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Biomedical subjects

A D Postle

Publications and source records attributed to A D Postle.

At least 19 recordsLinked to original sources
PubMed2004

Mass spectroscopic analysis of phosphatidylinositol synthesis using 6-deuteriated-myo-inositol: comparison of the molecular specificities and acyl remodelling mechanisms in mouse tissues and cultured cells.

Mammalian cell PtdIns (phosphatidylinositol) in vivo is enriched in the sn-1-stearoyl 2-arachidonoyl species, the physiological precursor of phosphatidylinositol 4,5-bisphosphate. Mechanisms regulating this specificity are unclear but are typically lost for cells in culture. We used ESI-MS (tandem electrospray ionization-mass spectrometry) to determine the molecular species of PtdIns synthesized by mouse tissues in vivo compared with cultured cells in vitro. After incorporation of deuteriated myo-d(6)-inositol over 3 h, endogenous and newly synthesized PtdIns and lysoPtdIns species were quantified from precursor scans of m/z 241(-) and m/z 247(-) respectively. PtdIns was synthesized as a wide range of species irrespective of the final membrane composition. Analyses of isotope enrichments argued against acyl remodelling as the major regulatory mechanism: composition of the lysoPtdIns pool under all conditions reflected that of either endogenous or newly synthesized PtdIns and was always at equilibrium. The kinetics of PtdIns synthesis, together with the prolonged time scale required for achieving final equilibrium compositions suggest that selective transport between membranes and/or hydrolysis of selected molecular species are the most probable mechanisms regulating compositions of PtdIns and, ultimately, phosphatidylinositol 4,5-bisphosphate.

Acylation↗
PubMed2004

Lipidomic analysis of the molecular specificity of a cholinephosphotransferase in situ.

Dynamic lipidomics using ESI-MS (tandem electrospray ionization mass spectrometry) of 9-deuterated choline (choline-d(9)) incorporation into mammalian cell PtdCho (phosphatidylcholine) permits assessment of the molecular specificity of synthesis. Bulk cell PtdCho synthesis occurs in spatially distinct locations, using separate CPTs (1,2 diacylglycerol CDP:choline cholinephosphotransferases). We assessed whether in vitro molecular selectivity of DAG (diacylglycerol) incorporation between CPTs is manifest in situ, by monitoring choline-d(9) incorporation into PtdCho and lyso-PtdCho molecular species over 3 h in control cells and in CHO-K1 cells overexpressing hCEPT1. Compared with controls, the basal molecular species composition of hCEPT1 overexpressors was significantly enriched in arachidonate. This was not due to net accretion of cellular PtdCho arguing against effects of inadequate unsaturated PtdCho degradation or remodelling. Rather, time-course analyses of PtdCho and lyso-PtdCho pools showed that both arachidonate-containing DAG incorporation and turnover of PtdCho is increased in hCEPT1 overexpressors. Increased choline-d(9) incorporation into arachidonyl lyso-PtdCho shows that both phospholipase A(1)- and A(2)-mediated turnover is involved. Spatially distinct molecular specificity of DAG incorporation into cellular PtdCho at the level of hCEPT1 exists in situ.

Animals↗
PubMed2004

Use of mass spectrometry-based lipidomics to probe PITPalpha (phosphatidylinositol transfer protein alpha) function inside the nuclei of PITPalpha+/+ and PITPalpha-/- cells.

The mammalian phospholipid exchange protein PITPalpha (phosphatidylinositol transfer protein alpha), found in both extranuclear and endonuclear compartments, is thought in part to facilitate nuclear import of the PtdIns (phosphatidylinositol) consumed in the generation of proliferation-associated endonuclear diacylglycerol accumulations. Unlike phosphatidylcholine, endonuclear PtdIns is not synthesized in situ. However, despite progressive postnatal lethality of PITPalpha ablation in mice, PITPalpha(-/-) MEF (mouse embryonic fibroblasts) lack an obviously impaired proliferative capacity. We used ESI-MS (tandem electrospray ionization-MS) to monitor incorporation of the deuterated phospholipid precursors, choline-d(9) and inositol-d(6), into molecular species of whole cell and endonuclear phosphatidylcholine and PtdIns over 24 h to assess the contribution of PITPalpha to the nuclear import of PtdIns into MEF cells. In cells labelled for 1, 3, 6, 12 and 24 h fractional inositol-d(6) incorporation into whole-cell PtdIns species was consistently higher in PITPalpha(-/-) MEF implying greater flux through its biosynthetic pathway. Moreover, endonuclear accumulation of PtdIns-d(6) was apparent in the PITPalpha(-/-) cells and mirrored that in PITPalpha(+/+) cells. Together, these results suggest that the essential endonuclear PtdIns import via PITPalpha can be accommodated by other mechanisms.

