Science.gov

Sample records for cell membrane fraction

  1. Lipids that determine detergent resistance of MDCK cell membrane fractions.

    PubMed

    Manni, Marco M; Cano, Ainara; Alonso, Cristina; Goñi, Félix M

    2015-10-01

    A comparative lipidomic study has been performed of whole Madin-Darby canine kidney epithelial cells and of the detergent-resistant membrane fraction (DRM) obtained after treating the cells with the non-ionic detergent Triton X-100. The DRM were isolated following a standard procedure that is extensively used in cell biology studies. Significant differences were found in the lipid composition of the whole cells and of DRM. The latter were enriched in all the analyzed sphingolipid classes: sphingomyelins, ceramides and hexosylceramides. Diacylglycerols were also preferentially found in DRM. The detergent-resistant fraction was also enriched in saturated over unsaturated fatty acyl chains, and in sn-1 acyl chains containing 16 carbon atoms, over the longer and shorter ones. The glycerophospholipid species phosphatidylethanolamines and phosphatidylinositols, that were mainly unsaturated, did not show a preference for DRM. Phosphatidylcholines were an intermediate case: the saturated, but not the unsaturated species were found preferentially in DRM. The question remains on whether these DRM, recovered from detergent-membrane mixtures by floatation over a sucrose gradient, really correspond to membrane domains existing in the cell membrane prior to detergent treatment.

  2. Comparative studies of two membrane fractions isolated from chemotrophically and phototrophically grown cells of Rhodopseudomonas capsulata.

    PubMed Central

    Garcia, A F; Drews, G; Reidl, H H

    1981-01-01

    Light and heavy membrane fractions have been isolated by equilibrium sucrose density centrifugation from Rhodopseudomonas capsulata 938 GCM grown aerobically in the dark (chemotrophically) and anaerobically in the light (phototrophically). The densities of the light and heavy fractions from phototrophic cells were 1.1004 to 1.1006 and 1.1478, respectively, and the densities of the light and heavy fractions from chemotrophic cells were 1.0957 to 1.0958 and 1.1315, respectively. Both fractions were active in photochemical and respiratory functions and in electron transport-coupled phosphorylation. The light membrane fraction isolated from chemotrophic cells contained the reaction center and the light-harvesting pigment-protein complex B 870, but not the variable light-harvesting complex B 800-850. A small amount of the complex B 800-850 was present in the light fraction isolated from phototrophically grown cells, but it was not energetically coupled to the photosynthetic apparatus. From inhibitor studies, difference spectroscopy, and measurement of enzyme activities it was tentatively concluded that the light membrane fraction contains only the reduced nicotinamide adenine dinucleotide-oxidizing electron transport chain having a KCN-insensitive, low-potential cytochrome c oxidase, whereas the heavy fraction contains additionally the succinate dehydrogenase and a high-potential cytochrome b terminal oxidase sensitive to KCN. The light membrane fraction was more labile than the heavy fraction in terms of phosphorylating activity. PMID:7204341

  3. Chemical modification and immunogenicity of membrane fractions from mouse tumour cells.

    PubMed Central

    Staab, H. J.; Anderer, F. A.

    1978-01-01

    A crude membrane fraction isolated from mouse tumour cells was treated with various chemicals. The effects on the immunogenicity of the membrane sample were tested in syngeneic mice for tumour protection, using a challenge dose of 10(5) viable tumour cells. Best protection was obtained after immunization of mice with a membrane sample modified with dimethylsulphate. Up to 60% of the animals remained tumour free, and the tumour-bearing animals showed a greatly increased mean survival time. The post-challenge sera contained no detectable amounts of cytotoxic antibodies. The membrane sample isolated from tumour cells which had been modified with dimethylsulphate showed less immunogenicity than the modified cells or the membrane fraction from unmodified cells. PMID:215180

  4. Tissue-culture cell fractionation. Fractionation of membranes from tissue-culture cells homogenized by glycerol-induced lysis.

    PubMed

    Graham, J M; Sandall, J K

    1979-07-15

    1. The disruption of various types of tissue-culture cells by (a) incubation in solutions of 1.2 M-glycerol and (b) transfer of the glycerol-loaded cells to relatively hypo-osmotic solutions of 0.25 M-sucrose was studied. 2. Bivalent cations (2mM-Mg2+) were generally included to preserve the nuclei, but some cells (polyoma-virus-transformed baby-hamster kidney cells) failed to be disrupted adequately under these conditions. 3. Other cells (mouse-embryo fibroblasts) required additional gentle Dounce homogenization to effect complete cell breakage. 4. Purification of the whole homogenate was carried out by a combination of differential centrifugation and sedimentation or flotation through sucrose gradients. 5. Enzyme analysis showed that plasma-membrane, endoplasmic-reticulum and mitochondrial fractions were obtained in good yield and purity.

  5. Fraction Reduction in Membrane Systems

    PubMed Central

    Zhang, Hong

    2014-01-01

    Fraction reduction is a basic computation for rational numbers. P system is a new computing model, while the current methods for fraction reductions are not available in these systems. In this paper, we propose a method of fraction reduction and discuss how to carry it out in cell-like P systems with the membrane structure and the rules with priority designed. During the application of fraction reduction rules, synchronization is guaranteed by arranging some special objects in these rules. Our work contributes to performing the rational computation in P systems since the rational operands can be given in the form of fraction. PMID:24772037

  6. Optimisation of the Factor VIII yield in mammalian cell cultures by reducing the membrane bound fraction.

    PubMed

    Kolind, Mille Petersen; Nørby, Peder Lisby; Berchtold, Martin Werner; Johnsen, Laust Bruun

    2011-02-20

    In vivo, clotting Factor VIII (FVIII) circulates in plasma bound to von Willebrand factor (vWF), and the vWF:FVIII complex prevents binding of FVIII to phosphatidylserine (PS). Activation of FVIII by thrombin releases FVIII from vWF, and subsequently FVIII binds to PS exposed on activated platelets and forms the tenase complex together with clotting Factor IX. In vitro, during serum free production of recombinant FVIII (rFVIII), production cells also expose PS, and since vWF is not present to hinder interaction of secreted rFVIII with PS, rFVIII is partly associated with the cell membrane of the production cells. Recently, we showed that as much as 90% of secreted rFVIII is bound to transiently transfected production cells during serum free conditions. In this study, we investigated the effect of including vWF in the serum free medium, and demonstrate that addition of vWF results in release of active membrane bound rFVIII to the culture medium. Moreover, the attachment of rFVIII to cell membranes of un-transfected HEK293 cells was studied in the presence of compounds that competes for interactions between rFVIII and PS. Competitive assays between iodinated rFVIII (¹²⁵I-rFVIII) and annexin V or ortho-phospho-L-serine (OPLS) demonstrated that annexin V and OPLS were able to reduce the membrane bound fraction of rFVIII by 70% and 30%, respectively. Finally, adding OPLS to CHO cells stably expressing FVIII increased the yield by 50%. Using this new knowledge, the recovery of rFVIII could be increased considerably during serum free production of this therapeutic protein.

  7. Fractional order models of viscoelasticity as an alternative in the analysis of red blood cell (RBC) membrane mechanics.

    PubMed

    Craiem, Damian; Magin, Richard L

    2010-01-20

    New lumped-element models of red blood cell mechanics can be constructed using fractional order generalizations of springs and dashpots. Such 'spring-pots' exhibit a fractional order viscoelastic behavior that captures a wide spectrum of experimental results through power-law expressions in both the time and frequency domains. The system dynamics is fully described by linear fractional order differential equations derived from first order stress-strain relationships using the tools of fractional calculus. Changes in the composition or structure of the membrane are conveniently expressed in the fractional order of the model system. This approach provides a concise way to describe and quantify the biomechanical behavior of membranes, cells and tissues.

  8. Fractional order models of viscoelasticity as an alternative in the analysis of red blood cell (RBC) membrane mechanics

    PubMed Central

    Craiem, Damian; Magin, Richard L

    2011-01-01

    New lumped-element models of red blood cell mechanics can be constructed using fractional order generalizations of springs and dashpots. Such ‘spring-pots’ exhibit a fractional order viscoelastic behavior that captures a wide spectrum of experimental results through power-law expressions in both the time and frequency domains. The system dynamics is fully described by linear fractional order differential equations derived from first order stress–strain relationships using the tools of fractional calculus. Changes in the composition or structure of the membrane are conveniently expressed in the fractional order of the model system. This approach provides a concise way to describe and quantify the biomechanical behavior of membranes, cells and tissues. PMID:20090192

  9. Isolation of Endoplasmic Reticulum, Mitochondria, and Mitochondria-Associated Membrane and Detergent Resistant Membrane Fractions from Transfected Cells and from Human Cytomegalovirus-Infected Primary Fibroblasts.

    PubMed

    Williamson, Chad D; Wong, Daniel S; Bozidis, Petros; Zhang, Aiping; Colberg-Poley, Anamaris M

    2015-09-01

    Increasingly mechanistic virology studies require dependable and sensitive methods for isolating purified organelles containing functional cellular sub-domains. The mitochondrial network is, in part, closely apposed to the endoplasmic reticulum (ER). The mitochondria-associated membrane (MAM) fraction provides direct physical contact between the ER and mitochondria. Characterization of the dual localization and trafficking of human cytomegalovirus (HCMV) UL37 proteins required establishing protocols in which the ER and mitochondria could be reliably separated. Because of its documented role in lipid and ceramide transfer from the ER to mitochondria, a method to purify MAM from infected cells was also developed. Two robust procedures were developed to efficiently isolate mitochondria, ER, and MAM fractions while providing substantial protein yields from HCMV-infected primary fibroblasts and from transfected HeLa cells. Furthermore, this unit includes protocols for isolation of detergent resistant membranes from subcellular fractions as well as techniques that allow visualization of the mitochondrial network disruption that occurs in permissively infected cells by their optimal resolution in Percoll gradients.

  10. Isolation of Endoplasmic Reticulum, Mitochondria, and Mitochondria-Associated Membrane and Detergent Resistant Membrane Fractions from Transfected Cells and from Human Cytomegalovirus-Infected Primary Fibroblasts

    PubMed Central

    Williamson, Chad D.; Wong, Daniel S.; Bozidis, Petros; Zhang, Aiping; Colberg-Poley, Anamaris M.

    2015-01-01

    Increasingly mechanistic virology studies require dependable and sensitive methods for isolating purified organelles containing functional cellular sub-domains. The mitochondrial network is, in part, closely apposed to the endoplasmic reticulum (ER). The mitochondria-associated membrane (MAM) fraction provides direct physical contact between the ER and mitochondria. Characterization of the dual localization and trafficking of human cytomegalovirus (HCMV) UL37 proteins required establishing protocols in which the ER and mitochondria could be reliably separated. Because of its documented role in lipid and ceramide transfer from the ER to mitochondria, a method to purify MAM from infected cells was also developed. Two robust procedures were developed to efficiently isolate mitochondria, ER, and MAM fractions while providing substantial protein yields from HCMV-infected primary fibroblasts and from transfected HeLa cells. Furthermore, this unit includes protocols for isolation of detergent resistant membranes from subcellular fractions as well as techniques that allow visualization of the mitochondria network disruption that occurs in permissively infected cells by their optimal resolution in Percoll gradients. PMID:26331984

  11. Enrichment of distinct microfilament-associated and GTP-binding-proteins in membrane/microvilli fractions from lymphoid cells

    PubMed Central

    Hao, Jian-Jiang; Wang, Guanghui; Pisitkun, Trairak; Patino-Lopez, Genaro; Nagashima, Kunio; Knepper, Mark A.; Shen, Rong-Fong; Shaw, Stephen

    2008-01-01

    Summary Lymphocyte microvilli mediate initial adhesion to endothelium during lymphocyte transition from blood into tissue but their molecular organization is incompletely understood. We modified a shear-based procedure to prepare biochemical fractions enriched for membrane/microvilli (MMV) from both human peripheral blood T-lymphocytes (PBT) and a mouse pre-B lymphocyte line (300.19). Enrichment of proteins in MMV relative to post nuclear lysate was determined by LC/MS/MS analysis and label-free quantitation. Subsequent analysis emphasized the 291 proteins shared by PBT and 300.19 and estimated by MS peak area to be highest abundance. Validity of the label-free quantitation was confirmed by many internal consistencies and by comparison with Western blot analyses. The MMV fraction was enriched primarily for subsets of cytoskeletal proteins, transmembrane proteins and G-proteins, with similar patterns in both lymphoid cell types. The most enriched cytoskeletal proteins were microfilament-related proteins NHERF1, Ezrin/Radixin/Moesin (ERMs), ADF/cofilin and Myosin1G. Other microfilament proteins such as talin, gelsolin, myosin II and profilin were markedly reduced in MMV, as were intermediate filament- and microtubule-related proteins. Heterotrimeric G-proteins and some small G-proteins (especially Ras and Rap1) were enriched in the MMV preparation. Two notable general observations also emerged. There was less overlap between the two cells in their transmembrane proteins than in other classes of proteins, consistent with a special role of plasma membrane proteins in differentiation. Second, unstimulated primary T-lymphocytes have an unusually high concentration of actin and other microfilament related proteins, consistent with the singular role of actin-mediated motility in the immunological surveillance performed by these primary cells. Lymphocyte microvilli initiate adhesion to endothelium during movement from blood into tissue. Using LC/MS/MS and label

  12. Isolation and Analysis of Detergent-Resistant Membrane Fractions.

    PubMed

    Aureli, Massimo; Grassi, Sara; Sonnino, Sandro; Prinetti, Alessandro

    2016-01-01

    The hypothesis that the Golgi apparatus is capable of sorting proteins and sending them to the plasma membrane through "lipid rafts," membrane lipid domains highly enriched in glycosphingolipids, sphingomyelin, ceramide, and cholesterol, was formulated by van Meer and Simons in 1988 and came to a turning point when it was suggested that lipid rafts could be isolated thanks to their resistance to solubilization by some detergents, namely Triton X-100. An incredible number of papers have described the composition and properties of detergent-resistant membrane fractions. However, the use of this method has also raised the fiercest criticisms. In this chapter, we would like to discuss the most relevant methodological aspects related to the preparation of detergent-resistant membrane fractions, and to discuss the importance of discriminating between what is present on a cell membrane and what we can prepare from cell membranes in a laboratory tube.

  13. Quantitative proteomics of fractionated membrane and lumen exosome proteins from isogenic metastatic and nonmetastatic bladder cancer cells reveal differential expression of EMT factors.

    PubMed

    Jeppesen, Dennis Kjølhede; Nawrocki, Arkadiusz; Jensen, Steffen Grann; Thorsen, Kasper; Whitehead, Bradley; Howard, Kenneth A; Dyrskjøt, Lars; Ørntoft, Torben Falck; Larsen, Martin R; Ostenfeld, Marie Stampe

    2014-03-01

    Cancer cells secrete soluble factors and various extracellular vesicles, including exosomes, into their tissue microenvironment. The secretion of exosomes is speculated to facilitate local invasion and metastatic spread. Here, we used an in vivo metastasis model of human bladder carcinoma cell line T24 without metastatic capacity and its two isogenic derivate cell lines SLT4 and FL3, which form metastases in the lungs and liver of mice, respectively. Cultivation in CLAD1000 bioreactors rather than conventional culture flasks resulted in a 13- to 16-fold increased exosome yield and facilitated quantitative proteomics of fractionated exosomes. Exosomes from T24, SLT4, and FL3 cells were partitioned into membrane and luminal fractions and changes in protein abundance related to the gain of metastatic capacity were identified by quantitative iTRAQ proteomics. We identified several proteins linked to epithelial-mesenchymal transition, including increased abundance of vimentin and hepatoma-derived growth factor in the membrane, and casein kinase II α and annexin A2 in the lumen of exosomes, respectively, from metastatic cells. The change in exosome protein abundance correlated little, although significant for FL3 versus T24, with changes in cellular mRNA expression. Our proteomic approach may help identification of proteins in the membrane and lumen of exosomes potentially involved in the metastatic process.

  14. Identification of novel autophagic Radix Polygalae fraction by cell membrane chromatography and UHPLC-(Q)TOF-MS for degradation of neurodegenerative disease proteins

    PubMed Central

    Wu, An-Guo; Kam-Wai Wong, Vincent; Zeng, Wu; Liu, Liang; Yuen-Kwan Law, Betty

    2015-01-01

    With its traditional use in relieving insomnia and anxiety, our previous study has identified onjisaponin B from Radix Polygalae (RP), as a novel autophagic enhancer with potential neuroprotective effects. In current study, we have further identified a novel active fraction from RP, contains 17 major triterpenoid saponins including the onjisaponin B, by the combinational use of cell membrane chromatography (CMC) and ultra-performance liquid chromatography coupled to (quadrupole) time-of-flight mass spectrometry {UHPLC-(Q)TOF-MS}. By exhibiting more potent autophagic effect in cells, the active fraction enhances the clearance of mutant huntingtin, and reduces protein level and aggregation of α-synuclein in a higher extent when compared with onjisaponin B. Here, we have reported for the first time the new application of cell-based CMC and UHPLC-(Q)TOF-MS analysis in identifying new autophagy inducers with neuroprotective effects from Chinese medicinal herb. This result has provided novel insights into the possible pharmacological actions of the active components present in the newly identified active fraction of RP, which may help to improve the efficacy of the traditional way of prescribing RP, and also provide new standard for the quality control of decoction of RP or its medicinal products in the future. PMID:26598009

  15. Cellulose-Based Membranes for Solutes Fractionation

    NASA Astrophysics Data System (ADS)

    Anokhina, T. S.; Yushkin, A. A.; Volkov, V. V.; Antonov, S. V.; Volkov, A. V.

