1,721,045 research outputs found
Phosphatidylcholine biosynthesis inside the nucleus: is it involved in regulating cell proliferation?
No full textDynamic lipidomics with stable isotope labeling
No full textIncorporation of stable isotope labelled precursors enables estimation of the kinetics of lipid synthesis and turnover (dynamic lipidomics) in the clinical as well the experimental setting. Recent advances in tandem mass spectrometry extend the analytical possibilities from measurements of isotope enrichments to determinations of intact substrates. Incorporations of deuteriated choline, ethanolamine and inositol can be determined by precursor and neutral loss scans of phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol, respectively. This experimental approach provides information on the kinetics of individual phospholipid molecular species and has considerable potential to probe diseases of lipid metabolism in vivo
Mass spectrometry determination of endonuclear phospholipid composition and dynamics
No full textMammalian cell lipid analyses using tandem electrospray ionization mass spectrometry, in conjunction with stable isotope labeling, permit unparalleled access to membrane phospholipid molecular species compositions and turnover. Lipidomic data from isolable compartments of lipid second messenger generation, such as membrane-free nuclei, can provide dynamic insights into the topology of phospholipid turnover. For example, ESI-MS/MS precursor scans of characteristic phosphocholine m/z 184+ fragments reveal a highly saturated endonuclear phosphatidylcholine pool with homeostatic maintenance properties. A spatially distinct CDPcholine pathway yields, within minutes of choline-d9 labeling, unsaturated endonuclear phosphatidylcholines progressively remodeled to more saturated species evidenced by tracking the deuteriated headgroup through precursor scans of phosphocholine-d9 (m/z 193+ fragment). Among the other endonuclear phospholipids, diacyl phosphatidylethanolamines (neutral loss of m/z 141+) are also highly saturated compared with those of whole cell whereas, phophatidylinositols (precursor scans of m/z 241? fragment) are essentially identical in nuclei and whole cells. Moreover, the pattern of myo-inositol-d6 acquisition into endonuclear phosphatidylinositol (precursor scans of m/z 247? fragment) is inconsistent with compartment-specific synthesis. Endonuclear sphingomyelins (seen in precursor scans of m/z 184+ and confirmed from precursor scans of m/z 168? fragments) are enriched but similar in composition to whole cell species whereas endonuclear phosphatidylserines (neutral loss of m/z 87?) are more saturated than their whole cell counterparts. The focus of described methodologies emphasize their value in probing the compositions and dynamics of endonuclear phospholipids, but in principle may be extended to exploration of other isolable compartments including ER or plasma membranes
Dynamic lipidomic insights into phosphatidylcholine synthesis from organelle to organism
No full textRecent technical improvement and technological innovation in small molecule mass spectrometry have provided powerful tools for the intensive metabolomic biochemical investigations that will necessarily characterise the "post-genomic" era of biomedical research. For membrane phospholipids, use of tandem electrospray ionisation mass spectrometry (ESI-MS/MS) exploiting precursor scanning of class-specific diagnostic fragments, can provide detailed quantitative profiles at the level of individual molecular species for many hundreds of unique lipids. Such "snapshot" measurements provide little information concerning metabolic flux. However, recent use of metabolic labelling with stable isotope derivatives of phospholipid headgroups combined with precursor scans of unlabelled and labelled fragments have yielded considerable insight into phosphatidylcholine metabolism in vivo. Here, we briefly review some of the recent work on pathways of phosphatidylcholine metabolism ranging from studies at subcellular organelle level through to whole organism. The sensitivity, specificity and suitability of this powerful methodological approach to numerous questions of phospholipid metabolism place ESI-MS/MS at the very heart of dynamic lipidomics in the foreseeable future
Analysis of the regulation of surfactant phosphatidylcholine metabolism using stable isotopes
No full textThe pathways and mechanisms that regulate pulmonary surfactant synthesis, processing, secretion and catabolism have been extensively characterised using classical biochemical and analytical approaches. These have constructed a model, largely in experimental animals, for surfactant phospholipid metabolism in the alveolar epithelial cell whereby phospholipid synthesised on the endoplasmic reticulum is selectively transported to lamellar body storage vesicles, where it is subsequently processed before secretion into the alveolus. Surfactant phospholipid is a complex mixture of individual molecular species defined by the combination of esterified fatty acid groups and a comprehensive description of surfactant phospholipid metabolism requires consideration of the interactions between such molecular species. However, until recently, lipid analytical techniques have not kept pace with the considerable advances in understanding of the enzymology and molecular biology of surfactant metabolism. Refinements in electrospray ionisation mass spectrometry (ESI-MS) can now provide very sensitive platforms for the rapid characterisation of surfactant phospholipid composition in molecular detail. The combination of ESI-MS and administration of phospholipid substrates labelled with stable isotopes extends this analytical approach to the quantification of synthesis and turnover of individual molecular species of surfactant phospholipid. As this methodology does not involve radioactivity, it is ideally suited to application in clinical studies. This review will provide an overview of the metabolic processes that regulate the molecular specificity of surfactant phosphatidylcholine together with examples of how the application of stable isotope technologies in vivo has, for the first time, begun to explore regulation of the molecular specificity of surfactant synthesis in human subjects
Regulation of lung surfactant phospholipid synthesis and metabolism