Animals↗
PubMed2003

The effect of eicosapentaenoic acid on rat lymphocyte proliferation depends upon its position in dietary triacylglycerols.

Animal and human studies have shown that greatly increasing the amount of fish oil [rich in long-chain (n-3) PUFA] in the diet can decrease lymphocyte functions. The effects of a more modest provision of long-chain (n-3) PUFA and whether eicosapentaenoic acid (20:5) and docosahexaenoic acid (22:6) have the same effects as one another are unclear. Whether the position of 20:5 or 22:6 in dietary triacylglycerols (TAG) influences their incorporation into immune cells and their subsequent functional effects is not known. In this study, male weanling rats were fed for 6 wk one of 9 diets that contained 178 g lipid/kg and that differed in the type of (n-3) PUFA and in the position of these in dietary TAG. The control diet contained 4.4 g alpha-linolenic acid (18:3)/100 g total fatty acids. In the other diets, 20:5 or 22:6 replaced a portion (50 or 100%) of 18:3, and were in the sn-2 or the sn-1(3) position of dietary TAG. There were significant dose-dependent increases in the proportion of 20:5 or 22:6 in spleen mononuclear cell phospholipids when 20:5 or 22:6 was fed. These increases were at the expense of arachidonic acid and were largely independent of the position of 20:5 or 22:6 in dietary TAG. Spleen lymphocyte proliferation increased dose dependently when 20:5 was fed in the sn-1(3) position of dietary TAG. There were no significant differences in interleukin-2, interferon-gamma or interleukin-10 production among spleen cells from rats fed the different diets. Prostaglandin E(2) production by spleen mononuclear cells was decreased by inclusion of either 20:5 or 22:6 in the diet in the sn-1(3) position. Thus, incorporation of 20:5 or 22:6 into spleen mononuclear cell phospholipids is not influenced by the position in dietary TAG. However, the pattern of incorporation may be influenced, and there are some differential functional effects of the position of long-chain (n-3) PUFA in dietary TAG. A moderate increase in the intake of 20:5 at the sn-1(3) position of dietary TAG increases lymphocyte proliferation.

Animals↗
PubMed2002

Electrospray ionisation mass spectrometric analysis of lipid restructuring in the carp (Cyprinus carpio L.) during cold acclimation.

Cold acclimation of carp from 30 degrees C to 10 degrees C causes a restructuring of liver microsomal phospholipids characterised by increased proportions of monounsaturated fatty acid in phosphatidylcholine (PC) and phosphatidylethanolamine (PE). Here, we have used electrospray ionisation mass spectrometry (ESI-MS) to determine the patterns of alteration to individual molecular species compositions of PC, PE and phosphatidylinositol (PI) in response to gradually decreasing temperature. The results demonstrate that cold induces precise changes to a limited number of phospholipid species, and that these changes are distinct and different for each phospholipid class. The major change for PC was increased 16:1/22:6, but for PE the species that increased was 18:1/22:6. By contrast, the PI species that increased during cold acclimation were characterised by an sn-1 monounsaturated fatty acid in combination with arachidonoyl or eicosapentaenoyl fatty acid at the sn-2 position. Analysis of acyl distribution indicates that cold only caused the accumulation of monounsaturated fatty acids at the sn-1 and not at the sn-2 position of phospholipids. These results highlight the tight and restricted range of modifications that membranes make to their phospholipid composition in response to thermal stress.

Animals↗
PubMed2001

A comparison of the molecular species compositions of mammalian lung surfactant phospholipids.

