse transcription-polymerase chain reaction. The presence of functional receptors was shown by inositol trisphosphate accumulation and a rise in the concentration of intracellular free calcium evoked by platelet-activating factor in myo-[2-3H]inositol-labelled and fura-2-loaded cells, respectively. Platelet-activating factor evoked rapid and concentration-dependent increases in the concentration of intracellular free calcium and inositol trisphosphate that were inhibited by the platelet-activating factor receptor antagonist WEB 2086, indicating that the responses are receptor mediated. Inositol trisphosphate accumulation evoked by platelet-activating factor was unaffected by pretreatment with pertussis toxin. Platelet-activating factor also stimulated the tyrosine phosphorylation of at least two major proteins of 80 kDa and 44 kDa; the smaller protein is an isoform of mitogen-activated protein kinase. These results show that functional platelet-activating factor receptors are located on the endometrial epithelial cell line HEC-1B and are linked to inositol lipid hydrolysis, calcium mobilization and tyrosine kinase activity.
Oilseed germination is characterized by the mobilization of storage lipids as a carbon source for the germinating seedling. In spite of the importance of lipid mobilization, its mechanism is only partially understood. Recent data suggest that a novel degradation mechanism is initiated by a 13-lipoxygenase during germination, using esterified fatty acids specifically as substrates. This 13-lipoxygenase reaction leads to a transient accumulation of ester lipid hydroperoxides in the storage lipids, and the corresponding oxygenated fatty acid moieties are preferentially removed by specific lipases. The free hydroperoxy fatty acids are subsequently reduced to their hydroxy derivatives, which might in turn undergo beta-oxidation.
The lateral mobility of plasma membrane lipids was analyzed during first cleavage of Xenopus laevis eggs by fluorescence photobleaching recovery (FPR) measurements, using the lipid analogs 5-(N-hexadecanoyl)aminofluorescein ("HEDAF") and 5-(N-tetradecanoyl)aminofluorescein ("TEDAF") as probes. The preexisting plasma membrane of the animal side showed an inhomogeneous, dotted fluorescence pattern after labeling and the lateral mobility of both probes used was below the detection limits of the FPR method (D much less than 10(-10) cm2/sec). In contrast, the preexisting plasma membrane of the vegetal side exhibited homogeneous fluorescence and the lateral diffusion coefficient of both probes used was relatively high (HEDAF, D = 2.8 X 10(-8) cm2/sec; TEDAF, D = 2.4 X 10(-8) cm2/sec). In the cleaving egg visible transfer of HEDAF or TEDAF from prelabeled plasma membrane to the new membrane in the furrow did not occur, even on the vegetal side. Upon labeling during cleavage, however, the new membrane was uniformly labeled and both probes were mobile, as in the vegetal preexisting plasma membrane. These data show that the membrane of the dividing Xenopus egg comprises three macrodomains: (i) the animal preexisting plasma membrane; (ii) the vegetal preexisting plasma membrane; (iii) the new furrow membrane.
The floating membrane vesicle is fixed by the counter solution flow in different points along the radius of a cylinder electrophoretic chamber, which permits to measure the vesicle electrophoretic mobility (EM). Close state condition of the chamber is provided for by the capillary system successively connected with the chamber. Relationship between EM of bimolecular lipid membranes (BLM) and pH and ionic concentration of aqueous solution qualitatively coincides with similar relationship for liposomes. The EM value of BLM essentially decreases in solution containing polyene antibiotics nystatine and levorin when derivative of cholesterol having 3betaOH-groups is present in the membrane.
In this paper we review the current status of research on fatty acid absorption and conversion to diacylglycerol in the midgut. We further discuss how diacylglycerol may leave the midgut and associate with lipophorin in hemolymph. We review the present understanding of the role of the lipid transfer particle and lipophorin receptors in lipid delivery between lipophorin and tissues. Finally, we discuss recent studies on the mobilization of diacylglycerol from the fat body in response to adipokinetic hormone. Several suggestions for exciting areas of future research are described.