    This work was focused on investigation of industrial cellophane film as a membrane material for solvent nanofiltration. The effect of conditioning of cellophane membranes by stepwise changing of composition of ethanol-water binary mixtures (from ethanol to water and from water to ethanol) was studied. It was shown that such treatment leads to an increase of ethanol permeability more than two orders of magnitude over initial untreated film samples. Treated cellophane membranes possess the ethanol permeability coefficient comparable with the values for highly permeability glassy polymers. Investigation of cellophane swelling in water ethanol solutions allowed to conclude that during the treatment formation of porous in the film takes place due to increase of inter chain distances. Observed high ethanol permeability connected with the fact that formed porous structure remains after the replacement of water with ethanol. Also it was shown that rejection coefficients of a number of dyes (MW 350) were in good agreement with the degree of hydrophobicity/hydrophilicity and ability of the solvent to form hydrogen bonding with the solute molecules. It was demonstrated that cellulose-based membranes can be complimentary for other type of the membranes in fractionation of multi-components solutions.

  16. Modulation of hyaluronan synthase activity in cellular membrane fractions.

    PubMed

    Vigetti, Davide; Genasetti, Anna; Karousou, Evgenia; Viola, Manuela; Clerici, Moira; Bartolini, Barbara; Moretto, Paola; De Luca, Giancarlo; Hascall, Vincent C; Passi, Alberto

    2009-10-30

    Hyaluronan (HA), the only non-sulfated glycosaminoglycan, is involved in morphogenesis, wound healing, inflammation, angiogenesis, and cancer. In mammals, HA is synthesized by three homologous HA synthases, HAS1, HAS2, and HAS3, that polymerize the HA chain using UDP-glucuronic acid and UDP-N-acetylglucosamine as precursors. Since the amount of HA is critical in several pathophysiological conditions, we developed a non-radioactive assay for measuring the activity of HA synthases (HASs) in eukaryotic cells and addressed the question of HAS activity during intracellular protein trafficking. We prepared three cellular fractions: plasma membrane, cytosol (containing membrane proteins mainly from the endoplasmic reticulum and Golgi), and nuclei. After incubation with UDP-sugar precursors, newly synthesized HA was quantified by polyacrylamide gel electrophoresis of fluorophore-labeled saccharides and high performance liquid chromatography. This new method measured HAS activity not only in the plasma membrane fraction but also in the cytosolic membranes. This new technique was used to evaluate the effects of 4-methylumbeliferone, phorbol 12-myristate 13-acetate, interleukin 1beta, platelet-derived growth factor BB, and tunicamycin on HAS activities. We found that HAS activity can be modulated by post-translational modification, such as phosphorylation and N-glycosylation. Interestingly, we detected a significant increase in HAS activity in the cytosolic membrane fraction after tunicamycin treatment. Since this compound is known to induce HA cable structures, this result links HAS activity alteration with the capability of the cell to promote HA cable formation.

  17. Composite fuel cell membranes

    DOEpatents

    Plowman, K.R.; Rehg, T.J.; Davis, L.W.; Carl, W.P.; Cisar, A.J.; Eastland, C.S.

    1997-08-05

    A bilayer or trilayer composite ion exchange membrane is described suitable for use in a fuel cell. The composite membrane has a high equivalent weight thick layer in order to provide sufficient strength and low equivalent weight surface layers for improved electrical performance in a fuel cell. In use, the composite membrane is provided with electrode surface layers. The composite membrane can be composed of a sulfonic fluoropolymer in both core and surface layers.

  18. Composite fuel cell membranes

    DOEpatents

    Plowman, Keith R.; Rehg, Timothy J.; Davis, Larry W.; Carl, William P.; Cisar, Alan J.; Eastland, Charles S.

    1997-01-01

    A bilayer or trilayer composite ion exchange membrane suitable for use in a fuel cell. The composite membrane has a high equivalent weight thick layer in order to provide sufficient strength and low equivalent weight surface layers for improved electrical performance in a fuel cell. In use, the composite membrane is provided with electrode surface layers. The composite membrane can be composed of a sulfonic fluoropolymer in both core and surface layers.

  19. Gas phase fractionation method using porous ceramic membrane

    DOEpatents

    Peterson, Reid A.; Hill, Jr., Charles G.; Anderson, Marc A.

    1996-01-01

    Flaw-free porous ceramic membranes fabricated from metal sols and coated onto a porous support are advantageously used in gas phase fractionation methods. Mean pore diameters of less than 40 .ANG., preferably 5-20 .ANG. and most preferably about 15 .ANG., are permeable at lower pressures than existing membranes. Condensation of gases in small pores and non-Knudsen membrane transport mechanisms are employed to facilitate and increase membrane permeability and permselectivity.

  20. Menin localization in cell membrane compartment

    PubMed Central

    He, Xin; Wang, Lei; Yan, Jizhou; Yuan, Chaoxing; Witze, Eric S.; Hua, Xianxin

    2016-01-01

    ABSTRACT Menin is encoded by the MEN1 gene, which is mutated in an inherited human syndrome, multiple endocrine neoplasia type 1(MEN1). Menin is primarily nuclear protein, acting as a tumor suppressor in endocrine organs, but as an oncogenic factor in the mixed lineage leukemia, in a tissue-specific manner. Recently, the crystal structures of menin with different binding partners reveal menin as a key scaffold protein that functionally interacts with various partners to regulate gene transcription in the nucleus. However, outside the nucleus, menin also regulates multiple signaling pathways that traverse the cell surface membrane. The precise nature regarding to how menin associates with the membrane fraction is poorly understood. Here we show that a small fraction of menin associates with the cell membrane fraction likely via serine palmitoylation. Moreover, the majority of the membrane-associated menin may reside inside membrane vesicles, as menin is protected from trypsin-mediated proteolysis, but disruption of the membrane fraction using detergent abolishes the detection. Consistently, cellular staining for menin also reveals the distribution of menin in the cell membrane and the punctate-like cell organelles. Our findings suggest that part of intracellular menin associates with the cell membrane peripherally as well as resides within the membrane vesicles. PMID:26560942

  1. Free-flow electrophoresis for fractionation of Arabidopsis thaliana membranes.

    PubMed

    Bardy, N; Carrasco, A; Galaud, J P; Pont-Lezica, R; Canut, H

    1998-06-01

    Highly purified tonoplast and plasma membrane vesicles were isolated from microsomes of Arabidopsis thaliana by preparative free-flow electrophoresis. The most electronegative fractions were identified as tonoplast using nitrate-inhibited Mg2+-ATPase as enzyme marker. The least electronegative fractions were identified as plasma membrane using glucan-synthase II, UDPG: sterol-glucosyl-transferase, and vanadate-inhibited Mg2+-ATPase as enzyme markers. Other membrane markers, latent inosine-5'-diphosphatase (Golgi), NADPH-cytochrome-c reductase (endoplasmic reticulum) and cytochrome-c oxidase (mitochondria) were recovered in the fractions intermediate between tonoplast and plasma membrane. Immunoblot analysis of membrane fractions by antibodies directed against tonoplast and plasma membrane proteins confirmed the nature and the purity of the isolated membranes. The cytoskeletal protein actin, which was also identified by immunoblotting, was found to be specifically attached to the plasma membrane vesicles. The structural and functional integrity of the isolated membranes from Arabidopsis thaliana is discussed in the light of results obtained for the location of receptors and enzymes, or for the determination of ligand binding activity.

  2. Membrane in cancer cells

    SciTech Connect

    Galeotti, T.; Cittadini, A.; Neri, G.; Scarpa, A.

    1988-01-01

    This book contains papers presented at a conference on membranes in cancer cells. Topics covered include Oncogenies, hormones, and free-radical processes in malignant transformation in vitro and Superoxide onion may trigger DNA strand breaks in human granulorytes by acting as a membrane target.

  3. Membrane Cells for Brine Electrolysis.

    ERIC Educational Resources Information Center

    Tingle, M.

    1982-01-01

    Membrane cells were developed as alternatives to mercury and diaphragm cells for the electrolysis of brine. Compares the three types of cells, focusing on the advantages and disadvantages of membrane cells. (JN)

  4. Membrane Capacitive Memory Alters Spiking in Neurons Described by the Fractional-Order Hodgkin-Huxley Model

    PubMed Central

    Weinberg, Seth H.

    2015-01-01

    Excitable cells and cell membranes are often modeled by the simple yet elegant parallel resistor-capacitor circuit. However, studies have shown that the passive properties of membranes may be more appropriately modeled with a non-ideal capacitor, in which the current-voltage relationship is given by a fractional-order derivative. Fractional-order membrane potential dynamics introduce capacitive memory effects, i.e., dynamics are influenced by a weighted sum of the membrane potential prior history. However, it is not clear to what extent fractional-order dynamics may alter the properties of active excitable cells. In this study, we investigate the spiking properties of the neuronal membrane patch, nerve axon, and neural networks described by the fractional-order Hodgkin-Huxley neuron model. We find that in the membrane patch model, as fractional-order decreases, i.e., a greater influence of membrane potential memory, peak sodium and potassium currents are altered, and spike frequency and amplitude are generally reduced. In the nerve axon, the velocity of spike propagation increases as fractional-order decreases, while in a neural network, electrical activity is more likely to cease for smaller fractional-order. Importantly, we demonstrate that the modulation of the peak ionic currents that occurs for reduced fractional-order alone fails to reproduce many of the key alterations in spiking properties, suggesting that membrane capacitive memory and fractional-order membrane potential dynamics are important and necessary to reproduce neuronal electrical activity. PMID:25970534

  5. Membrane capacitive memory alters spiking in neurons described by the fractional-order Hodgkin-Huxley model.

    PubMed

    Weinberg, Seth H

    2015-01-01

    Excitable cells and cell membranes are often modeled by the simple yet elegant parallel resistor-capacitor circuit. However, studies have shown that the passive properties of membranes may be more appropriately modeled with a non-ideal capacitor, in which the current-voltage relationship is given by a fractional-order derivative. Fractional-order membrane potential dynamics introduce capacitive memory effects, i.e., dynamics are influenced by a weighted sum of the membrane potential prior history. However, it is not clear to what extent fractional-order dynamics may alter the properties of active excitable cells. In this study, we investigate the spiking properties of the neuronal membrane patch, nerve axon, and neural networks described by the fractional-order Hodgkin-Huxley neuron model. We find that in the membrane patch model, as fractional-order decreases, i.e., a greater influence of membrane potential memory, peak sodium and potassium currents are altered, and spike frequency and amplitude are generally reduced. In the nerve axon, the velocity of spike propagation increases as fractional-order decreases, while in a neural network, electrical activity is more likely to cease for smaller fractional-order. Importantly, we demonstrate that the modulation of the peak ionic currents that occurs for reduced fractional-order alone fails to reproduce many of the key alterations in spiking properties, suggesting that membrane capacitive memory and fractional-order membrane potential dynamics are important and necessary to reproduce neuronal electrical activity.

  6. Inhibition of adhesion of Clostridium difficile to human intestinal cells after treatment with serum and intestinal fluid isolated from mice immunized with nontoxigenic C. difficile membrane fraction.

    PubMed

    Senoh, Mitsutoshi; Iwaki, Masaaki; Yamamoto, Akihiko; Kato, Haru; Fukuda, Tadashi; Shibayama, Keigo

    2015-04-01

    Diarrhea and pseudomembrane colitis caused by Clostridium difficile infection is a global health concern because of the high recurrence rate after standard antibiotic therapy. Vaccination presents a powerful countermeasure against disease recurrence. In this study, mice vaccinated with the nontoxigenic C. difficile membrane fraction generated a marked immune response to the antigen, as demonstrated by the serum IgG and intestinal fluid IgA levels. Significantly, pretreatment with harvested IgG- and IgA-containing fluids was sufficient to prevent in vitro adhesion of C. difficile to human Caco-2 intestinal cells. These results highlight the potential of nontoxigenic C. difficile membrane fraction as a vaccine candidate for C. difficile infection.

  7. Fuel cell membrane humidification

    DOEpatents

    Wilson, Mahlon S.

    1999-01-01

    A polymer electrolyte membrane fuel cell assembly has an anode side and a cathode side separated by the membrane and generating electrical current by electrochemical reactions between a fuel gas and an oxidant. The anode side comprises a hydrophobic gas diffusion backing contacting one side of the membrane and having hydrophilic areas therein for providing liquid water directly to the one side of the membrane through the hydrophilic areas of the gas diffusion backing. In a preferred embodiment, the hydrophilic areas of the gas diffusion backing are formed by sewing a hydrophilic thread through the backing. Liquid water is distributed over the gas diffusion backing in distribution channels that are separate from the fuel distribution channels.

  8. [Effect of different organic fraction on membrane flux declines].

    PubMed

    Zhou, Xian-Jiao; Dong, Bing-Zhi

    2009-02-15

    Organic matter in the tap water was isolated into strongly hydrophobic acids, weakly hydrophobic acids, charged hydrophilic and neutral hydrophilic by DAX-8, XAD-4 and IRA-958 synthetic resins. Filtration tests using polyvinylidene fluoride (PVDF), polyethersulphone (PES) and cellulose acetate (CA) membranes were conducted to investigate the contribution of different organic fractions to membrane fouling. The results show that in filtration of raw water, flux declines with PES, PVDF and CA membrane are 67%, 59% and 19% of the initial flux, indicating that the more hydrophobic membrane resulted in more severe fouling. For the effect of different fractions on flux, flux decline with neutral hydrophilic is 41%-75% of the initial flux, whereas weakly hydrophobic acids is 6%-33%, suggesting that neutral hydrophilic has a great impact on filtration flux. Among three membranes tested, CA membrane shows the lowest flux decline compared with other membranes in spite of rejection of as high as 14.69% of neutral hydrophilic, suggesting that the extent of flux decline may not be associated with the total amount of NOM removed. The mechanism of fouling was discussed and found that the neutral hydrophilic fraction with greater than 3 x 10(4) of molecular weight caused a significant flux decline, through blocking the pore for the MF or UF having greater relative molecular mass cut-off (MWCO), but resulted in a little impact on flux with the UF having lower MWCO, through forming cake layer on the surface of membrane due to not entering the inside of pore.

  9. Peptide separations by on-line MudPIT compared to isoelectric focusing in an off-gel format: application to a membrane-enriched fraction from C2C12 mouse skeletal muscle cells.

    PubMed

    Elschenbroich, Sarah; Ignatchenko, Vladimir; Sharma, Parveen; Schmitt-Ulms, Gerold; Gramolini, Anthony O; Kislinger, Thomas

    2009-10-01

    High-resolution peptide separation is pivotal for successful shotgun proteomics. The need for capable techniques propels invention and improvement of ever more sophisticated approaches. Recently, Agilent Technologies has introduced the OFFGEL fractionator, which conducts peptide separation by isoelectric focusing in an off-gel setup. This platform has been shown to accomplish high resolution of peptides for diverse sample types, yielding valuable advantages over comparable separation techniques. In this study, we deliver the first comparison of the newly emerging OFFGEL approach to the well-established on-line MudPIT platform. Samples from a membrane-enriched fraction isolated from murine C2C12 cells were subjected to replicate analysis by OFFGEL (12 fractions, pH 3-10) followed by RP-LC-MS/MS or 12-step on-line MudPIT. OFFGEL analyses yielded 1398 proteins (identified by 10,269 peptides), while 1428 proteins (11,078 peptides) were detected with the MudPIT approach. Thus, our data shows that both platforms produce highly comparable results in terms of protein/peptide identifications and reproducibility for the sample type analyzed. We achieve more accurate peptide focusing after OFFGEL fractionation with 88% of all peptides binned to a single fraction, as compared to 61% of peptides detected in only one step in MudPIT analyses. Our study suggests that both platforms are equally capable of high quality peptide separation of a sample with medium complexity, rendering them comparably valuable for comprehensive proteomic analyses.

  10. Biological Fuel Cells and Membranes.

    PubMed

    Ghassemi, Zahra; Slaughter, Gymama

    2017-01-17

    Biofuel cells have been widely used to generate bioelectricity. Early biofuel cells employ a semi-permeable membrane to separate the anodic and cathodic compartments. The impact of different membrane materials and compositions has also been explored. Some membrane materials are employed strictly as membrane separators, while some have gained significant attention in the immobilization of enzymes or microorganisms within or behind the membrane at the electrode surface. The membrane material affects the transfer rate of the chemical species (e.g., fuel, oxygen molecules, and products) involved in the chemical reaction, which in turn has an impact on the performance of the biofuel cell. For enzymatic biofuel cells, Nafion, modified Nafion, and chitosan membranes have been used widely and continue to hold great promise in the long-term stability of enzymes and microorganisms encapsulated within them. This article provides a review of the most widely used membrane materials in the development of enzymatic and microbial biofuel cells.