While dipalmitoyl phosphatidylcholine (PC16:0/16:0) is essential for pulmonary surfactant function, roles for other individual molecular species of surfactant phospholipids have not been established. If any phospholipid species other than PC16:0/16:0 is important for surfactant function, then it may be conserved across animal species. Consequently, we have quantified, by electrospray ionisation mass spectrometry, molecular species compositions of phosphatidylcholine (PC), phosphatidylglycerol (PG) and phosphatidylinositol (PI) in surfactants from human, rabbit, rat and guinea pig lungs. While PC compositions displayed only relatively minor variations across the animal species studied, there were wide variations of PG and PI concentrations and compositions. Human surfactant PG and PI were enriched in the same three monounsaturated species (PG16:0/18:1, PG18:1/18:1 and PG18:0/18:1) with minimal amounts of PG16:0/16:0 or polyunsaturated species, while all animal surfactant PG contained increased concentrations of PG16:0/16:0 and PG16:0/18:2. Animal surfactant PIs were essentially monounsaturated except for a high content of PI18:0/20:4 (29%) in the rat. As these four surfactants all maintain appropriate lung function of the respective animal species, then all their varied compositions of acidic phospholipids must be adequate at promoting the processes of adsorption, film refinement, respreading and collapse characteristic of surfactant. We conclude that this effectively monounsaturated composition of anionic phospholipid molecular species is a common characteristic of mammalian surfactants.

Animals↗
PubMed2001

Pulmonary and gastric surfactants. A comparison of the effect of surface requirements on function and phospholipid composition.

Surfactant 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.

Animals↗
PubMed2001

Pulmonary surfactant in birds: coping with surface tension in a tubular lung.

As birds have tubular lungs that do not contain alveoli, avian surfactant predominantly functions to maintain airflow in tubes rather than to prevent alveolar collapse. Consequently, we have evaluated structural, biochemical, and functional parameters of avian surfactant as a model for airway surfactant in the mammalian lung. Surfactant was isolated from duck, chicken, and pig lung lavage fluid by differential centrifugation. Electron microscopy revealed a uniform surfactant layer within the air capillaries of the bird lungs, and there was no tubular myelin in purified avian surfactants. Phosphatidylcholine molecular species of the various surfactants were measured by HPLC. Compared with pig surfactant, both bird surfactants were enriched in dipalmitoylphosphatidylcholine, the principle surface tension-lowering agent in surfactant, and depleted in palmitoylmyristoylphosphatidylcholine, the other disaturated phosphatidylcholine of mammalian surfactant. Surfactant protein (SP)-A was determined by immunoblot analysis, and SP-B and SP-C were determined by gel-filtration HPLC. Neither SP-A nor SP-C was detectable in either bird surfactant, but both preparations of surfactant contained SP-B. Surface tension function was determined using both the pulsating bubble surfactometer (PBS) and capillary surfactometer (CS). Under dynamic cycling conditions, where pig surfactant readily reached minimal surface tension values below 5 mN/m, neither avian surfactant reached values below 15 mN/m within 10 pulsations. However, maximal surface tension of avian surfactant was lower than that of porcine surfactant, and all surfactants were equally efficient in the CS. We conclude that a surfactant composed primarily of dipalmitoylphosphatidylcholine and SP-B is adequate to maintain patency of the air capillaries of the bird lung.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed2000

Highly saturated endonuclear phosphatidylcholine is synthesized in situ and colocated with CDP-choline pathway enzymes.