The time course of the accumulation of triacylglycerols (TAGs) in Rhodococcus opacus PD630 or of TAGs plus polyhydroxyalkanoates (PHA) in Rhodococcus ruber NCIMB 40126 with gluconate or glucose as carbon source, respectively, was studied. In addition, we examined the mobilization of these storage compounds in the absence of a carbon source. R. opacus accumulated TAGs only after the exhaustion of ammonium in the medium, and, with a fixed concentration of the carbon source, the amounts of TAGs in the cells increased with decreasing concentrations of ammonium in the medium. When these cells were incubated in the absence of an additional carbon source, about 90% of these TAGs were mobilized and used as endogenous carbon source, particularly if ammonium was available. R. ruber accumulated a copolyester consisting of 3-hydroxybutyrate and 3-hydroxyvalerate already during the early exponential growth phase, whereas TAGs were synthesized and accumulated mainly during the late exponential and stationary growth phases. In the stationary growth phase, synthesis of TAGs continued, whereas PHA was partially mobilized. In the absence of an additional carbon source but in the presence of ammonium, mobilization of TAGs started first and was then paralleled by the mobilization of PHA, resulting in an approximately 90% and 80% decrease of these storage compounds, respectively. During the accumulation phase, interesting shifts in the composition of the two storage compounds occurred, indicating that the substrates of the PHA synthase and the TAG synthesizing enzymes were provided to varying extents, depending on whether the cells were in the early or late exponential or in the stationary growth phase.
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The rat pericoronary adipose tissue was perfused in the presence of either the liposynthetic hormone insulin or the lipolytic hormone noradrenaline. Insulin perfusion associated with a) larger adipocyte mean sectional diameter in comparison with noradrenaline perfusion; b) glycogen deposition; c) appearance of small fat globules at discrete sites at the periphery of the main lipid drop. The two latter phenomena were apparently dose-dependent. Massive lipid deposition was induced by addition of triglycerides to the perfusion medium and this associated with appearance of prominent endoplasmic reticulum in the cytoplasm. In noradrenaline-perfused adipose tissue many small lipid droplets surrounded the central lipid deposit and the endoplasmic reticulum was in the form of both thin long, dashed cisternae sometime surrounding lipid droplets and grouped, anastomosing tubular cisternae. The present work shows that the perfused white adipose tissue of the heart is a suitable model to study, in situ, the morphological effects of hormones in adipocytes.
Three different nitroxides were used to probe either the head group (Tempil stearate) or acyl chain region (Spin labeled cholestane (ChSl) and methyl ester of 5 doxyl palmitate (MeFASL(10,3))) of human plasma low density lipoproteins (LDL) and very low density lipoproteins (VLDL). The ESR data were compared with the simulated spectra which assume rapid anisotropic motion of nitroxide. The results indicate that in the head group region of both LDL and VLDL only the slowing down of the rotational motion occurred when temperature was lowered and the whole region showed up as a unique compartment. On the other hand, the acyl chain region, probed with MeFASL(10,3), behaved as one compartment at physiological temperatures, while at lower temperatures coexistence of fluid and immobilized components were observed. The ESR spectra of lipoproteins labeled with Cholestane showed even higher sensitivity to the mobility constraints. Here, the LDL spectra revealed a drastic immobilization of ChSl axial rotation already at physiological temperatures. The results of these experiments were discussed in terms of core phase transition and/or lipid-protein interactions.
Increasing evidence points to a major role for free radicals in the pathogenesis of alcohol-induced liver injury. In vitro, free radicals may be generated during ethanol metabolism by the further metabolism of acetaldehyde by molybdenum-dependent oxidases such as xanthine oxidase. Ferritin iron mobilized by such free radicals may serve as catalytic iron. Increased stores of ferritin iron and induction of microsomal P-450 reductase activity are mechanisms by which chronic alcohol feeding may potentiate the acute effects of alcohol.