  11. Biological Fuel Cells and Membranes

    PubMed Central

    Ghassemi, Zahra; Slaughter, Gymama

    2017-01-01

    Biofuel cells have been widely used to generate bioelectricity. Early biofuel cells employ a semi-permeable membrane to separate the anodic and cathodic compartments. The impact of different membrane materials and compositions has also been explored. Some membrane materials are employed strictly as membrane separators, while some have gained significant attention in the immobilization of enzymes or microorganisms within or behind the membrane at the electrode surface. The membrane material affects the transfer rate of the chemical species (e.g., fuel, oxygen molecules, and products) involved in the chemical reaction, which in turn has an impact on the performance of the biofuel cell. For enzymatic biofuel cells, Nafion, modified Nafion, and chitosan membranes have been used widely and continue to hold great promise in the long-term stability of enzymes and microorganisms encapsulated within them. This article provides a review of the most widely used membrane materials in the development of enzymatic and microbial biofuel cells. PMID:28106711

  12. The First Cell Membranes

    NASA Astrophysics Data System (ADS)

    Deamer, David; Dworkin, Jason P.; Sandford, Scott A.; Bernstein, Max P.; Allamandola, Louis J.

    2002-12-01

    Organic compounds are synthesized in the interstellar medium and can be delivered to planetary surfaces such as the early Earth, where they mix with endogenous species. Some of these compounds are amphiphilic, having polar and nonpolar groups on the same molecule. Amphiphilic compounds spontaneously self-assemble into more complex structures such as bimolecular layers, which in turn form closed membranous vesicles. The first forms of cellular life required self-assembled membranes that were likely to have been produced from amphiphilic compounds on the prebiotic Earth. Laboratory simulations show that such vesicles readily encapsulate functional macromolecules, including nucleic acids and polymerases. The goal of future investigations will be to fabricate artificial cells as models of the origin of life.

  13. The First Cell Membranes

    NASA Technical Reports Server (NTRS)

    Deamer, David; Dworkin, Jason P.; Sandford, Scott A.; Bernstein, Max P.; Allamandola, Louis J.

    2004-01-01

    Organic compounds are synthesized in the interstellar medium and can be delivered to planetary surfaces such as the early Earth, where they mix with endogenous organic mixtures. Some of these compounds are amphiphilic, having polar and non-polar groups on the same molecule. Amphiphilic compounds spontaneously self-assembly into more complex structures such as bimolecular layers, which in turn form closed membranous vesicles. The first forms of cellular life required self-assembled membranes that were likely to be available on the prebiotic Earth. Laboratory simulations show that such vesicles readily encapsulate functional macromolecules, including nucleic acids and polymerases. A goal of future investigations is to fabricate artificial cells as models of the origin of life.

  14. Cell Membrane Softening in Cancer Cells

    NASA Astrophysics Data System (ADS)

    Schmidt, Sebastian; Händel, Chris; Käs, Josef

    Biomechanical properties are useful characteristics and regulators of the cell's state. Current research connects mechanical properties of the cytoskeleton to many cellular processes but does not investigate the biomechanics of the plasma membrane. We evaluated thermal fluctuations of giant plasma membrane vesicles, directly derived from the plasma membranes of primary breast and cervical cells and observed a lowered rigidity in the plasma membrane of malignant cells compared to non-malignant cells. To investigate the specific role of membrane rigidity changes, we treated two cell lines with the Acetyl-CoA carboxylase inhibitor Soraphen A. It changed the lipidome of cells and drastically increased membrane stiffness by up regulating short chained membrane lipids. These altered cells had a decreased motility in Boyden chamber assays. Our results indicate that the thermal fluctuations of the membrane, which are much smaller than the fluctuations driven by the cytoskeleton, can be modulated by the cell and have an impact on adhesion and motility.

  15. Isolation of plasma membrane fractions from the intestinal epithelial model T84.

    PubMed

    Kaoutzani, P; Parkos, C A; Delp-Archer, C; Madara, J L

    1993-05-01

    The human intestinal epithelial cell line T84 is widely used as a model for studies of Cl- secretion and crypt cell biology. We report a fractionation approach that permits separation of purified apical and basolateral T84 plasma membrane domains. T84 cellular membranes were isolated by nitrogen cavitation and differential centrifugation from monolayers grown on permeable supports. Membranes were then fractionated by isopycnic sucrose density gradient sedimentation, and fractions were assessed, using enzymatic and Western blot techniques, for apical (alkaline phosphatase) and basolateral (Na(+)-K(+)-ATPase) plasma membrane markers and for cytosolic, lysosomal, Golgi, and mitochondrial markers. Buffer conditions were defined that permitted separation of enriched apical and basolateral markers. The validity of the selected markers for the apical and basolateral domains was verified by selective apical and basolateral surface labeling studies using trace iodinated wheat germ agglutinin or biotinylation. This approach allows for separation of apical and basolateral plasma membranes of T84 cells for biochemical analyses and should thus be of broad utility in studies of this model polarized and transporting epithelium.

  16. [The influence of N-, S-containing chinasolone derivatives (NC-224) on the biochemical and physicochemical parameters of membrane endoplasmatic reticulum and nuclear chromatine fractions of rats liver cells in conditions of its injury by tetrachloromethane].

    PubMed

    Gubs'kyî, Iu I; Goriushko, G G; Belenichev, I F; Kovalenko, S I; Litvinova, N V; Marchenko, O M; Kurapova, T M; Babenko, L P; Velychko, O M

    2010-01-01

    Using biochemical and physicochemical methods of investigation in vivo, the effect of the substance NC-224, N-, S-chinasolone-derivative, on the lipoperoxidation activity in rat liver endoplasmatic reticulum membranes and nuclear chromatin fractions under tetrachloromethane intoxication have been studied. It was shown that NC-224 has pronounced antioxidant activity which is the biochemical basis of the substance membrane- and genome-protective effects and its ability to restore physicochemical properties of the surface and hydrophobic zones of hepatocyte membranes and structural parameter nuclear chromatin fractions in the conditions of chemical liver injury.

  17. Origin of subdiffusion of water molecules on cell membrane surfaces

    PubMed Central

    Yamamoto, Eiji; Akimoto, Takuma; Yasui, Masato; Yasuoka, Kenji

    2014-01-01

    Water molecules play an important role in providing unique environments for biological reactions on cell membranes. It is widely believed that water molecules form bridges that connect lipid molecules and stabilize cell membranes. Using all-atom molecular dynamics simulations, we show that translational and rotational diffusion of water molecules on lipid membrane surfaces exhibit subdiffusion and aging. Moreover, we provide evidence that both divergent mean trapping time (continuous-time random walk) and long-correlated noise (fractional Brownian motion) contribute to this subdiffusion. These results suggest that subdiffusion on cell membranes causes the water retardation, an enhancement of cell membrane stability, and a higher reaction efficiency. PMID:24739933

  18. Fuel cell with ionization membrane

    NASA Technical Reports Server (NTRS)

    Hartley, Frank T. (Inventor)

    2007-01-01

    A fuel cell is disclosed comprising an ionization membrane having at least one area through which gas is passed, and which ionizes the gas passing therethrough, and a cathode for receiving the ions generated by the ionization membrane. The ionization membrane may include one or more openings in the membrane with electrodes that are located closer than a mean free path of molecules within the gas to be ionized. Methods of manufacture are also provided.

  19. Role of polyadenylic acid in a deoxyribonucleic acid-membrane fraction extracted from pneumococci.

    PubMed Central

    Firshein, W; Meyer, B; Epner, E; Viggiani, J

    1976-01-01

    After the addition of radioactive polyadenylic acid to cell suspensions of pneumocci, part of the radioactivity becomes associated with a deoxyribonucleic acid (DNA)-membrane fraction extracted from the cells. A variety of techniques show that a portion of this associated radioactivity may represent oligoadenylates complexed to DNA, probaby as part of a ribonucleic acid (RNA) component. Polyadenylic acid, which had previously been shown to enhance DNA synthesis in cell suspensions (Firshein and Benson, 1968), also enhances the extent of DNA synthesis by the DNA-membrane fraction in vitro under specific conditions of concentration and conformation. The mechanism of action of this enhancement may be related to the ability of oligoadenylates to increase the number of initiation sites for DNA replication by stimulating the production of an RNA primer, thus providing additional 3'-OH groups with which DNA polymerase can react. PMID:6428

  20. Derivation of extracellular fluid volume fraction and equivalent dielectric constant of the cell membrane from dielectric properties of the human body. Part 2: A preliminary study for tracking the progression of surgical tissue injury.

    PubMed

    Tatara, T; Tsuzaki, K

    2000-07-01

    A study is conducted to determine whether the extracellular fluid (ECF) volume fraction and equivalent dielectric constant of the cell membrane epsilon m, derived from the dielectric properties of the human body can track the progression of surgical tissue injury. Frequency-dependent dielectric constants and electrical conductivities of body segments are obtained at surgical (trunk) and non-surgical sites (arm and leg) from five patients who have undergone oesophageal resections, before and at the end of surgery and on the day after the operation. The ECF volume fraction and the equivalent epsilon m of body segments are estimated by fitting the dielectric data for body segments to the cell suspension model incorporating fat tissue, and their time-course changes are compared between body segments. By the day after the operation, the estimated ECF volume fraction has increased in all body segments compared with that before surgery, by 0.13 in the arm, 0.16 in the trunk and 0.14 in the leg (p < 0.05), indicating postoperative fluid accumulation in the extracellular space. In contrast, the estimated equivalent epsilon m shows a different time course between body segments on the day after the operation, characterised by a higher change ratio of epsilon m of the trunk (1.34 +/- 0.66, p < 0.05), from that of the arm (0.66 +/- 0.34) and leg (0.61 +/- 0.11). The results suggest that the equivalent epsilon m of a body segment at a surgical site can track pathophysiological cell changes following surgical tissue injury.

  1. Role of deoxyribonucleic acid ligase in a doxyribonucleic acid membrane fraction extracted from pneumococci.

    PubMed Central

    Greene, M; Firshein, W

    1976-01-01

    Deoxyribonucleic acid (DNA) ligase has been detected in a DNA membrane fraction extracted from Pneumococcus. The specific activity of the enzyme in this fraction is 10-fold greater than in the remaining cell extract. It remains firmly bound (with other enzymes) to the complex after a purification procedure in which a considerable percentage of the macromolecules are dissociated. The ligase acts in two ways in the DNA membrane fraction in vitro. One, it catalyzes the linkage of small-molecular-weight pieces of newly synthesized DNA into heavier-molecular-weight DNA strands as shown by others (M Gellert, 1976; R. Okazaki, A. Sugino, S. Hirose, T. Okazaki, Y. Imae, R. Kainuma-Kuroda, T. Ogawa, M. Arisawa, and Y. Kurosowa, 1973; B. Olivera and I. Lehman, 14; and A. Sugino, S. Hirose, and R. Okazaki, 1972) and, two, it protects DNA from degradation by deoxyribonucleases. This latter effect is due to a competition between the ability of the nucleases to degrade DNA and the ability of DNA ligase to seal the nicks produced by these degradative enzymes. The ligase acts cooperatively with other enzymes in the DNA membrane fraction to synthesize DNA. PMID:4433

  2. In vitro enzymatic reduction kinetics of mineral oxides by membrane fractions from Shewanella oneidensis MR-1

    NASA Astrophysics Data System (ADS)

    Ruebush, Shane S.; Icopini, Gary A.; Brantley, Susan L.; Tien, Ming

    2006-01-01

    This study documents the first example of in vitro solid-phase mineral oxide reduction by enzyme-containing membrane fractions. Previous in vitro studies have only reported the reduction of aqueous ions. Total membrane (TM) fractions from iron-grown cultures of Shewanella oneidensis MR-1 were isolated and shown to catalyze the reduction of goethite, hematite, birnessite, and ramsdellite/pyrolusite using formate. In contrast, nicotinamide adenine dinucleotide (NADH) and succinate cannot function as electron donors. The significant implications of observations related to this cell-free system are: (i) both iron and manganese mineral oxides are reduced by the TM fraction, but aqueous U(VI) is not; (ii) TM fractions from anaerobically grown, but not aerobically grown, cells can reduce the mineral oxides; (iii) electron shuttles and iron chelators are not needed for this in vitro reduction, documenting conclusively that reduction can occur by direct contact with the mineral oxide; (iv) electron shuttles and EDTA stimulate the in vitro Fe(III) reduction, documenting that exogenous molecules can enhance rates of enzymatic mineral reduction; and (v) multiple membrane components are involved in solid-phase oxide reduction. The membrane fractions, consisting of liposomes of cytoplasmic and outer membrane segments, contain at least 100 proteins including the enzyme that oxidizes formate, formate dehydrogenase. Mineral oxide reduction was inhibited by the addition of detergent Triton X-100, which solubilizes membranes and their associated proteins, consistent with the involvement of multiple electron carriers that are disrupted by detergent addition. In contrast, formate dehydrogenase activity was not inhibited by Triton X-100. The addition of anthraquinone-2,6-disulfonate (AQDS) and menaquinone-4 was unable to restore activity; however, menadione (MD) restored 33% of the activity. The addition of AQDS and MD to reactions without added detergent increased the rate of goethite

  3. In Vitro Enzymatic Reduction Kinetics of Mineral Oxides by Membrane Fractions from Shewanella oneidensis MR-1

    SciTech Connect

    Ruebush,S.; Icopini, G.; Brantley, S.; Tien, M.

    2006-01-01

    This study documents the first example of in vitro solid-phase mineral oxide reduction by enzyme-containing membrane fractions. Previous in vitro studies have only reported the reduction of aqueous ions. Total membrane (TM) fractions from iron-grown cultures of Shewanella oneidensis MR-1 were isolated and shown to catalyze the reduction of goethite, hematite, birnessite, and ramsdellite/pyrolusite using formate. In contrast, nicotinamide adenine dinucleotide (NADH) and succinate cannot function as electron donors. The significant implications of observations related to this cell-free system are: (i) both iron and manganese mineral oxides are reduced by the TM fraction, but aqueous U(VI) is not; (ii) TM fractions from anaerobically grown, but not aerobically grown, cells can reduce the mineral oxides; (iii) electron shuttles and iron chelators are not needed for this in vitro reduction, documenting conclusively that reduction can occur by direct contact with the mineral oxide; (iv) electron shuttles and EDTA stimulate the in vitro Fe(III) reduction, documenting that exogenous molecules can enhance rates of enzymatic mineral reduction; and (v) multiple membrane components are involved in solid-phase oxide reduction. The membrane fractions, consisting of liposomes of cytoplasmic and outer membrane segments, contain at least 100 proteins including the enzyme that oxidizes formate, formate dehydrogenase. Mineral oxide reduction was inhibited by the addition of detergent Triton X-100, which solubilizes membranes and their associated proteins, consistent with the involvement of multiple electron carriers that are disrupted by detergent addition. In contrast, formate dehydrogenase activity was not inhibited by Triton X-100. The addition of anthraquinone-2,6-disulfonate (AQDS) and menaquinone-4 was unable to restore activity; however, menadione (MD) restored 33% of the activity. The addition of AQDS and MD to reactions without added detergent increased the rate of goethite

  4. The Molecules of the Cell Membrane.

    ERIC Educational Resources Information Center

    Bretscher, Mark S.

    1985-01-01

    Cell membrane molecules form a simple, two-dimensional liquid controlling what enters and leaves the cell. Discusses cell membrane molecular architecture, plasma membranes, epithelial cells, cycles of endocytosis and exocytosis, and other topics. Indicates that some cells internalize, then recycle, membrane area equivalent to their entire surface…

  5. ISOLATION OF PLASMA MEMBRANE FRAGMENTS FROM HELA CELLS

    PubMed Central

    Boone, Charles W.; Ford, Lincoln E.; Bond, Howard E.; Stuart, Donald C.; Lorenz, Dianne

    1969-01-01

    A method for isolating plasma membrane fragments from HeLa cells is described. The procedure starts with the preparation of cell membrane "ghosts," obtained by gentle rupture of hypotonically swollen cells, evacuation of most of the cell contents by repeated washing, and isolation of the ghosts on a discontinuous sucrose density gradient. The ghosts are then treated by minimal sonication (5 sec) at pH 8.6, which causes the ghost membranes to pinch off into small vesicles but leaves any remaining larger intracellular particulates intact and separable by differential centrifugation. The ghost membrane vesicles are then subjected to isopycnic centrifugation on a 20–50% w/w continuous sucrose gradient in tris-magnesium buffer, pH 8.6. A band of morphologically homogeneous smooth vesicles, derived principally from plasma membrane, is recovered at 30–33% (peak density = 1.137). The plasma membrane fraction contained a Na-K-activated ATPase activity of 1.5 µmole Pi/hr per mg, 3% RNA, and 13.8% of the NADH-cytochrome c reductase activity of a heavier fraction from the same gradient which contained mitochondria and rough endoplasmic vesicles. The plasma membranes of viable HeLa cells were marked with 125I-labeled horse antibody and followed through the isolation procedure. The specific antibody binding of the plasma membrane vesicle fraction was increased 49-fold over that of the original whole cells. PMID:4239370

  6. Tannin-rich fraction from pomegranate rind damages membrane of Listeria monocytogenes.

    PubMed

    Li, Guanghui; Xu, Yunfeng; Wang, Xin; Zhang, Baigang; Shi, Chao; Zhang, Weisong; Xia, Xiaodong

    2014-04-01

    Pomegranate rind has been reported to inhibit several foodborne pathogens, and its antimicrobial activity has been attributed mainly to its tannin fraction. This study aimed to investigate the antimicrobial activity of the tannin-rich fraction from pomegranate rind (TFPR) against Listeria monocytogenes and its mechanism of action. The tannin-related components of TFPR were analyzed by high-performance liquid chromatography and liquid chromatography-mass spectrometry, and the minimum inhibitory concentration (MIC) of TFPR was determined using the agar dilution method. Extracellular potassium concentration, the release of cell constituents, intra- and extracellular ATP concentrations, membrane potential, and intracellular pH (pHin) were measured to elucidate a possible antibacterial mechanism. Punicalagin (64.2%, g/g) and ellagic acid (3.1%, g/g) were detected in TFPR, and the MICs of TFPR were determined to be 1.25-5.0 mg/mL for different L. monocytogenes strains. Treatment with TFPR induced a decrease of the intracellular ATP concentration, an increase of the extracellular concentrations of potassium and ATP, and the release of cell constituents. A reduction of pHin and cell membrane hyperpolarization were observed after treatment. Electron microscopic observations showed that the cell membrane structures of L. monocytogenes were apparently impaired by TFPR. It is concluded that TFPR could destroy the integrity of the cell membrane of L. monocytogenes, leading to a loss of cell homeostasis. These findings indicate that TFPR has the potential to be used as a food preservative in order to control L. monocytogenes contamination in food and reduce the risk of listeriosis.