No full textThe alveolar type II epithelial (ATII) cell is highly specialised for the synthesis and storage, in intracellular lamellar bodies, of phospholipid destined for secretion as pulmonary surfactant into the alveolus. Regulation of the enzymology of surfactant phospholipid synthesis and metabolism has been extensively characterised at both molecular and functional levels, but understanding of surfactant phospholipid metabolism in vivo in either healthy or, especially, diseased lungs is still relatively poorly understood. This review will integrate recent advances in the enzymology of surfactant phospholipid metabolism with metabolic studies in vivo in both experimental animals and human subjects. It will highlight developments in the application of stable isotope-labelled precursor substrates and mass spectrometry to probe lung phospholipid metabolism in terms of individual molecular lipid species and identify areas where a more comprehensive metabolic model would have considerable potential for direct application to disease states
Selective changes to phosphatidylcholine and phosphatidylethanolamine molecular species in the developing fetal guinea pig liver and plasma
No full textThe molecular species composition of membrane phospholipids influences the activities of integral proteins and cell signalling pathways. We determined the effect of increasing gestational age on fetal guinea pig liver phosphatidylcholine (PC) and phosphatidylethanolamine (PE), and plasma PC molecular species composition. The livers were collected from fetuses (n = 5/time point) at 5 day intervals between 40 and 65 days of gestation, and at term (68 days). Hepatic PC and PE molecular species composition was determined by electrospray ionisation mass spectrometry. An increasing gestational age was accompanied by selective changes in individual molecular species. The proportion of the sn-1 18:0 species increased relative to the sn-1 16:0 species in liver PC, but not PE, with an increasing gestational age. 1-O-alkyl-2-acyl PC species concentrations decreased significantly between 40 and 45 days of gestation (40%), and 65 and 68 days (54%). Total 1-O-alkenyl-2-acyl PE species concentration increased between days 60 and 65, due to a rise in 1-O-16:0 alkyl/20:4 content, and then decreased until term. Between day 40 and term, PC and PE sn-2 18:2n-6 species concentrations increased 3-fold. PC16:0/18:2 increased gradually throughout gestation, while PC18:0/18:2 content only increased after day 65. The overall increase in PE18:2n-6 content was due to PE18:0/18:2 alone. The composition of plasma PC essentially reflected hepatic PC. Overall, these data suggest differential regulation of hepatic PC and PE molecular species composition during development which is essentially independent of the maternal fatty acid supply
Testing the hypothesis that amphiphilic antineoplastic lipid analogues act through reduction of membrane curvature elastic stress
No full textThe alkyllysophospholipid (ALP) analogues Mitelfosine and Edelfosine are anticancer drugs whose mode of action is still the subject of debate. It is agreed that the primary interaction of these compounds is with cellular membranes. Furthermore, the membrane-associated protein CTP: phosphocholine cytidylyltransferase (CCT) has been proposed as the critical target. We present the evaluation of our hypothesis that ALP analogues disrupt membrane curvature elastic stress and inhibit membrane-associated protein activity (e.g. CCT), ultimately resulting in apoptosis. This hypothesis was tested by evaluating structure-activity relationships of ALPs from the literature. In addition we characterized the lipid typology, cytotoxicity and critical micelle concentration of novel ALP analogues that we synthesized. Overall we find the literature data and our experimental data provide excellent support for the hypothesis, which predicts that the most potent ALP analogues will be type I lipids
Chapter 9 Polyunsaturated fatty acids, brain phospholipids and the fetal alcohol syndrome
No full textAlthough to date these observations in the fetal guinea pig have not been substantiated in humans, they provide strong preliminary evidence that impaired accumulation of DHA into brain phospholipids may be one important mechanism in the pathogenesis of FAS. Furthermore, since increasing DHA availability to the mother, and presumably the fetus, appeared to produce a reduction in the severity of ethanol-induced neurological damage it is possible that maternal DHA supplementation may provide a therapeutic strategy in humans FAS. Alternatively, the prolonged period of postnatal brain development in the human may permit appropriate dietary DHA supplementation of the affected infants after delivery, which would potentially be a more practical clinical intervention.<br/
Pulmonary and gastric surfactants. A comparison of the effect of surface requirements on function and phospholipid composition
No full textSurfactant is present in the alveoli and conductive airways of mammalian lungs. The presence of surface active agents was, moreover, demonstrated for avian tubular lungs and for the stomach and intestine. As the surface characteristics of these organs differ from each other, their surfactants possess distinct biochemical and functional characteristics. In the stomach so-called ‘gastric surfactant’ forms a hydrophobic barrier to protect the mucosa against acid back-diffusion. For this purpose gastric mucosal cells secrete unsaturated phosphatidylcholines (PC), but no dipalmitoyl-PC (PC16:0/16:0). By contrast, surfactant from conductive airways, lung alveoli and tubular avian lungs contain PC16:0/16:0 as their main component in similar concentrations. Hence, there is no biochemical relation between gastric and pulmonary surfactant. Alveolar surfactant, being designed for preventing alveolar collapse under the highly dynamic conditions of an oscillating alveolus, easily reaches values of <5 mN/m upon cyclic compression. Surfactants from tubular air-exposed structures, however, like the conductive airways of mammalian lungs and the exclusively tubular avian lung, display inferior compressibility as they only reach minimal surface tension values of approximately 20 mN/m. Hence, the highly dynamic properties of alveolar surfactant do not apply for surfactants designed for air–liquid interfaces of tubular lung structures
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