Chromatin-associated phospholipids are well recognized. A report that catalytically active endonuclear CTP:choline-phosphate cytidylyltransferase alpha is necessary for cell survival questions whether endonuclear, CDP-choline pathway phosphatidylcholine synthesis may occur in situ. We report that chromatin from human IMR-32 neuroblastoma cells possesses such a biosynthetic pathway. First, membrane-free nuclei retain all three CDP-choline pathway enzymes in proportions comparable with the content of chromatin-associated phosphatidylcholine. Second, following supplementation of cells with deuterated choline and using electrospray ionization mass spectrometry, both the time course and molecular species labeling pattern of newly synthesized endonuclear and whole cell phosphatidylcholine revealed the operation of spatially separate, compositionally distinct biosynthetic routes. Specifically, endogenous and newly synthesized endonuclear phosphatidylcholine species are both characterized by a high degree of diacyl/alkylacyl chain saturation. This unusual species content and synthetic pattern (evident within 10 min of supplementation) are maintained through cell growth arrest by serum depletion and when proliferation is restored, suggesting that endonuclear disaturated phosphatidylcholine enrichment is essential and closely regulated. We propose that endonuclear phosphatidylcholine synthesis may regulate periodic nuclear accumulations of phosphatidylcholine-derived lipid second messengers. Furthermore, our estimates of saturated phosphatidylcholine nuclear volume occupancy of around 10% may imply a significant additional role in regulating chromatin structure.

Choline-Phosphate Cytidylyltransferase↗
PubMed2000

Phospholipid molecular species of bronchoalveolar lavage fluid after local allergen challenge in asthma.

Electrospray ionization mass spectrometry was used to quantify phosphatidylcholine (PC) and phosphatidylglycerol (PG) molecular species in bronchoalveolar lavage fluid (BALF) from control and mild asthmatic subjects after local allergen challenge. BALF was obtained from 5 control and 13 asthmatic subjects before and 24 h after segmental allergen and saline challenge. There were no differences in the ratio of total PC to total PG or in the molecular species composition of PC or PG between the asthmatic and control groups under basal conditions. Allergen challenge in asthmatic but not in control volunteers caused a significant increase in the PC-to-PG ratio because of increased concentrations of PC species containing linoleic acid (16:0/18:2 PC, 18:0/18:2 PC, and 18:1/18:2 PC). These molecular species were characteristic of plasma PC analyzed from the same subjects, strongly suggesting that the altered PC composition in BALF in asthmatic subjects after allergen challenge was due to infiltration of plasma lipoprotein, not to catabolism of surfactant phospholipid. Interactions between surfactant and lipoprotein infiltrate may contribute to surfactant dysfunction and potentiate disease severity in asthma.

Allergens↗
PubMed2000

Altered airway surfactant phospholipid composition and reduced lung function in asthma.

Pulmonary surfactant in bronchoalveolar lavage fluid (BALF) and induced sputum from adults with stable asthma (n = 36) and healthy controls (n = 12) was analyzed for phospholipid and protein compositions and function. Asthmatic subjects were graded as mild, moderate, or severe. Phospholipid compositions of BALF and sputum from control subjects were similar and characteristic of surfactant. For asthmatic subjects, the proportion of dipalmitoyl phosphatidylcholine (16:0/16:0PC), the major phospholipid in surfactant, decreased in sputum (P < 0.05) but not in BALF. In BALF, mole percent 16:0/16:0PC correlated with surfactant function measured in a capillary surfactometer, and sputum mole percent 16:0/16:0PC correlated with lung function (forced expiratory volume in 1 s). Neither surfactant protein A nor total protein concentration in either BALF or sputum was altered in asthma. These results suggest altered phospholipid composition and function of airway (sputum) but not alveolar (BALF) surfactant in stable asthma. Such underlying surfactant dysfunction may predispose asthmatic subjects to further surfactant inhibition by proteins or aeroallergens in acute asthma episodes and contribute to airway closure in asthma. Consequently, administration of an appropriate therapeutic surfactant could provide clinical benefit in asthma.

Adolescent↗
PubMed2000

Exogenous surfactant supplementation in infants with respiratory syncytial virus bronchiolitis.