  7. Physical principles of membrane remodelling during cell mechanoadaptation

    PubMed Central

    Kosmalska, Anita Joanna; Casares, Laura; Elosegui-Artola, Alberto; Thottacherry, Joseph Jose; Moreno-Vicente, Roberto; González-Tarragó, Víctor; del Pozo, Miguel Ángel; Mayor, Satyajit; Arroyo, Marino; Navajas, Daniel; Trepat, Xavier; Gauthier, Nils C.; Roca-Cusachs, Pere

    2015-01-01

    Biological processes in any physiological environment involve changes in cell shape, which must be accommodated by their physical envelope—the bilayer membrane. However, the fundamental biophysical principles by which the cell membrane allows for and responds to shape changes remain unclear. Here we show that the 3D remodelling of the membrane in response to a broad diversity of physiological perturbations can be explained by a purely mechanical process. This process is passive, local, almost instantaneous, before any active remodelling and generates different types of membrane invaginations that can repeatedly store and release large fractions of the cell membrane. We further demonstrate that the shape of those invaginations is determined by the minimum elastic and adhesive energy required to store both membrane area and liquid volume at the cell–substrate interface. Once formed, cells reabsorb the invaginations through an active process with duration of the order of minutes. PMID:26073653

  8. High-speed pectic enzyme fractionation by immobilised metal ion affinity membranes.

    PubMed

    Camperi, S A; Grasselli, M; Cascone, O

    2000-01-01

    Immobilised metal ion affinity polysulfone hollow-fibre membranes, with a high capacity for protein adsorption, were prepared and their utilisation for commercial pectic enzyme fractionation was studied. The pass-through fraction containing pectinlyase is useful for fruit-juice clarification without methanol production on account of pectinesterase being retained by the IDA-Cu2+ membrane.

  9. Proteomics and Phosphoproteomics Analysis of Human Lens Fiber Cell Membranes

    PubMed Central

    Wang, Zhen; Han, Jun; David, Larry L.; Schey, Kevin L.

    2013-01-01

    Purpose. The human lens fiber cell insoluble membrane fraction contains important membrane proteins, cytoskeletal proteins, and cytosolic proteins that are strongly associated with the membrane. The purpose of this study was to characterize the lens fiber cell membrane proteome and phosphoproteome from human lenses. Methods. HPLC-mass spectrometry–based multidimensional protein identification technology (MudPIT), without or with phosphopeptide enrichment, was applied to study the proteome and phosphoproteome of lens fiber cell membranes, respectively. Results. In total, 951 proteins were identified, including 379 integral membrane and membrane-associated proteins. Enriched gene categories and pathways based on the proteomic analysis include carbohydrate metabolism (glycolysis/gluconeogenesis, pentose phosphate pathway, pyruvate metabolism), proteasome, cell-cell signaling and communication (GTP binding, gap junction, focal adhesion), glutathione metabolism, and actin regulation. The combination of TiO2 phosphopeptide enrichment and MudPIT analysis revealed 855 phosphorylation sites on 271 proteins, including 455 phosphorylation sites that have not been previously identified. PKA, PKC, CKII, p38MAPK, and RSK are predicted as the major kinases for phosphorylation on the sites identified in the human lens membrane fraction. Conclusions. The results presented herein significantly expand the characterized proteome and phosphoproteome of the human lens fiber cell and provide a valuable reference for future research in studies of lens development and disease. PMID:23349431

  10. Hydrophilic fraction of natural organic matter causing irreversible fouling of microfiltration and ultrafiltration membranes.

    PubMed

    Yamamura, Hiroshi; Okimoto, Kenji; Kimura, Katsuki; Watanabe, Yoshimasa

    2014-05-01

    Although membrane filtration is a promising technology in the field of drinking water treatment, persistent membrane fouling remains a major disadvantage. For more efficient operation, causative agents of membrane fouling need to be identified. Membrane fouling can be classified into physically reversible and irreversible fouling on basis of the removability of the foulants by physical cleaning. Four types of natural organic matter (NOM) in river water used as a source of drinking water were fractionated into hydrophobic and hydrophilic fractions, and their potential to develop irreversible membrane fouling was evaluated by a bench-scale filtration experiment together with spectroscopic and chromatographic analyses. In this study, only dissolved NOM was investigated without consideration of interactions of NOM fractions with particulate matter. Results demonstrated that despite identical total organic carbon (TOC), fouling development trends were significantly different between hydrophilic and hydrophobic fractions. The hydrophobic fractions did not increase membrane resistance, while the hydrophilic fractions caused severe loss of membrane permeability. These results were identical with the case when the calcium was added to hydrophobic and hydrophilic fractions. The largest difference in NOM characteristics between hydrophobic and hydrophilic fractions was the presence or absence of macromolecules; the primary constituent causing irreversible fouling was inferred to be "biopolymers", including carbohydrates and proteins. In addition, the results demonstrated that the extent of irreversible fouling was considerably different depending on the combination of membrane materials and NOM characteristics. Despite identical nominal pore size (0.1 μm), a polyvinylidene fluoride (PVDF) membrane was found to be more rapidly fouled than a PE membrane. This is probably explained by the generation of strong hydrogen bonding between hydroxyl groups of biopolymers and fluorine

  11. Corrugated Membrane Fuel Cell Structures

    SciTech Connect

    Grot, Stephen

    2013-09-30

    One of the most challenging aspects of traditional PEM fuel cell stacks is the difficulty achieving the platinum catalyst utilization target of 0.2 gPt/kWe set forth by the DOE. Good catalyst utilization can be achieved with state-of-the-art catalyst coated membranes (CCM) when low catalyst loadings (<0.3 mg/cm2) are used at a low current. However, when low platinum loadings are used, the peak power density is lower than conventional loadings, requiring a larger total active area and a larger bipolar plate. This results in a lower overall stack power density not meeting the DOE target. By corrugating the fuel cell membrane electrode structure, Ion Power?s goal is to realize both the Pt utilization targets as well as the power density targets of the DOE. This will be achieved by demonstrating a fuel cell single cell (50 cm2) with a twofold increase in the membrane active area over the geometric area of the cell by corrugating the MEA structure. The corrugating structure must be able to demonstrate the target properties of < 10 mOhm-cm2 electrical resistance at > 20 psi compressive strength over the active area, in combination with offering at least 80% of power density that can be achieved by using the same MEA in a flat plate structure. Corrugated membrane fuel cell structures also have the potential to meet DOE power density targets by essentially packaging more membrane area into the same fuel cell volume as compared to conventional stack constructions.

  12. Membrane fractions active in poliovirus RNA replication contain VPg precursor polypeptides

    SciTech Connect

    Takegami, T.; Semler, B.L.; Anderson, C.W.; Wimmer, E.

    1983-01-01

    The poliovirus specific polypeptide P3-9 is of special interest for studies of viral RNA replication because it contains a hydrophobic region and, separated by only seven amino acids from that region, the amino acid sequence of the genome-linked protein VPg. Membraneous complexes of poliovirus-infected HeLa cells that contain poliovirus RNA replicating proteins have been analyzed for the presence of P3-9 by immunoprecipitation. Incubation of a membrane fraction rich in P3-9 with proteinase leaves the C-terminal 69 amino acids of P3-9 intact, an observation suggesting that this portion is protected by its association with the cellular membrane. These studies have also revealed two hitherto undescribed viral polypeptides consisting of amino acid sequences of the P2 andf P3 regions of the polyprotein. Sequence analysis by stepwise Edman degradation show that these proteins are 3b/9 (M/sub r/77,000) and X/9 (M/sub r/50,000). 3b/9 and X/9 are membrane bound and are turned over rapidly and may be direct precursors to proteins P2-X and P3-9 of the RNA replication complex. P2-X, a polypeptide void of hydrophobic amino acid sequences but also found associated with membranes, is rapidly degraded when the membraneous complex is treated with trypsin. It is speculated that P2-X is associated with membranes by its affinity to the N-terminus of P3-9.

  13. Membrane proteins of dense lysosomes from Chinese hamster ovary cells

    SciTech Connect

    Chance, S.C.

    1987-01-01

    In this work membrane proteins from lysosomes were studied in order to gain more information on the biogenesis and intracellular sorting of this class of membrane proteins. Membrane proteins were isolated from a purified population of lysosomes. These proteins were then examined for various co- and post-translational modifications which could serve as potential intracellular sorting signals. Biochemical analysis using marker enzymatic activities detected no plasma membrane, Golgi, endoplasmic reticulum, peroxisomes, mitochondria, or cytosol. Analysis after incorporation of ({sup 3}H)thymidine or ({sup 3}H)uridine detected no nuclei or ribosomes. A fraction containing integral membrane proteins was obtained from the dense lysosomes by extraction with Triton X-114. Twenty-three polypeptides which incorporated both ({sup 35}S)methionine and ({sup 3}H)leucine were detected by SDS PAGE in this membrane fraction, and ranged in molecular weight from 30-130 kDa. After incorporation by cells of various radioactive metabolic precursors, the membrane fraction from dense lysosomes was examined and was found to be enriched in mannose, galactose, fucose, palmitate, myristate, and sulfate, but was depleted in phosphate. The membrane fraction from dense lysosomes was then analyzed by SDS PAGE to determine the apparent molecular weights of modified polypepties.

  14. Plasma membrane associated membranes (PAM) from Jurkat cells contain STIM1 protein is PAM involved in the capacitative calcium entry?

    PubMed

    Kozieł, Katarzyna; Lebiedzinska, Magdalena; Szabadkai, Gyorgy; Onopiuk, Marta; Brutkowski, Wojciech; Wierzbicka, Katarzyna; Wilczyński, Grzegorz; Pinton, Paolo; Duszyński, Jerzy; Zabłocki, Krzysztof; Wieckowski, Mariusz R

    2009-12-01

    A proper cooperation between the plasma membrane, the endoplasmic reticulum and the mitochondria seems to be essential for numerous cellular processes involved in Ca(2+) signalling and maintenance of Ca(2+) homeostasis. A presence of microsomal and mitochondrial proteins together with those characteristic for the plasma membrane in the fraction of the plasma membrane associated membranes (PAM) indicates a formation of stabile interactions between these three structures. We isolated the plasma membrane associated membranes from Jurkat cells and found its significant enrichment in the plasma membrane markers including plasma membrane Ca(2+)-ATPase, Na(+), K(+)-ATPase and CD3 as well as sarco/endoplasmic reticulum Ca(2+) ATPase as a marker of the endoplasmic reticulum membranes. In addition, two proteins involved in the store-operated Ca(2+) entry, Orai1 located in the plasma membrane and an endoplasmic reticulum protein STIM1 were found in this fraction. Furthermore, we observed a rearrangement of STIM1-containing protein complexes isolated from Jurkat cells undergoing stimulation by thapsigargin. We suggest that the inter-membrane compartment composed of the plasma membrane and the endoplasmic reticulum, and isolated as a stabile plasma membrane associated membranes fraction, might be involved in the store-operated Ca(2+) entry, and their formation and rebuilding have an important regulatory role in cellular Ca(2+) homeostasis.

  15. Focus on membrane differentiation and membrane domains in the prokaryotic cell.

    PubMed

    Boekema, Egbert J; Scheffers, Dirk-Jan; van Bezouwen, Laura S; Bolhuis, Henk; Folea, I Mihaela

    2013-01-01

    A summary is presented of membrane differentiation in the prokaryotic cell, with an emphasis on the organization of proteins in the plasma/cell membrane. Many species belonging to the Eubacteria and Archaea have special membrane domains and/or membrane proliferation, which are vital for different cellular processes. Typical membrane domains are found in bacteria where a specific membrane protein is abundantly expressed. Lipid rafts form another example. Despite the rareness of conventional organelles as found in eukaryotes, some bacteria are known to have an intricate internal cell membrane organization. Membrane proliferation can be divided into curvature and invaginations which can lead to internal compartmentalization. This study discusses some of the clearest examples of bacteria with such domains and internal membranes. The need for membrane specialization is highest among the heterogeneous group of bacteria which harvest light energy, such as photosynthetic bacteria and halophilic archaea. Most of the highly specialized membranes and domains, such as the purple membrane, chromatophore and chlorosome, are found in these autotrophic organisms. Otherwise the need for membrane differentiation is lower and variable, except for those structures involved in cell division. Microscopy techniques have given essential insight into bacterial membrane morphology. As microscopy will further contribute to the unraveling of membrane organization in the years to come, past and present technology in electron microscopy and light microscopy is discussed. Electron microscopy was the first to unravel bacterial morphology because it can directly visualize membranes with inserted proteins, which no other technique can do. Electron microscopy techniques developed in the 1950s and perfected in the following decades involve the thin sectioning and freeze fractioning of cells. Several studies from the golden age of these techniques show amazing examples of cell membrane morphology

  16. HeLa cell plasma membranes. I. 5'-Nucleotidase and ouabain-sensitive ATPase as markers for plasma membranes.

    PubMed

    Johnsen, S; Stokke, T; Prydz, H

    1974-11-01

    A method for the preparation of HeLa cell plasma membrane ghosts is described. The purity of the plasma membrane fraction was examined by phase contrast and electron microscopy, by chemical analysis, and by assay of marker enzymes. Data on the composition of the plasma membrane fraction are given. It was observed that the distribution pattern of 5'-nucleotidase activity among the subcellular fractions differed from that of ouabain-sensitive ATPase. In addition, the specific activity of 5'-nucleotidase did not follow the distribution of the membrane ghosts. Thus, this enzyme would seem unsuitable as a plasma membrane marker. A complete balance sheet for marker enzyme activities during the fractionation is necessary for the calculation of increase in specific activity because the activities of both 5'-nucleotidase and ouabain-sensitive ATPase might change during the fractionation procedures.

  17. Strategies for cell membrane functionalization

    PubMed Central

    Armstrong, James PK

    2016-01-01

    The ability to rationally manipulate and augment the cytoplasmic membrane can be used to overcome many of the challenges faced by conventional cellular therapies and provide innovative opportunities when combined with new biotechnologies. The focus of this review is on emerging strategies used in cell functionalization, highlighting both pioneering approaches and recent developments. These will be discussed within the context of future directions in this rapidly evolving field. PMID:27229904

  18. Microalgae fractionation using steam explosion, dynamic and tangential cross-flow membrane filtration.

    PubMed

    Lorente, E; Hapońska, M; Clavero, E; Torras, C; Salvadó, J

    2017-03-24

    In this study, the microalga Nannochloropsis gaditana was subjected to acid catalysed steam explosion treatment and the resulting exploded material was subsequently fractionated to separate the different fractions (lipids, sugars and solids). Conventional and vibrational membrane setups were used with several polymeric commercial membranes. Two different routes were followed: 1) filtration+lipid solvent extraction and 2) lipid solvent extraction+filtration. Route 1 revealed to be much better since the used membrane for filtration was able to permeate the sugar aqueous phase and retained the fraction containing lipids; after this, an extraction required a much lower amount of solvent and a better recovering yield. Filtration allowed complete lipid rejection. Dynamic filtration improved permeability compared to the tangential cross-flow filtration. Best membrane performance was achieved using a 5000Da membrane with the dynamic system, obtaining a permeability of 6L/h/m(2)/bar.