The effects of lysophosphatidylcholine (lysoPC) on human erythrocyte (RBC) ghost morphology, transmembrane protein and lipid lateral mobilities, and membrane lipid composition were studied in order to elucidate mechanisms by which lysoPC immobilizes ghost membrane components [Golan, D. E., Brown, C. S., Cianci, C. M. L., Furlong, S. T., & Caulfield, J. P. (1986) J. Cell Biol. 103, 819-828]. Under standardized conditions 1.0-1.5 micrograms/mL egg lysoPC lysed 50% of RBCs and induced, in some ghosts, the formation of large patches of wrinkled membrane. Patches exhibited complete immobilization of glycophorin and band 3 and partial immobilization of the phospholipid analogue fluorescein phosphatidylethanolamine (Fl-PE), whereas adjacent smooth membrane areas manifested only partial immobilization of proteins and no immobilization of Fl-PE. Supralytic concentrations of lysoPC induced both progressive, homogeneous wrinkling of RBC ghost membranes and concentration-dependent decreases in the lateral mobilities of glycophorin, band 3, and Fl-PE. Complete immobilization of glycophorin and band 3 occurred at 8.4 micrograms/mL lysoPC and of Fl-PE at 16.8 micrograms/mL lysoPC. Monopalmitoylphosphatidylcholine (MPPC), the major component of egg lysoPC, induced both membrane wrinkling and a concentration-dependent decrease in Fl-PE mobility, with complete immobilization at 10 micrograms/mL. Other synthetic lysoPCs did not completely immobilize Fl-PE, although some caused membrane wrinkling. MPPC was incorporated into ghost membranes with a linear dependence (r = 0.97) on MPPC concentration. Relative to total membrane lipid, the lysoPC mole fraction increased from 0.2 +/- 0.1% at 0 micrograms/mL MPPC to 25 +/- 2% at 16 micrograms/mL MPPC.(ABSTRACT TRUNCATED AT 250 WORDS)
Stable bilayer lipid membrane with mobile Platear-Gibbs border have been formed. The predominant condition of the formation is the presence of lipid coverage on the teflon surface near the hole. The formation process includes transformation of the initial planar lipid bilayer into cupola-shaped one by bowing of the lipid bilayer due to hydrostatic pressure, movement of the PGb along the teflon surface. The bilayer area estimated by electric capacitance increases from 0.1 x 10(-8) F to 21 x 10(-8) F. Electric conductance of the lipid bilayer has not changed except for the phase transition and membrane collapse. The electric capacitance of the BLM formed from hydrogenated egg lecithin was changed by cooling between 60 degrees and 40 degrees C with the maximum at about phase transition range. The individual membrane sustains several scannings of the temperature without disruption which is an evidence of the stability of the cupola-shaped membranes.
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The determination of FA in cold water marine samples is challenging because of the presence of large proportions of a variety of labile PUFA. This study was undertaken to establish optimal methods for FA analysis in various sample types present in the marine environment. Several techniques used in FA analysis, including lipid fractionation, FAME formation, and picolinyl ester synthesis, were examined. Neutral lipids, acetone-mobile polar lipids, and phospholipids (PL) were readily separated from each other on columns of activated silica gel, but recoveries of PL were reduced. Deactivation of the silica gel with 20% w/w water produced variable recoveries of PL (66 +/- 22%). FAME formation with BF3 gave optimal recoveries, and a method to remove hydrocarbon contamination from these samples before GC analysis using column chromatography was optimized. Picolinyl derivatives of FA are useful in structural determinations with MS, and a new base-catalyzed transesterification method of their synthesis from FAME was developed. Finally, a series of calculations, combining FA proportions with acyl lipid class concentrations, was designed to estimate FA concentrations. In algae and animal samples, these estimates were in good agreement with actual FA concentrations determined by internal standards.
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The major feature of sickle cell anemia is the tendency of erythrocytes to sickle when exposed to decreased oxygen tension and to unsickle when reoxygenated. Irreversible sickle cells (ISCs) are sickle erythrocytes which retain bipolar elongated shapes despite reoxygenation. ISCs are believed to owe their biophysical abnormalities to acquired membrane alterations which decrease membrane deformability. While increased membrane surface viscosity has been measured in ISCs, the lateral dynamics of membrane lipids in these cells have not heretofore been examined. We have measured the lateral diffusion of the lipid analog 3,3'-dioctadecylindocyanine iodide (DiI) in the plasma membrane of intact normal erythrocytes, reversible sickle cells (RSCs), and irreversible sickle cells by fluorescence photobleaching recovery (FPR). The diffusion coefficients +/- standard errors of the mean of DiI in intact normal red blood cells (RBCs), RSCs, and ISCs at 37 degrees C are (8.06 +/- 0.29) X 10(-9) cm2 X s-1, (7.74 +/- 0.22) X 10(-9) cm2 X s-1, and (7.29 +/- 0.24) X 10(-9) cm2 X s-1, respectively. A similar decrease in the diffusion coefficient of DiI in the plasma membranes of the three cell types was observed at 4, 10, 17, 23, and 30 degrees C. ANOVA analysis of the changes in DiI diffusion showed significant differences between the RBC and ISC membranes at all temperatures examined. The characteristic breaks in Arrhenius plots of the diffusion coefficients for the RBCs, RSCs, and ISCs occurred at 20, 19, and 18.6 degrees C, respectively. Photobleaching recovery data were used to estimate (Boullier, J.A., Melnykovich, G. and Barisas, B.G. (1982) Biochim. Biophys. Acta 692, 278-286) the microviscosities of the plasma membranes of the three cell types at 25 degrees C. We find significant differences between our microviscosity values and those obtained in previous fluorescence depolarization studies. However, both methods indicate qualitatively similar differences in membrane microviscosity among the various cell types.