Infants with respiratory syncytial virus (RSV) bronchiolitis are deficient in surfactant, both in quantity and ability to reduce surface tension. New evidence suggests surfactant has a role in maintaining the patency of conducting airways, which has implications for RSV bronchiolitis. A randomized, controlled pilot study was undertaken to assess the effects of exogenous surfactant supplementation to RSV-positive infants on pulmonary mechanics, indices of gas exchange, and the phospholipid composition of bronchoalveolar lavage fluid (BALF). Nineteen ventilated infants (median corrected age 4 wk) received either two doses of surfactant (Survanta, 100 mg/kg) within 24 and 48 h of mechanical ventilation (n = 9), or air placebo (n = 10). Static lung compliance and resistance of infants in the placebo but not in the surfactant-treated group became progressively worse over the first 30 h following enrollment. Although no significant acute changes in gas exchange parameters were seen following surfactant, infants in the surfactant group showed a more rapid improvement in oxygenation and ventilation indices over the first 60 h of ventilation. Surfactant status was assessed from the concentration ratio in BALF of the disaturated phospholipid species dipalmitoylphosphatidylcholine to that of the monounsaturated species palmitoyloleoylphosphatidylcholine. This ratio correlated with both lung compliance (positively) and resistance (negatively), and over time increased in the treated group and declined in placebo infants. The data from this pilot study suggest that functional surfactant has a role in maintaining small airway patency as well as lung compliance in infants infected with RSV and an outcome study is now warranted.

Bronchiolitis, Viral↗
PubMed1999

Deficient hydrophilic lung surfactant proteins A and D with normal surfactant phospholipid molecular species in cystic fibrosis.

Chronic bacterial colonization of the lungs, with an excessive inflammatory response, is the major cause of morbidity and mortality in cystic fibrosis. Lung surfactant exhibits a spectrum of potential immunomodulatory properties: phospholipid components inhibit cellular inflammatory responses, whereas the hydrophilic surfactant proteins A (SP-A) and D (SP-D) are integral components of the innate host defense response of the lungs against bacterial infection. Consequently, alteration to the relative proportions of lung surfactant components may alter the susceptibility of the lungs to bacterial colonization. In this study, bronchoalveolar lavage (BAL) samples were collected at diagnostic fiberoptic bronchoscopy from 11 control children, 13 children with cystic fibrosis, and 11 children with acute lung infection. Electrospray ionization mass spectrometry analysis demonstrated negligible changes to the molecular species or total BAL concentrations of phosphatidylcholine, phosphatidylglycerol, or phosphatidylinositol among the three subject groups. In contrast, median SP-A concentration was decreased (P < 0.001) in the cystic fibrosis group (2.65 microg/ml) compared with control (12.35 microg/ml) and infection (9.76 microg/ml) groups. Median SP-D was also decreased (P < 0.05) in the infection (12.17 ng/ml) compared with the control group (641 ng/ml), and was below assay limits for the majority of cystic fibrosis children (P < 0. 001). This dramatic decrease of hydrophilic surfactant proteins in the presence of normal surfactant phospholipid may be one mechanism underlying the relative ineffectiveness of the cellular inflammatory response in killing invading bacteria in the lungs of patients with cystic fibrosis.

Bacterial Infections↗
PubMed1998

Pregnancy-associated adaptations to hepatic phosphatidylcholine biosynthesis in the guinea-pig.

Pregnancy is associated with increased phosphatidylcholine (PC) 16:0/22:6 and PC16:0/20:4 concentrations in rat liver and plasma, guinea-pig liver, and in plasma in women. These changes may be related to supply of polyunsaturated fatty acids (PUFA) to the fetus. For the rat, these adaptations to hepatic PC composition are regulated by modifications to synthesis de novo from choline. However, it is not known whether these adaptations are applicable to other species. Consequently, we have determined biochemical mechanisms for regulation of hepatic PC synthesis in the pregnant guinea pig. The PUFA content of guinea-pig liver PC synthesised de novo did not change significantly during pregnancy. [Methyl-14C]methionine incorporation into PC in vivo, however, showed increased PC16:0/22:6 and PC16:0/20:4 contents. [Methyl-14C]methionine incorporation into PC over 6 hr in vivo increased during early pregnancy, while PC synthesis de novo did not change. In contrast to the rat, modulation of PE N-methylation is a primary mechanism for regulating the PUFA content of hepatic PC in the pregnant guinea-pig. The use of distinct metabolic strategies to achieve comparable pregnancy-associated adaptations to hepatic PC composition between these animal species suggests both evolutionary convergence and a fundamental the role for PC16:0/22:6 and PC16:0/20:4 in PUFA metabolism during gestation.

Adaptation, Physiological↗