  19. Isolation of a hemidesmosome-rich fraction from a human squamous cell carcinoma cell line

    SciTech Connect

    Hirako, Yoshiaki; Yonemoto, Yuki; Yamauchi, Tomoe; Nishizawa, Yuji; Kawamoto, Yoshiyuki; Owaribe, Katsushi

    2014-06-10

    Hemidesmosomes are cell-to-matrix adhesion complexes anchoring keratinocytes to basement membranes. For the first time, we present a method to prepare a fraction from human cultured cells that are highly enriched in hemidesmosomal proteins. Using DJM-1 cells derived from human squamous cell carcinoma, accumulation of hemidesmosomes was observed when these cells were cultured for more than 10 days in a commercial serum-free medium without supplemental calcium. Electron microscopy demonstrated that numerous electron-dense adhesion structures were present along the basal cell membranes of DJM-1 cells cultured under the aforementioned conditions. After removing cellular materials using an ammonia solution, hemidesmosomal proteins and deposited extracellular matrix were collected and separated by electrophoresis. There were eight major polypeptides, which were determined to be plectin, BP230, BP180, integrin α6 and β4 subunits, and laminin-332 by immunoblotting and mass spectrometry. Therefore, we designated this preparation as a hemidesmosome-rich fraction. This fraction contained laminin-332 exclusively in its unprocessed form, which may account for the promotion of laminin deposition, and minimal amounts of Lutheran blood group protein, a nonhemidesmosomal transmembrane protein. This hemidesmosome-rich fraction would be useful not only for biological research on hemidesmosomes but also for developing a serum test for patients with blistering skin diseases. - Highlights: • A defined condition promoted accumulation of hemidesmosomes in human cultured cells. • A fraction isolated from the cells contained eight major polypeptides. • The polypeptides were the five major hemidesmosome proteins and laminin-332. • The cultured cells deposited laminin-332 in its unprocessed form under the condition. • We report a method to prepare a fraction highly enriched in hemidesmosome proteins.

  20. Effect of membrane length, membrane resistance, and filtration conditions on the fractionation of milk proteins by microfiltration.

    PubMed

    Piry, A; Heino, A; Kühnl, W; Grein, T; Ripperger, S; Kulozik, U

    2012-04-01

    We investigated the fractionation of casein micelles and the whey protein β-lactoglobulin (β-LG) of skim milk by crossflow microfiltration (0.1 μm) for the first time by a novel approach as a function of membrane length and membrane resistance. A special module was constructed with 4 sections and used to assess the effects of membrane length by measuring flux and β-LG permeation (or transmission) as a function of transmembrane pressure and membrane length. Depending on the position, the membranes were partly controlled by a deposit layer. A maximum for β-LG mass flow through the various membrane sections was found, depending on the position along the membrane. To study the effect of convective flow toward the membrane, membranes with 4 different intrinsic permeation resistances were assessed in terms of the permeation and fouling effects along the flow channel. From these findings, we derived a ratio between transmembrane pressure and membrane resistance, which was useful in reducing the effect of deposit formation and, thus, to optimize the protein permeation. In addition, the fouling effect was investigated in terms of reversible and irreversible fouling and, in addition, by differentiation between pressure-induced fouling and adsorption-induced (pressure-independent) fouling, again as a function of membrane length.

  1. Investigation of hybrid membrane-sorption technologies for air fractionating

    NASA Astrophysics Data System (ADS)

    Laguntsov, N. I.; Kurchatov, I. M.; Korolev, M. V.; Tishin, A. V.

    2016-09-01

    Main aim of the work is to develop and to research single-circuit hybrid membrane- sorption system for enriching air with oxygen. The developed system allows to produce air, enriched with air up to 50%, purified from dust of any size, including nanoscale dust received in consequence of sorbent abrasion. In the course of the work the research of existing systems for air enrichment with oxygen, and the possibility of combining two methods of gas separation: membrane, and sorption, was conducted. The developed system differs from its analogues in that it has improved energy efficiency compared to methods of the membrane and sorption separation. Also work presents method of cyclogram determining of the hybrid system. In this methodic an algorithm for calculating of the cycles number, and determining of the stages duration in order to obtain the desired performance was presented.

  2. Lateral organization of membranes and cell shapes.

    PubMed Central

    Markin, V S

    1981-01-01

    The relations among membrane structure, mechanical properties, and cell shape have been investigated. The fluid mosaic membrane models used contains several components that move freely in the membrane plane. These components interact with each other and determine properties of the membrane such as curvature and elasticity. A free energy equation is postulated for such a multicomponent membrane and the condition of free energy minimum is used to obtain differential equations relating the distribution of membrane components and the local membrane curvature. The force that moves membrane components along the membrane in a variable curvature field is calculated. A change in the intramembrane interactions can bring about phase separation or particle clustering. This, in turn, may strongly affect the local curvature. The numerical solution of the set of equations for the two dimensional case allows determination of the cell shape and the component distribution along the membrane. The model has been applied to describe certain erythrocytes shape transformations. PMID:7284547

  3. Cell sorting by one gravity SPLITT fractionation.

    PubMed

    Benincasa, Maria-Anna; Moore, Lee R; Williams, P Stephen; Poptic, Earl; Carpino, Francesca; Zborowski, Maciej

    2005-08-15

    The need for innovative separative techniques suitable for the fractionation of biomaterials prompted this investigation into the performance of the gravitational split-flow thin channel (G-SPLITT) system as a cell sorter. The rigorous mathematical description of the separation mechanism allows achievement of fast separation of several million myeloma cells from healthy splenocytes using flow conditions calculated from theory. Separation in G-SPLITT is based on differences in sedimentation rate. For accurate prediction of the optimal working conditions, this parameter was directly measured by cell tracking velocimetry rather than relying on a measure of diameter (by Multisizer) and an assumed density for each cell population. We also discuss the influence of different flow conditions on the effectiveness of separation.

  4. Antioxidant activities of bambara groundnut (Vigna subterranea) protein hydrolysates and their membrane ultrafiltration fractions.

    PubMed

    Arise, Abimbola K; Alashi, Adeola M; Nwachukwu, Ifeanyi D; Ijabadeniyi, Oluwatosin A; Aluko, Rotimi E; Amonsou, Eric O

    2016-05-18

    In this study, the bambara protein isolate (BPI) was digested with three proteases (alcalase, trypsin and pepsin), to produce bambara protein hydrolysates (BPHs). These hydrolysates were passed through ultrafiltration membranes to obtain peptide fractions of different sizes (<1, 1-3, 3-5 and 5-10 kDa). The hydrolysates and their peptide fractions were investigated for antioxidant activities. The membrane fractions showed that peptides with sizes <3 kDa had significantly (p < 0.05) reduced surface hydrophobicity when compared with peptides >3 kDa. This is in agreement with the result obtained for the ferric reducing power, metal chelating and hydroxyl radical scavenging activities where higher molecular weight peptides exhibited better activity (p < 0.05) when compared to low molecular weight peptide fractions. However, for all the hydrolysates, the low molecular weight peptides were more effective diphenyl-1-picrylhydrazyl (DPPH) radical scavengers but not superoxide radicals when compared to the bigger peptides. In comparison with glutathione (GSH), BPHs and their membrane fractions had better (p < 0.05) reducing power and ability to chelate metal ions except for the pepsin hydrolysate and its membrane fractions that did not show any metal chelating activity. However, the 5-10 kDa pepsin hydrolysate peptide fractions had greater (88%) hydroxyl scavenging activity than GSH, alcalase and trypsin hydrolysates (82%). These findings show the potential use of BPHs and their peptide fraction as antioxidants in reducing food spoilage or management of oxidative stress-related metabolic disorders.

  5. Detergent fractionation with subsequent subtractive suppression hybridization as a tool for identifying genes coding for plasma membrane proteins.

    PubMed

    Lange, Andreas; Kistler, Claudia; Jutzi, Tanja B; Bazhin, Alexandr V; Klemke, Claus Detlev; Schadendorf, Dirk; Eichmüller, Stefan B

    2009-06-01

    The identification of tumor-specific proteins located at the plasma membrane is hampered by numerous methodological pitfalls many of which are associated with the post-translational modification of such proteins. Here, we present a new combination of detergent fractionation of cells and of subtractive suppression hybridization (SSH) to gain overexpressed genes coding for membrane-associated or secreted proteins. Fractionation of subcellular components by digitonin allowed sequestering mRNA of the rough Endoplasmatic reticulum and thereby increasing the percentage of sequences coding for membrane-bound proteins. Fractionated mRNAs from the cutaneous T-cell lymphoma (CTCL) cell line HuT78 and from normal peripheral blood monocytes were used for SSH leading to the enrichment of sequences overexpressed in the tumor cells. We identified some 21 overexpressed genes, among them are GPR137B, FAM62A, NOMO1, HSP90, SLIT1, IBP2, CLIF, IRAK and ARC. mRNA expression was tested for selected genes in CTCL cell lines, skin specimens and peripheral blood samples from CTCL patients and healthy donors. Several of the detected sequences are clearly related to cancer, but have not yet been associated with CTCL. qPCR confirmed an enrichment of these mRNAs in the rough endoplasmic reticulum fraction. RT-PCR confirmed the expression of these genes in skin specimens and peripheral blood of CTCL patients. Western blotting verified protein expression of HSP90 and IBP2 in HuT78. GPR137B could be detected by immunohistology in HuT78 and in keratinocytes of dysplastic epidermis, but also in sweat glands of healthy skin. In summary, we developed a new technique, which allows identifying overexpressed genes coding preferentially for membrane-associated proteins.

  6. Fuel-Cell Structure Prevents Membrane Drying

    NASA Technical Reports Server (NTRS)

    Mcelroy, J.

    1986-01-01

    Embossed plates direct flows of reactants and coolant. Membrane-type fuel-cell battery has improved reactant flow and heat removal. Compact, lightweight battery produces high current and power without drying of membranes.

  7. Actinide transport across cell membranes.

    PubMed

    Bulman, R A; Griffin, R J

    1980-01-01

    Protactinium uptake into the normal liver does not exceed 3%, but when the phospholipid levels in the liver are elevated by administration of thioacetamide this uptake increases to 31%. Phosphatidic acid, which is absent from the normal liver, has been shown to extract protactinium into organic solvents. However, phosphatidylserine, a component of normal liver cell membranes, does not extract protactinium. It might be conjectured that this is why so little protactinium is taken up by the normal liver. The hypothesis is advanced that phosphatidylserine, which is known to complex plutonium, americium and curium, may regulate the uptake of these elements by liver.

  8. Chemical degradation mechanisms of membranes for alkaline membrane fuel cells

    SciTech Connect

    Choe, Yoong-Kee; Henson, Neil J.; Kim, Yu Seung

    2015-12-31

    Chemical degradation mechanisms of membranes for alkaline membrane fuel cells have been investigated using density functional theory (DFT). We have elucidated that the aryl-ether moiety of membranes is one of the weakest site against attack of hydroxide ions. The results of DFT calculations for hydroxide initiated aryl-ether cleavage indicated that the aryl-ether cleavage occurred prior to degradation of cationic functional group. Such a weak nature of the aryl-ether group arises from the electron deficiency of the aryl group as well as the low bond dissociation energy. The DFT results suggests that removal of the aryl-ether group in the membrane should enhance the stability of membranes under alkaline conditions. In fact, an ether fee poly(phenylene) membrane exhibits excellent stability against the attack from hydroxide ions.

  9. Characterisation of cell-wall polysaccharides from mandarin segment membranes.

    PubMed

    Coll-Almela, Luis; Saura-López, Domingo; Laencina-Sánchez, José; Schols, Henk A; Voragen, Alfons G J; Ros-García, José María

    2015-05-15

    In an attempt to develop a process of enzymatic peeling of mandarin segments suitable for use on an industrial scale, the cell wall fraction of the segment membrane of Satsuma mandarin fruits was extracted to obtain a chelating agent-soluble pectin fraction (ChSS), a dilute sodium hydroxide-soluble pectin fraction (DASS), a 1M sodium hydroxide-soluble hemicellulose fraction (1MASS), a 4M sodium hydroxide-soluble hemicellulose fraction (4MASS) and a cellulose-rich residue (3.1, 0.9, 0.4, 0.7 and 1.6%w/w of fresh membrane, respectively). The ChSS pectin consisted mainly of galacturonic acid followed by arabinose and galactose. The DASS fraction contained less galacturonic acid and more neutral sugars than ChSS. Eighty-nine percent of the galacturonic acid present in the segment membranes was recovered in the above two pectin fractions. The two hemicellulosic fractions consisted of two different molecular weight populations, which also differed in their sugar composition. Arabinose, xylose, mannose, galactose and glucose were the main sugar constituents of these hemicellulose fractions. In addition to an (arabino)xylan and a xyloglucan, the presence of an arabinogalactan is suggested by the sugar composition of both hemicelluloses. The pectin fractions were also characterised by their degradability by the pectic enzymes polygalacturonase, pectinmethylesterase and rhamnogalacturonan hydrolase. However the degree of degradation of the pectin fractions by enzymes differed, and the amount of the polymeric materials resistant to further degradation and the oligomeric products also differed. Using pectic enzymes it is possible to obtain peeled mandarin segments ready to eat or for canning.

  10. [Isolation and characteristics of the plasma membrane fraction from the swine myometrium].

    PubMed

    Kondratiuk, T P; Bychenok, S F; Prishchepa, L A; Babich, L G; Kurskiĭ, M D

    1986-01-01

    An accelerated method is developed for isolating a fraction of plasma membranes of pig myometrium using ultracentrifugation within the sucrose density gradient (15% and 30%). The membranes possessed the high activity of 5'-nucleotidase and Na+, K+-ATPase and the low activity of rhotenon-insensitive NADH-cytochrome c reductase. The vesicularized preparations of plasma membranes are able of ATP-dependent accumulation of Ca2+ (7.5 +/- 0.3 nmol. 45Ca2+ per 1 mg of protein for 15 min). Phosphate increases the calcium accumulation in the presence of ATP and Mg2+. Ionophore A 23187 promotes a complete and rapid release of the previously active-accumulated calcium. The release of 45Ca2+ accumulated by the membrane fraction may be reached by introduction of 1 mM EGTA or DS-Na into the incubation medium, that evidences for the cation accumulation inside closed structures. Using concanavalin-A-sepharose 4B it is shown that 60% of membrane vesicles are turned inside out. The low saponine concentrations (0.0005%) which inhibit Ca2+-accumulation by plasma membranes but not by the endoplasmic reticulum inhibit this process by 60-70% in preparations of the isolated membrane fraction. The method has certain advantages over the previously applied methods used for isolating of plasma membrane fragments from smooth muscles.

  11. In-membrane micro fuel cell

    DOEpatents

    Omosebi, Ayokunle; Besser, Ronald

    2016-09-06

    An in-membrane micro fuel cell comprises an electrically-insulating membrane that is permissive to the flow of cations, such as protons, and a pair of electrodes deposited on channels formed in the membrane. The channels are arranged as conduits for fluids, and define a membrane ridge between the channels. The electrodes are porous and include catalysts for promoting the liberation of a proton and an electron from a chemical species and/or or the recombination of a proton and an electron with a chemical specie. The fuel cell may be provided a biosensor, an electrochemical sensor, a microfluidic device, or other microscale devices fabricated in the fuel cell membrane.

  12. Membrane tether formation from blebbing cells.

    PubMed Central

    Dai, J; Sheetz, M P

    1999-01-01

    Membrane tension has been proposed to be important in regulating cell functions such as endocytosis and cell motility. The apparent membrane tension has been calculated from tether forces measured with laser tweezers. Both membrane-cytoskeleton adhesion and membrane tension contribute to the tether force. Separation of the plasma membrane from the cytoskeleton occurs in membrane blebs, which could remove the membrane-cytoskeleton adhesion term. In renal epithelial cells, tether forces are significantly lower on blebs than on membranes that are supported by cytoskeleton. Furthermore, the tether forces are equal on apical and basolateral blebs. In contrast, tether forces from membranes supported by the cytoskeleton are greater in apical than in basolateral regions, which is consistent with the greater apparent cytoskeletal density in the apical region. We suggest that the tether force on blebs primarily contains only the membrane tension term and that the membrane tension may be uniform over the cell surface. Additional support for this hypothesis comes from observations of melanoma cells that spontaneously bleb. In melanoma cells, tether forces on blebs are proportional to the radius of the bleb, and as large blebs form, there are spikes in the tether force in other cell regions. We suggest that an internal osmotic pressure inflates the blebs, and the pressure calculated from the Law of Laplace is similar to independent measurements of intracellular pressures. When the membrane tension term is subtracted from the apparent membrane tension over the cytoskeleton, the membrane-cytoskeleton adhesion term can be estimated. In both cell systems, membrane-cytoskeleton adhesion was the major factor in generating the tether force. PMID:10585959

  13. A membrane reservoir at the cell surface

    PubMed Central

    Figard, Lauren; Sokac, Anna Marie

    2014-01-01

    Cell surface expansion is a necessary part of cell shape change. One long-standing hypothesis proposes that membrane for this expansion comes from the flattening out of cell surface projections such as microvilli and membrane folds. Correlative EM data of cells undergoing phagocytosis, cytokinesis, and morphogenesis has hinted at the existence of such an unfolding mechanism for decades; but unfolding has only recently been confirmed using live-cell imaging and biophysical approaches. Considering the wide range of cells in which plasma membrane unfolding has now been reported, it likely represents a fundamental mechanism of cell shape change. PMID:24844289

  14. Polymer electrolyte membrane assembly for fuel cells

    NASA Technical Reports Server (NTRS)

    Yen, Shiao-Ping S. (Inventor); Kindler, Andrew (Inventor); Yavrouian, Andre (Inventor); Halpert, Gerald (Inventor)

    2000-01-01

    An electrolyte membrane for use in a fuel cell can contain sulfonated polyphenylether sulfones. The membrane can contain a first sulfonated polyphenylether sulfone and a second sulfonated polyphenylether sulfone, wherein the first sulfonated polyphenylether and the second sulfonated polyphenylether sulfone have equivalent weights greater than about 560, and the first sulfonated polyphenylether and the second sulfonated polyphenylether sulfone also have different equivalent weights. Also, a membrane for use in a fuel cell can contain a sulfonated polyphenylether sulfone and an unsulfonated polyphenylether sulfone. Methods for manufacturing a membrane electrode assemblies for use in fuel cells can include roughening a membrane surface. Electrodes and methods for fabricating such electrodes for use in a chemical fuel cell can include sintering an electrode. Such membranes and electrodes can be assembled into chemical fuel cells.

  15. Polymer electrolyte membrane assembly for fuel cells

    NASA Technical Reports Server (NTRS)

    Yen, Shiao-Ping S. (Inventor); Kindler, Andrew (Inventor); Yavrouian, Andre (Inventor); Halpert, Gerald (Inventor)

    2002-01-01

    An electrolyte membrane for use in a fuel cell can contain sulfonated polyphenylether sulfones. The membrane can contain a first sulfonated polyphenylether sulfone and a second sulfonated polyphenylether sulfone, wherein the first sulfonated polyphenylether and the second sulfonated polyphenylether sulfone have equivalent weights greater than about 560, and the first sulfonated polyphenylether and the second sulfonated polyphenylether sulfone also have different equivalent weights. Also, a membrane for use in a fuel cell can contain a sulfonated polyphenylether sulfone and an unsulfonated polyphenylether sulfone. Methods for manufacturing a membrane electrode assemblies for use in fuel cells can include roughening a membrane surface. Electrodes and methods for fabricating such electrodes for use in a chemical fuel cell can include sintering an electrode. Such membranes and electrodes can be assembled into chemical fuel cells.

  16. [Germ cell membrane lipids in spermatogenesis].

    PubMed

    Wang, Ting; Shi, Xiao; Quan, Song

    2016-05-01

    Spermatogenesis is a complex developmental process in which a diploid progenitor germ cell transforms into highly specialized spermatozoa. During spermatogenesis, membrane remodeling takes place, and cell membrane permeability and liquidity undergo phase-specific changes, which are all associated with the alteration of membrane lipids. Lipids are important components of the germ cell membrane, whose volume and ratio fluctuate in different phases of spermatogenesis. Abnormal lipid metabolism can cause spermatogenic dysfunction and consequently male infertility. Germ cell membrane lipids are mainly composed of cholesterol, phospholipids and glycolipids, which play critical roles in cell adhesion and signal transduction during spermatogenesis. An insight into the correlation of membrane lipids with spermatogenesis helps us to better understand the mechanisms of spermatogenesis and provide new approaches to the diagnosis and treatment of male infertility.

  17. Fuel cell ion-exchange membrane investigation

    NASA Technical Reports Server (NTRS)

    Toy, M. S.

    1972-01-01

    The present deficiencies in the fluorocarbon sulfonic acid membrane used as the solid polymer electrolyte in the H2/O2 fuel cell are studied. Considered are: Adhesives selection, elastomeric formulations, scavenger exploration, and membrane characterization. The significant data are interpreted and recommendations are given for both short and long range further investigations in two of the four major areas: membrane adhesives and membrane stabilization.

  18. Fuel cell and membrane therefore

    SciTech Connect

    Aindow, Tai-Tsui

    2016-08-09

    A fuel cell includes first and second flow field plates, and an anode electrode and a cathode electrode between the flow field plates. A polymer electrolyte membrane (PEM) is arranged between the electrodes. At least one of the flow field plates influences, at least in part, an in-plane anisotropic physical condition of the PEM that varies in magnitude between a high value direction and a low value direction. The PEM has an in-plane physical property that varies in magnitude between a high value direction and a low value direction. The PEM is oriented with its high value direction substantially aligned with the high value direction of the flow field plate.

  19. A vacuolar-type proton pump in a vesicle fraction enriched with potassium transporting plasma membranes from tobacco hornworm midgut

    SciTech Connect

    Wieczorek, H.; Weerth, S.; Schindlbeck, M.; Klein, U.

    1989-07-05

    Mg-ATP dependent electrogenic proton transport, monitored with fluorescent acridine orange, 9-aminoacridine, and oxonol V, was investigated in a fraction enriched with potassium transporting goblet cell apical membranes of Manduca sexta larval midgut. Proton transport and the ATPase activity from the goblet cell apical membrane exhibited similar substrate specificity and inhibitor sensitivity. ATP and GTP were far better substrates than UTP, CTP, ADP, and AMP. Azide and vanadate did not inhibit proton transport, whereas 100 microM N,N'-dicyclohexylcarbodiimide and 30 microM N-ethylmaleimide were inhibitors. The pH gradient generated by ATP and limiting its hydrolysis was 2-3 pH units. Unlike the ATPase activity, proton transport was not stimulated by KCl. In the presence of 20 mM KCl, a proton gradient could not be developed or was dissipated. Monovalent cations counteracted the proton gradient in an order of efficacy like that for stimulation of the membrane-bound ATPase activity: K+ = Rb+ much greater than Li+ greater than Na+ greater than choline (chloride salts). Like proton transport, the generation of an ATP dependent and azide- and vanadate-insensitive membrane potential (vesicle interior positive) was prevented largely by 100 microM N,N'-dicyclohexylcarbodiimide and 30 microM N-ethylmaleimide. Unlike proton transport, the membrane potential was not affected by 20 mM KCl. In the presence of 150 mM choline chloride, the generation of a membrane potential was suppressed, whereas the pH gradient increased 40%, indicating an anion conductance in the vesicle membrane. Altogether, the results led to the following new hypothesis of electrogenic potassium transport in the lepidopteran midgut. A vacuolar-type electrogenic ATPase pumps protons across the apical membrane of the goblet cell, thus energizing electroneutral proton/potassium antiport. The result is a net active and electrogenic potassium flux.

  20. Kallikrein and Renin in the Membrane Fractions of the Rat Kidney.

    DTIC Science & Technology

    1980-05-23

    and Renin _____ ___ i.;? - 2 SUMMARY Plasma membrane (P-H) and endoplasmic reticulum (ER) enriched fractions were isolated from the homogenized rat...The inhibitor of proteases, p-methylsulfonylfluoride inhibited 77-80% all kallikrein preparations at 3 w*1 concentration . Activation of renin Renin ...fold although only at a concentration 5-10 times higher than used with kallikrein. The relative rate of activation of renin in the ER fraction was

  1. Proton Exchange Membranes for Fuel Cells

    SciTech Connect

    Devanathan, Ramaswami

    2010-11-01

    Proton exchange membrane, also known as polymer electrolyte membrane, fuel cells (PEMFCs) offer the promise of efficient conversion of chemical energy of fuel, such as hydrogen or methanol, into electricity with minimal pollution. Their widespread use to power zero-emission automobiles as part of a hydrogen economy can contribute to enhanced energy security and reduction in greenhouse gas emissions. However, the commercial viability of PEMFC technology is hindered by high cost associated with the membrane electrode assembly (MEA) and poor membrane durability under prolonged operation at elevated temperature. Membranes for automotive fuel cell applications need to perform well over a period comparable to the life of an automotive engine and under heavy load cycling including start-stop cycling under sub-freezing conditions. The combination of elevated temperature, changes in humidity levels, physical stresses and harsh chemical environment contribute to membrane degradation. Perfluorinated sulfonic acid (PFSA)-based membranes, such as Nafion®, have been the mainstay of PEMFC technology. Their limitations, in terms of cost and poor conductivity at low hydration, have led to continuing research into membranes that have good proton conductivity at elevated temperatures above 120 °C and under low humidity conditions. Such membranes have the potential to avoid catalyst poisoning, simplify fuel cell design and reduce the cost of fuel cells. Hydrocarbon-based membranes are being developed as alternatives to PFSA membranes, but concerns about chemical and mechanical stability and durability remain. Novel anhydrous membranes based on polymer gels infused with protic ionic liquids have also been recently proposed, but considerable fundamental research is needed to understand proton transport in novel membranes and evaluate durability under fuel cell operating conditions. In order to advance this promising technology, it is essential to rationally design the next generation

  2. Functional dynamics of cell surface membrane proteins

    NASA Astrophysics Data System (ADS)

    Nishida, Noritaka; Osawa, Masanori; Takeuchi, Koh; Imai, Shunsuke; Stampoulis, Pavlos; Kofuku, Yutaka; Ueda, Takumi; Shimada, Ichio

    2014-04-01

    Cell surface receptors are integral membrane proteins that receive external stimuli, and transmit signals across plasma membranes. In the conventional view of receptor activation, ligand binding to the extracellular side of the receptor induces conformational changes, which convert the structure of the receptor into an active conformation. However, recent NMR studies of cell surface membrane proteins have revealed that their structures are more dynamic than previously envisioned, and they fluctuate between multiple conformations in an equilibrium on various timescales. In addition, NMR analyses, along with biochemical and cell biological experiments indicated that such dynamical properties are critical for the proper functions of the receptors. In this review, we will describe several NMR studies that revealed direct linkage between the structural dynamics and the functions of the cell surface membrane proteins, such as G-protein coupled receptors (GPCRs), ion channels, membrane transporters, and cell adhesion molecules.

  3. Membrane-filtered olive mill wastewater: Quality assessment of the dried phenolic-rich fraction

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A current trend in olive mill wastewater (OMWW) management is to not only decrease environmental pollution but also extract and utilize valuable by-products. Therefore, the objectives of this study were to explore different techniques for drying a phenolic-rich membrane filtration fraction of OMWW a...

  4. Mechanical degradation of fuel cell membranes under fatigue fracture tests

    NASA Astrophysics Data System (ADS)

    Khorasany, Ramin M. H.; Sadeghi Alavijeh, Alireza; Kjeang, Erik; Wang, G. G.; Rajapakse, R. K. N. D.

    2015-01-01

    The effects of cyclic stresses on the fatigue and mechanical stability of perfluorosulfonic acid (PFSA) membranes are experimentally investigated under standard fuel cell conditions. The experiments are conducted ex-situ by subjecting membrane specimens to cyclic uniaxial tension at controlled temperature and relative humidity. The fatigue lifetime is measured in terms of the number of cycles until ultimate fracture. The results indicate that the membrane fatigue lifetime is a strong function of the applied stress, temperature, and relative humidity. The fatigue life increases exponentially with reduced stresses in all cases. The effect of temperature is found to be more significant than that of humidity, with reduced fatigue life at high temperatures. The maximum membrane strain at fracture is determined to decrease exponentially with increasing membrane lifetime. At a given fatigue life, a membrane exposed to fuel cell conditions is shown to accommodate more plastic strain before fracture than one exposed to room conditions. Overall, the proposed ex-situ membrane fatigue experiment can be utilized to benchmark the fatigue lifetime of new materials in a fraction of the time and cost associated with conventional in-situ accelerated stress testing methods.

  5. Application of membrane processes in fractionation of elements in river water.

    PubMed

    Wu, N; Wyart, Y; Rose, J; Angeletti, B; Moulin, P

    2015-01-01

    The influence of wastewater treatment plant (WWTP) effluents from one microelectronic industrial zone on element concentrations and partitioning in river water was investigated. The stepwise membrane filtration is used to distinguish different size fractions including large particulate (>18 μm), particulate (0.2-18 μm), colloidal/nanoparticle (10 kDa-0.2 μm) and truly dissolved fractions (<10 kDa) in river water samples and WWTP effluents. Results demonstrated that anthropogenic inputs (WWTP effluents and industrial area) had an important influence on concentrations and partitioning of some elements in river water. Mass balance results showed that membrane filtration processes could realize a good fractionation for many elements (good recoveries) in water samples. Flux decline during 0.2 μm and 10 kDa filtrations were analyzed, and corresponding fouling mechanisms are discussed.

  6. Red cell membrane: past, present, and future

    PubMed Central

    Gallagher, Patrick G.

    2008-01-01

    As a result of natural selection driven by severe forms of malaria, 1 in 6 humans in the world, more than 1 billion people, are affected by red cell abnormalities, making them the most common of the inherited disorders. The non-nucleated red cell is unique among human cell type in that the plasma membrane, its only structural component, accounts for all of its diverse antigenic, transport, and mechanical characteristics. Our current concept of the red cell membrane envisions it as a composite structure in which a membrane envelope composed of cholesterol and phospholipids is secured to an elastic network of skeletal proteins via transmembrane proteins. Structural and functional characterization of the many constituents of the red cell membrane, in conjunction with biophysical and physiologic studies, has led to detailed description of the way in which the remarkable mechanical properties and other important characteristics of the red cells arise, and of the manner in which they fail in disease states. Current studies in this very active and exciting field are continuing to produce new and unexpected revelations on the function of the red cell membrane and thus of the cell in health and disease, and shed new light on membrane function in other diverse cell types. PMID:18988878

  7. Red cell membrane: past, present, and future.

    PubMed

    Mohandas, Narla; Gallagher, Patrick G

    2008-11-15

    As a result of natural selection driven by severe forms of malaria, 1 in 6 humans in the world, more than 1 billion people, are affected by red cell abnormalities, making them the most common of the inherited disorders. The non-nucleated red cell is unique among human cell type in that the plasma membrane, its only structural component, accounts for all of its diverse antigenic, transport, and mechanical characteristics. Our current concept of the red cell membrane envisions it as a composite structure in which a membrane envelope composed of cholesterol and phospholipids is secured to an elastic network of skeletal proteins via transmembrane proteins. Structural and functional characterization of the many constituents of the red cell membrane, in conjunction with biophysical and physiologic studies, has led to detailed description of the way in which the remarkable mechanical properties and other important characteristics of the red cells arise, and of the manner in which they fail in disease states. Current studies in this very active and exciting field are continuing to produce new and unexpected revelations on the function of the red cell membrane and thus of the cell in health and disease, and shed new light on membrane function in other diverse cell types.

  8. Membrane Stability during Biopreservation of Blood Cells

    PubMed Central

    Stoll, Christoph; Wolkers, Willem F.

    2011-01-01

    Summary Storage methods, which can be taken into consideration for red blood cells and platelets, include liquid storage, cryopreservation and freeze-drying. Red blood cells can be hypothermically stored at refrigerated temperatures, whereas platelets are chilling sensitive and therefore cannot be stored at temperatures below 20 °C. Here we give an overview of available cryopreservation and freeze-drying procedures for blood cells and discuss the effects of these procedures on cells, particularly on cellular membranes. Cryopreservation and freeze-drying may result in chemical and structural modifications of cellular membranes. Membranes undergo phase and permeability changes during freezing and drying. Cryo- and lyoprotective agents prevent membrane damage by different mechanisms. Cryoprotective agents are preferentially excluded from membrane surfaces. They decrease the activation energy for water transport during freezing and control the rate of cellular dehydration. Lyoprotectants are thought to stabilize membranes during drying by forming direct hydrogen bonding interactions with phospholipid head groups. In addition, lyoprotectants can form a glassy state at room temperature. Recently liposomes have been investigated to stabilize blood cells during freezing and freeze-drying. Liposomes modify the composition of cellular membranes by lipid and cholesterol transfer, which can stabilize or destabilize the low temperature response of cells. PMID:21566710

  9. Proton conducting membrane for fuel cells

    DOEpatents

    Colombo, Daniel G.; Krumpelt, Michael; Myers, Deborah J.; Kopasz, John P.

    2005-12-20

    An ion conducting membrane comprising dendrimeric polymers covalently linked into a network structure. The dendrimeric polymers have acid functional terminal groups and may be covalently linked via linking compounds, cross-coupling reactions, or copolymerization reactions. The ion conducting membranes may be produced by various methods and used in fuel cells.

  10. Proton conducting membrane for fuel cells

    DOEpatents

    Colombo, Daniel G.; Krumpelt, Michael; Myers, Deborah J.; Kopasz, John P.

    2007-03-27

    An ion conducting membrane comprising dendrimeric polymers covalently linked into a network structure. The dendrimeric polymers have acid functional terminal groups and may be covalently linked via linking compounds, cross-coupling reactions, or copolymerization reactions. The ion conducting membranes may be produced by various methods and used in fuel cells.

  11. Partitioning of exogenous delta-tocopherol between the triacylglycerol and membrane lipid fractions of chicken muscle.

    PubMed

    Sigfusson, Halldor; Hultin, Herbert O

    2002-11-20

    The partitioning of exogenous delta-tocopherol, added dissolved in ethanol, between the neutral triacylglycerols and membranes of chicken leg muscles was investigated. The two lipid fractions were separated using differential ultracentrifugation techniques. Triacylglycerols were obtained after high-speed centrifugation of the minced muscle at 130000 g for 30 min. Membranes were collected from a muscle-buffer homogenate (pH 7.5) between 10000 g for 20 min and 130000 g for 30 min. The triacylglycerols collected represented from 15 to 80% of the total triacylglycerols of the minced muscle, the yields increasing with increasing muscle triacylglycerol content. The phospholipids in the isolated membrane fraction represented from 20 to 35% of the total phospholipids of the muscle. At low muscle total lipid contents (3-5%), the added delta-tocopherol was present in approximately the same concentration in both muscle lipid fractions. At higher total lipid contents, achieved by adding exogenous triacylglycerols, the delta-tocopherol concentration in the membranes increased relative to that in the triacylglycerols.

  12. Membrane elastic properties and cell function.

    PubMed

    Pontes, Bruno; Ayala, Yareni; Fonseca, Anna Carolina C; Romão, Luciana F; Amaral, Racκele F; Salgado, Leonardo T; Lima, Flavia R; Farina, Marcos; Viana, Nathan B; Moura-Neto, Vivaldo; Nussenzveig, H Moysés

    2013-01-01

    Recent studies indicate that the cell membrane, interacting with its attached cytoskeleton, is an important regulator of cell function, exerting and responding to forces. We investigate this relationship by looking for connections between cell membrane elastic properties, especially surface tension and bending modulus, and cell function. Those properties are measured by pulling tethers from the cell membrane with optical tweezers. Their values are determined for all major cell types of the central nervous system, as well as for macrophage. Astrocytes and glioblastoma cells, which are considerably more dynamic than neurons, have substantially larger surface tensions. Resting microglia, which continually scan their environment through motility and protrusions, have the highest elastic constants, with values similar to those for resting macrophage. For both microglia and macrophage, we find a sharp softening of bending modulus between their resting and activated forms, which is very advantageous for their acquisition of phagocytic functions upon activation. We also determine the elastic constants of pure cell membrane, with no attached cytoskeleton. For all cell types, the presence of F-actin within tethers, contrary to conventional wisdom, is confirmed. Our findings suggest the existence of a close connection between membrane elastic constants and cell function.

  13. Membrane Elastic Properties and Cell Function

    PubMed Central

    Pontes, Bruno; Ayala, Yareni; Fonseca, Anna Carolina C.; Romão, Luciana F.; Amaral, Racκele F.; Salgado, Leonardo T.; Lima, Flavia R.; Farina, Marcos; Viana, Nathan B.; Moura-Neto, Vivaldo; Nussenzveig, H. Moysés

    2013-01-01

    Recent studies indicate that the cell membrane, interacting with its attached cytoskeleton, is an important regulator of cell function, exerting and responding to forces. We investigate this relationship by looking for connections between cell membrane elastic properties, especially surface tension and bending modulus, and cell function. Those properties are measured by pulling tethers from the cell membrane with optical tweezers. Their values are determined for all major cell types of the central nervous system, as well as for macrophage. Astrocytes and glioblastoma cells, which are considerably more dynamic than neurons, have substantially larger surface tensions. Resting microglia, which continually scan their environment through motility and protrusions, have the highest elastic constants, with values similar to those for resting macrophage. For both microglia and macrophage, we find a sharp softening of bending modulus between their resting and activated forms, which is very advantageous for their acquisition of phagocytic functions upon activation. We also determine the elastic constants of pure cell membrane, with no attached cytoskeleton. For all cell types, the presence of F-actin within tethers, contrary to conventional wisdom, is confirmed. Our findings suggest the existence of a close connection between membrane elastic constants and cell function. PMID:23844071

  14. Directing membrane chromatography to manufacture α1-antitrypsin from human plasma fraction IV.

    PubMed

    Fan, Jinxin; Luo, Jianquan; Song, Weijie; Chen, Xiangrong; Wan, Yinhua

    2015-12-04

    The surging demand for plasma proteins, mainly driven by the growing market and the development of new therapeutic indications, is promoting manufacturers to improve the throughput of plasma proteins. Due to the inherent convective mass transfer, membrane chromatography has been proved to be an efficient approach for extracting a small amount of target proteins from large-volume feed. In this study, α1-antitrypsin (AAT) was extracted from human plasma fraction IV by a two-step membrane chromatography. An anion-exchange membrane chromatography (AEMC) was used to capture the plasma proteins in bind/elute mode, and the obtained effluent was further polished by a hydrophobic interaction membrane chromatography (HIMC) in flow-through mode. Under optimal conditions, the recovery and purity of AAT achieved 87.0% and 0.58 AAT/protein (g/g) by AEMC, respectively. After the precise polishing by HIMC, the purity of AAT was 1.22 AAT/protein (g/g). The comparison results showed that membrane chromatography outperformed column chromatography in both steps because of its high throughput. This two-step membrane chromatography could obtain an AAT recovery of 83.3% and an activity recovery of 91.4%. The outcome of this work not only offers an alternative process for protein purification from plasma, but also provides guidelines for manufacturing product from a large-volume feed with multi-components by membrane chromatography.

  15. A membrane bending model of outer hair cell electromotility.

    PubMed Central

    Raphael, R M; Popel, A S; Brownell, W E

    2000-01-01

    We propose a new mechanism for outer hair cell electromotility based on electrically induced localized changes in the curvature of the plasma membrane (flexoelectricity). Electromechanical coupling in the cell's lateral wall is modeled in terms of linear constitutive equations for a flexoelectric membrane and then extended to nonlinear coupling based on the Langevin function. The Langevin function, which describes the fraction of dipoles aligned with an applied electric field, is shown to be capable of predicting the electromotility voltage displacement function. We calculate the electrical and mechanical contributions to the force balance and show that the model is consistent with experimentally measured values for electromechanical properties. The model rationalizes several experimental observations associated with outer hair cell electromotility and provides for constant surface area of the plasma membrane. The model accounts for the isometric force generated by the cell and explains the observation that the disruption of spectrin by diamide reduces force generation in the cell. We discuss the relation of this mechanism to other proposed models of outer hair cell electromotility. Our analysis suggests that rotation of membrane dipoles and the accompanying mechanical deformation may be the molecular mechanism of electromotility. PMID:10827967

  16. Advanced membrane electrode assemblies for fuel cells

    SciTech Connect

    Kim, Yu Seung; Pivovar, Bryan S

    2014-02-25

    A method of preparing advanced membrane electrode assemblies (MEA) for use in fuel cells. A base polymer is selected for a base membrane. An electrode composition is selected to optimize properties exhibited by the membrane electrode assembly based on the selection of the base polymer. A property-tuning coating layer composition is selected based on compatibility with the base polymer and the electrode composition. A solvent is selected based on the interaction of the solvent with the base polymer and the property-tuning coating layer composition. The MEA is assembled by preparing the base membrane and then applying the property-tuning coating layer to form a composite membrane. Finally, a catalyst is applied to the composite membrane.

  17. Advanced membrane electrode assemblies for fuel cells

    SciTech Connect

    Kim, Yu Seung; Pivovar, Bryan S.

    2012-07-24

    A method of preparing advanced membrane electrode assemblies (MEA) for use in fuel cells. A base polymer is selected for a base membrane. An electrode composition is selected to optimize properties exhibited by the membrane electrode assembly based on the selection of the base polymer. A property-tuning coating layer composition is selected based on compatibility with the base polymer and the electrode composition. A solvent is selected based on the interaction of the solvent with the base polymer and the property-tuning coating layer composition. The MEA is assembled by preparing the base membrane and then applying the property-tuning coating layer to form a composite membrane. Finally, a catalyst is applied to the composite membrane.

  18. A novel bioactive membrane by cell electrospinning.

    PubMed

    Chen, Haiping; Liu, Yuanyuan; Hu, Qingxi

    2015-11-01

    Electrospinning permits fabrication of biodegradable matrices that can resemble the both scale and mechanical behavior of the native extracellular matrix. However, achieving high-cellular density and infiltration of cells within matrices with traditional technique remain challenging and time consuming. The cell electrospinning technique presented in this paper can mitigate the problems associated with these limitations. Cells encapsulated by the material in the cell electrospinning technique survived well and distributed homogenously within the nanofibrous membrane, and their vitality was improved to 133% after being cultured for 28 days. The electrospun nanofibrous membrane has a certain degradation property and favorable cell-membrane interaction that supports the active biocompatibility of the membrane. Its properties are helpful for supporting cell attachment and growth, maintaining phenotypic shape, and secreting an ample amount of extracellular matrix (ECM). This novel membrane may be a potential application within the field of tissue engineering. The ability of cell electrospinning to microintegrate cells into a biodegradable fibrous matrix embodies a novel tissue engineering approach that could be applied to fabricate a high cell density elastic tissue mimetic.

  19. Cell-cell adhesion interface: rise of the lateral membrane

    PubMed Central

    Tang, Vivian

    2017-01-01

    The lateral membrane plays an important role in the mechanical stability of epithelial cell sheet in steady state. In addition, the lateral membrane is continuously remodeled during dynamic processes such as cell extrusion, cytokinesis, and intercellular cell movement. In wound healing, the lateral membrane must be built from flat and spread cells that had crawled into the area of the wound. Thus, forming the lateral membrane is a phenomenon that occurs not only in development but also during homeostatic maintenance and regeneration of differentiated epithelial tissues. PMID:28357057

  20. A membrane reservoir at the cell surface: unfolding the plasma membrane to fuel cell shape change.

    PubMed

    Figard, Lauren; Sokac, Anna Marie

    2014-01-01

    Cell surface expansion is a necessary part of cell shape change. One long-standing hypothesis proposes that membrane for this expansion comes from the flattening out of cell surface projections such as microvilli and membrane folds. Correlative EM data of cells undergoing phagocytosis, cytokinesis, and morphogenesis has hinted at the existence of such an unfolding mechanism for decades; but unfolding has only recently been confirmed using live-cell imaging and biophysical approaches. Considering the wide range of cells in which plasma membrane unfolding has now been reported, it likely represents a fundamental mechanism of cell shape change.

  1. Durability of PEM Fuel Cell Membranes

    NASA Astrophysics Data System (ADS)

    Huang, Xinyu; Reifsnider, Ken

    Durability is still a critical limiting factor for the commercialization of polymer electrolyte membrane (PEM) fuel cells, a leading energy conversion technology for powering future hydrogen fueled automobiles, backup power systems (e.g., for base transceiver station of cellular networks), portable electronic devices, etc. Ionic conducting polymer (ionomer) electrolyte membranes are the critical enabling materials for the PEM fuel cells. They are also widely used as the central functional elements in hydrogen generation (e.g., electrolyzers), membrane cell for chlor-alkali production, etc. A perfluorosulfonic acid (PFSA) polymer with the trade name Nafion® developed by DuPont™ is the most widely used PEM in chlor-alkali cells and PEM fuel cells. Similar PFSA membranes have been developed by Dow Chemical, Asahi Glass, and lately Solvay Solexis. Frequently, such membranes serve the dual function of reactant separation and selective ionic conduction between two otherwise separate compartments. For some applications, the compromise of the "separation" function via the degradation and mechanical failure of the electrolyte membrane can be the life-limiting factor; this is particularly the case for PEM in hydrogen/oxygen fuel cells.

  2. Activated Membrane Patches Guide Chemotactic Cell Motility

    PubMed Central

    Hecht, Inbal; Skoge, Monica L.; Charest, Pascale G.; Ben-Jacob, Eshel; Firtel, Richard A.; Loomis, William F.; Levine, Herbert; Rappel, Wouter-Jan

    2011-01-01

    Many eukaryotic cells are able to crawl on surfaces and guide their motility based on environmental cues. These cues are interpreted by signaling systems which couple to cell mechanics; indeed membrane protrusions in crawling cells are often accompanied by activated membrane patches, which are localized areas of increased concentration of one or more signaling components. To determine how these patches are related to cell motion, we examine the spatial localization of RasGTP in chemotaxing Dictyostelium discoideum cells under conditions where the vertical extent of the cell was restricted. Quantitative analyses of the data reveal a high degree of spatial correlation between patches of activated Ras and membrane protrusions. Based on these findings, we formulate a model for amoeboid cell motion that consists of two coupled modules. The first module utilizes a recently developed two-component reaction diffusion model that generates transient and localized areas of elevated concentration of one of the components along the membrane. The activated patches determine the location of membrane protrusions (and overall cell motion) that are computed in the second module, which also takes into account the cortical tension and the availability of protrusion resources. We show that our model is able to produce realistic amoeboid-like motion and that our numerical results are consistent with experimentally observed pseudopod dynamics. Specifically, we show that the commonly observed splitting of pseudopods can result directly from the dynamics of the signaling patches. PMID:21738453

  3. Vesicle trafficking and cell surface membrane patchiness.

    PubMed Central

    Tang, Q; Edidin, M

    2001-01-01

    Membrane proteins and lipids often appear to be distributed in patches on the cell surface. These patches are often assumed to be membrane domains, arising from specific molecular associations. However, a computer simulation (Gheber and Edidin, 1999) shows that membrane patchiness may result from a combination of vesicle trafficking and dynamic barriers to lateral mobility. The simulation predicts that the steady-state patches of proteins and lipids seen on the cell surface will decay if vesicle trafficking is inhibited. To test this prediction, we compared the apparent sizes and intensities of patches of class I HLA molecules, integral membrane proteins, before and after inhibiting endocytic vesicle traffic from the cell surface, either by incubation in hypertonic medium or by expression of a dominant-negative mutant dynamin. As predicted by the simulation, the apparent sizes of HLA patches increased, whereas their intensities decreased after endocytosis and vesicle trafficking were inhibited. PMID:11423406

  4. Characterization of a Partially Purified Adenosine Triphosphatase from a Corn Root Plasma Membrane Fraction 1

    PubMed Central

    Dupont, Frances M.; Burke, Linda L.; Spanswick, Roger M.

    1981-01-01

    The (K+,Mg2+)-ATPase was partially purified from a plasma membrane fraction from corn roots (WF9 × Mol7) and stored in liquid N2 without loss of activity. Specific activity was increased 4-fold over that of the plasma membrane fraction. ATPase activity resembled that of the plasma membrane fraction with certain alterations in cation sensitivity. The enzyme required a divalent cation for activity (Co2+ > Mg2+ > Mn2+ > Zn2+ > Ca2+) when assayed at 3 millimolar ATP and 3 millimolar divalent cation at pH 6.3. When assayed in the presence of 3 millimolar Mg2+, the enzyme was further activated by monovalent cations (K+, NH4+, Rb+ ≫ Na+, Cs+, Li+). The pH optima were 6.5 and 6.3 in the absence and presence of 50 millimolar KCl, respectively. The enzyme showed simple Michaelis-Menten kinetics for the substrate ATP-Mg, with a Km of 1.3 millimolar in the absence and 0.7 millimolar in the presence of 50 millimolar KCl. Stimulation by K+ approached simple Michaelis-Menten kinetics, with a Km of approximately 4 millimolar KCl. ATPase activity was inhibited by sodium orthovanadate. Half-maximal inhibition was at 150 and 35 micromolar in the absence and presence of 50 millimolar KCl. The enzyme required the substrate ATP. The rate of hydrolysis of other substrates, except UDP, IDP, and GDP, was less than 20% of ATP hydrolysis. Nucleoside diphosphatase activity was less than 30% of ATPase activity, was not inhibited by vanadate, was not stimulated by K+, and preferred Mn2+ to Mg2+. The results demonstrate that the (K+,Mg2+)-ATPase can be clearly distinguished from nonspecific phosphohydrolase and nucleoside diphosphatase activities of plasma membrane fractions prepared from corn roots. PMID:16661634

  5. Microalgal biomass production by using ultra- and nanofiltration membrane fractions of olive mill wastewater.

    PubMed

    Cicci, A; Stoller, M; Bravi, M

    2013-09-01

    Olive milling produces huge amounts of wastewater (OMWW) characterized by an extremely high organic load. Its polyphenols content is a hindrance to conventional biological treatment and to using it as growing medium for common microbial biomasses. The practice to dump it on soil is in conflict with the latest EU directives about waste management. OMWW can be effectively and efficiently treated by means of membrane technology to a fraction of the initial volume, but membrane processing concentrates still require treatment. Reversing the overall cost balance of membrane processing and subsequent treatment requires valorizing the concentrates through their reuse, as well as ensuring long-term service of the membrane system through effective wastewater pretreatment and sustainable, fouling-controlling, membrane operation conduite. Aim of this work is to reuse and valorize the ultra- and nanofiltration membrane concentrates as media for biomass production of microalgae and cyanobacteria. Scenedesmus dimorphus and Arthrospira platensis, usable as a food, feed, nutraceutical component or feedstock for biofuels, were selected for this investigation. Microalgal growth was experimentally determined and related to the composition of the concentrate-based media and to the irradiance distribution within the photobioreactor volume to decouple light limitation and medium chemical composition effects.

  6. Stretching micropatterned cells on a PDMS membrane.

    PubMed

    Carpi, Nicolas; Piel, Matthieu

    2014-01-22

    Mechanical forces exerted on cells and/or tissues play a major role in numerous processes. We have developed a device to stretch cells plated on a PolyDiMethylSiloxane (PDMS) membrane, compatible with imaging. This technique is reproducible and versatile. The PDMS membrane can be micropatterned in order to confine cells or tissues to a specific geometry. The first step is to print micropatterns onto the PDMS membrane with a deep UV technique. The PDMS membrane is then mounted on a mechanical stretcher. A chamber is bound on top of the membrane with biocompatible grease to allow gliding during the stretch. The cells are seeded and allowed to spread for several hours on the micropatterns. The sample can be stretched and unstretched multiple times with the use of a micrometric screw. It takes less than a minute to apply the stretch to its full extent (around 30%). The technique presented here does not include a motorized device, which is necessary for applying repeated stretch cycles quickly and/or computer controlled stretching, but this can be implemented. Stretching of cells or tissue can be of interest for questions related to cell forces, cell response to mechanical stress or tissue morphogenesis. This video presentation will show how to avoid typical problems that might arise when doing this type of seemingly simple experiment.

  7. Activation of intrinsic apoptotic signaling pathway in cancer cells by Cymbopogon citratus polysaccharide fractions.

    PubMed

    Thangam, Ramar; Sathuvan, Malairaj; Poongodi, Arasu; Suresh, Veeraperumal; Pazhanichamy, Kalailingam; Sivasubramanian, Srinivasan; Kanipandian, Nagarajan; Ganesan, Nalini; Rengasamy, Ramasamy; Thirumurugan, Ramasamy; Kannan, Soundarapandian

    2014-07-17

    Essential oils of Cymbopogon citratus were already reported to have wide ranging medical and industrial applications. However, information on polysaccharides from the plant and their anticancer activities are limited. In the present study, polysaccharides from C. citratus were extracted and fractionated by anion exchange and gel filtration chromatography. Two different polysaccharide fractions such as F1 and F2 were obtained, and these fractions were found to have distinct acidic polysaccharides as characterized by their molecular weight and sugar content. NMR spectral analysis revealed the presence of (1→4) linked b-d-Xylofuranose moiety in these polysaccharides. Using these polysaccharide fractions F1 and F2, anti-inflammatory and anticancer activities were evaluated against cancer cells in vitro and the mechanism of action of the polysaccharides in inducing apoptosis in cancer cells via intrinsic pathway was also proposed. Two different reproductive cancer cells such as Siha and LNCap were employed for in vitro studies on cytotoxicity, induction of apoptosis and apoptotic DNA fragmentation, changes in mitochondrial membrane potential, and profiles of gene and protein expression in response to treatment of cells by the polysaccharide fractions. These polysaccharide fractions exhibited potential cytotoxic and apoptotic effects on carcinoma cells, and they induced apoptosis in these cells through the events of up-regulation of caspase 3, down-regulation of bcl-2 family genes followed by cytochrome c release.

  8. Cellular enrichment through microfluidic fractionation based on cell biomechanical properties.

    PubMed

    Wang, Gonghao; Turbyfield, Cory; Crawford, Kaci; Alexeev, Alexander; Sulchek, Todd

    2015-10-01

    The biomechanical properties of populations of diseased cells are shown to have differences from healthy populations of cells, yet the overlap of these biomechanical properties can limit their use in disease cell enrichment and detection. We report a new microfluidic cell enrichment technology that continuously fractionates cells through differences in biomechanical properties, resulting in highly pure cellular subpopulations. Cell fractionation is achieved in a microfluidic channel with an array of diagonal ridges that are designed to segregate biomechanically distinct cells to different locations in the channel. Due to the imposition of elastic and viscous forces during cellular compression, which are a function of cell biomechanical properties including size and viscoelasticity, larger, stiffer and less viscos cells migrate parallel to the diagonal ridges and exhibit positive lateral displacement. On the other hand, smaller, softer and more viscous cells migrate perpendicular to the diagonal ridges due to circulatory flow induced by the ridges and result in negative lateral displacement. Multiple outlets are then utilized to collect cells with finer gradation of differences in cell biomechanical properties. The result is that cell fractionation dramatically improves cell separation efficiency compared to binary outputs and enables the measurement of subtle biomechanical differences within a single cell type. As a proof-of-concept demonstration, we mix two different leukemia cell lines (K562 and HL60) and utilize cell fractionation to achieve over 45-fold enhancement of cell populations, with high purity cellular enrichment (90% to 99%) of each cell line. In addition, we demonstrate cell fractionation of a single cell type (K562 cells) into subpopulations and characterize the variations of biomechanical properties of the separated cells with atomic force microscopy. These results will be beneficial to obtaining label-free separation of cellular mixtures, or to

  9. Cellular enrichment through microfluidic fractionation based on cell biomechanical properties

    PubMed Central

    Wang, Gonghao; Turbyfield, Cory; Crawford, Kaci; Alexeev, Alexander; Sulchek, Todd

    2017-01-01

    The biomechanical properties of populations of diseased cells are shown to have differences from healthy populations of cells, yet the overlap of these biomechanical properties can limit their use in disease cell enrichment and detection. We report a new microfluidic cell enrichment technology that continuously fractionates cells through differences in biomechanical properties, resulting in highly pure cellular subpopulations. Cell fractionation is achieved in a microfluidic channel with an array of diagonal ridges that are designed to segregate biomechanically distinct cells to different locations in the channel. Due to the imposition of elastic and viscous forces during cellular compression, which are a function of cell biomechanical properties including size and viscoelasticity, larger, stiffer and less viscos cells migrate parallel to the diagonal ridges and exhibit positive lateral displacement. On the other hand, smaller, softer and more viscous cells migrate perpendicular to the diagonal ridges due to circulatory flow induced by the ridges and result in negative lateral displacement. Multiple outlets are then utilized to collect cells with finer gradation of differences in cell biomechanical properties. The result is that cell fractionation dramatically improves cell separation efficiency compared to binary outputs and enables the measurement of subtle biomechanical differences within a single cell type. As a proof-of-concept demonstration, we mix two different leukemia cell lines (K562 and HL60) and utilize cell fractionation to achieve over 45-fold enhancement of cell populations, with high purity cellular enrichment (90% to 99%) of each cell line. In addition, we demonstrate cell fractionation of a single cell type (K562 cells) into subpopulations and characterize the variations of biomechanical properties of the separated cells with atomic force microscopy. These results will be beneficial to obtaining label-free separation of cellular mixtures, or to

  10. Solubilization and Partial Purification of the Adenosine Triphosphatase from a Corn Root Plasma Membrane Fraction

    PubMed Central

    Dupont, Frances M.; Leonard, Robert T.

    1980-01-01

    The K+-stimulated ATPase was partially purified from a plasma membrane fraction from corn roots (WF9 × Mo 17) by solubilization with 30 millimolar octyl-β-d-glucopyranoside followed by precipitation with dilute ammonium sulfate. The specific activity of the enzyme was increased about five times by this procedure. The molecular weight of the detergent-extracted ATPase complex was estimated to be at least 500,000 daltons by chromatography on a Bio-Gel A-5m column. Negative staining electron microscopy indicated that the detergent-extracted material consisted of amorphous particles, while the ammonium sulfate precipitate was composed of uniform vesicles with an average diameter of 100 nanometers. The protein composition of the ammonium sulfate precipitate was significantly different from that of the plasma membrane fraction when compared by sodium dodecyl sulfate gel electrophoresis. The characteristics of the partially purified ATPase resembled those of the plasma membrane associated enzyme. The ATPase required Mg2+, was further stimulated by K+, was almost completely inhibited by 0.1 millimolar diethylstilbestrol, and was not affected by 5.0 micrograms per milliliter oligomycin. Although the detergents sodium cholate, deoxycholate, Triton X-100 and Lubrol WX also solubilized some membrane protein, none solubilized the K+-stimulated ATPase activity. Low concentrations of each detergent, including octyl-β-d-glucopyranoside, activated the ATPase and higher concentrations inactivated the enzyme. These results suggest that the plasma membrane ATPase is a large, integral membrane protein or protein complex that requires lipids to maintain its activity. Images PMID:16661309

  11. Fuel cell subassemblies incorporating subgasketed thrifted membranes

    DOEpatents

    Iverson, Eric J.; Pierpont, Daniel M.; Yandrasits, Michael A.; Hamrock, Steven J.; Obradovich, Stephan J.; Peterson, Donald G.

    2016-03-01

    A fuel cell roll good subassembly is described that includes a plurality of individual electrolyte membranes. One or more first subgaskets are attached to the individual electrolyte membranes. Each of the first subgaskets has at least one aperture and the first subgaskets are arranged so the center regions of the individual electrolyte membranes are exposed through the apertures of the first subgaskets. A second subgasket comprises a web having a plurality of apertures. The second subgasket web is attached to the one or more first subgaskets so the center regions of the individual electrolyte membranes are exposed through the apertures of the second subgasket web. The second subgasket web may have little or no adhesive on the subgasket surface facing the electrolyte membrane.

  12. Fuel cell subassemblies incorporating subgasketed thrifted membranes

    DOEpatents

    Iverson, Eric J; Pierpont, Daniel M; Yandrasits, Michael A; Hamrock, Steven J; Obradovich, Stephan J; Peterson, Donald G

    2014-01-28

    A fuel cell roll good subassembly is described that includes a plurality of individual electrolyte membranes. One or more first subgaskets are attached to the individual electrolyte membranes. Each of the first subgaskets has at least one aperture and the first subgaskets are arranged so the center regions of the individual electrolyte membranes are exposed through the apertures of the first subgaskets. A second subgasket comprises a web having a plurality of apertures. The second subgasket web is attached to the one or more first subgaskets so the center regions of the individual electrolyte membranes are exposed through the apertures of the second subgasket web. The second subgasket web may have little or no adhesive on the subgasket surface facing the electrolyte membrane.

  13. Mechanical tension drives cell membrane fusion.

    PubMed

    Kim, Ji Hoon; Ren, Yixin; Ng, Win Pin; Li, Shuo; Son, Sungmin; Kee, Yee-Seir; Zhang, Shiliang; Zhang, Guofeng; Fletcher, Daniel A; Robinson, Douglas N; Chen, Elizabeth H

    2015-03-09

    Membrane fusion is an energy-consuming process that requires tight juxtaposition of two lipid bilayers. Little is known about how cells overcome energy barriers to bring their membranes together for fusion. Previously, we have shown that cell-cell fusion is an asymmetric process in which an "attacking" cell drills finger-like protrusions into the "receiving" cell to promote cell fusion. Here, we show that the receiving cell mounts a Myosin II (MyoII)-mediated mechanosensory response to its invasive fusion partner. MyoII acts as a mechanosensor, which directs its force-induced recruitment to the fusion site, and the mechanosensory response of MyoII is amplified by chemical signaling initiated by cell adhesion molecules. The accumulated MyoII, in turn, increases cortical tension and promotes fusion pore formation. We propose that the protrusive and resisting forces from fusion partners put the fusogenic synapse under high mechanical tension, which helps to overcome energy barriers for membrane apposition and drives cell membrane fusion.

  14. Analysis of plasma membrane phosphoinositides from fusogenic carrot cells

    SciTech Connect

    Wheeler, J.J.; Boss, W.F.

    1987-04-01

    Phosphatidylinositol monophosphate (PIP) and phosphatidylinositol bisphosphate (PIP/sub 2/) were found to be associated with the plasma membrane-rich fractions isolated by aqueous polymer two-phase partitioning from fusogenic cells. They represented at least 5% and 0.7% of the total inositol-labeled lipids in the plasma membrane-rich fractions, respectively, and were present in a ratio of about 7:1 (PIP:PIP/sub 2/). In addition, two unidentified inositol-labeled compounds, which together were approximately 3% of the inositol-labeled lipids, were found predominantly in the plasma membrane-rich fractions and migrated between PIP/sub 2/ and PIP. The R/sub f/s of these compounds were approximately 0.31 and 0.34 in the solvent system CHCl/sub 3/:MeOH:15N NH/sub 4/OH:H/sub 2/O (90:90:7:22) using LK5 plates presoaked in 1% potassium oxalate. These compounds incorporated /sup 32/P/sub i/, (/sup 3/H)inositol and were hydrolyzed in mild base. These data suggested that they were glycero-phospholipids. Although the compounds did not comigrate with lysoPIP obtained from bovine brain (R/sub f/ approx. 0.35), when endogenous PIP was hydrolyzed to lysoPIP, the breakdown product migrated in the region of the unidentified inositol lipids.

  15. Cell membrane softening in human breast and cervical cancer cells

    NASA Astrophysics Data System (ADS)

    Händel, Chris; Schmidt, B. U. Sebastian; Schiller, Jürgen; Dietrich, Undine; Möhn, Till; Kießling, Tobias R.; Pawlizak, Steve; Fritsch, Anatol W.; Horn, Lars-Christian; Briest, Susanne; Höckel, Michael; Zink, Mareike; Käs, Josef A.

    2015-08-01

    Biomechanical properties are key to many cellular functions such as cell division and cell motility and thus are crucial in the development and understanding of several diseases, for instance cancer. The mechanics of the cellular cytoskeleton have been extensively characterized in cells and artificial systems. The rigidity of the plasma membrane, with the exception of red blood cells, is unknown and membrane rigidity measurements only exist for vesicles composed of a few synthetic lipids. In this study, thermal fluctuations of giant plasma membrane vesicles (GPMVs) directly derived from the plasma membranes of primary breast and cervical cells, as well as breast cell lines, are analyzed. Cell blebs or GPMVs were studied via thermal membrane fluctuations and mass spectrometry. It will be shown that cancer cell membranes are significantly softer than their non-malignant counterparts. This can be attributed to a loss of fluid raft forming lipids in malignant cells. These results indicate that the reduction of membrane rigidity promotes aggressive blebbing motion in invasive cancer cells.

  16. Basement Membranes: Cell Scaffoldings and Signaling Platforms

    PubMed Central

    Yurchenco, Peter D.

    2011-01-01

    Basement membranes are widely distributed extracellular matrices that coat the basal aspect of epithelial and endothelial cells and surround muscle, fat, and Schwann cells. These extracellular matrices, first expressed in early embryogenesis, are self-assembled on competent cell surfaces through binding interactions among laminins, type IV collagens, nidogens, and proteoglycans. They form stabilizing extensions of the plasma membrane that provide cell adhesion and that act as solid-phase agonists. Basement membranes play a role in tissue and organ morphogenesis and help maintain function in the adult. Mutations adversely affecting expression of the different structural components are associated with developmental arrest at different stages as well as postnatal diseases of muscle, nerve, brain, eye, skin, vasculature, and kidney. PMID:21421915

  17. Molecular motors are differentially distributed on Golgi membranes from polarized epithelial cells

    PubMed Central

    1994-01-01

    Microtubules (MT) are required for the efficient transport of membranes from the trans-Golgi and for transcytosis of vesicles from the basolateral membrane to the apical cytoplasm in polarized epithelia. MTs in these cells are primarily oriented with their plus ends basally near the Golgi and their minus-ends in the apical cytoplasm. Here we report that isolated Golgi and Golgi-enriched membranes from intestinal epithelial cells possess the actin based motor myosin-I, the MT minus- end-directed motor cytoplasmic dynein and its in vitro motility activator dynactin (p150/Glued). The Golgi can be separated into stacks, possessing features of the Golgi cisternae, and small membranes enriched in the trans-Golgi network marker TGN 38/41. Whereas myosin-I is present on all membranes in the Golgi fraction, dynein is present only on the small membrane fraction. Dynein, like myosin-I, is associated with membranes as a cytoplasmic peripheral membrane protein. Dynein and myosin-I coassociate with membranes that bind to MTs and cross-link actin filaments and MTs in a nucleotide-dependent manner. We propose that cytoplasmic dynein moves Golgi membranes along MTs to the cell cortex where myosin-I provides local delivery through the actin- rich cytoskeleton to the apical membrane. PMID:8045931

  18. Heterogeneous distribution of enzymes among plasma-membrane fragments sedimenting with the microsomal fraction of rat liver

    PubMed Central

    Norris, Kenneth A.; Dobrota, Miloslav; Issa, Faiz S.; Hinton, Richard H.; Reid, Eric

    1974-01-01

    Plasma-membrane fragments recovered in the microsomal fraction of rat liver homogenates were shown to be heterogeneous in density. It was demonstrated that 5′-nucleotidase, the most commonly used plasma-membrane marker, is concentrated in the lightest subfraction. Two of the published procedures for the isolation of plasma-membrane fragments from the microsomal fraction (Touster et al., 1970; Hinton et al., 1971) are shown to give products which are not representative of all the plasma-membrane fragments of microsomal size, and it is argued that a third procedure (House & Weidemann, 1970) is likely to give a similar product. PMID:4377214

  19. Continuous flow magnetic cell fractionation based on antigen expression level.

    PubMed

    Schneider, Thomas; Moore, Lee R; Jing, Ying; Haam, Seungjoo; Williams, P Stephen; Fleischman, Aaron J; Roy, Shuvo; Chalmers, Jeffrey J; Zborowski, Maciej

    2006-07-31

    Cell separation is important in medical and biological research and plays an increasingly important role in clinical therapy and diagnostics, such as rare cancer cell detection in blood. The immunomagnetic labeling of cells with antibodies conjugated to magnetic nanospheres gives rise to a proportional relationship between the number of magnetic nanospheres attached to the cell and the cell surface marker number. This enables the potential fractionation of cell populations by magnetophoretic mobility (MM). We exploit this feature with our apparatus, the Dipole Magnet Flow Fractionator (DMFF), which consists of an isodynamic magnetic field, an orthogonally-oriented thin ribbon of cell suspension in continuous sheath flow, and ten outlet flows. From a sample containing a 1:1 mixture of immunomagnetically labeled (label+) and unlabeled (label-) cells, we achieved an increase in enrichment of the label+ cell fraction with increasing outlet numbers in the direction of the magnetic field gradient (up to 10-fold). The total recovery of the ten outlet fractions was 90.0+/-7.7%. The mean MM of label+ cells increased with increasing outlet number by up to a factor of 2.3. The postulated proportionality between the number of attached magnetic beads and the number of cell surface markers was validated by comparison of MM measured by cell tracking velocimetry (CTV) with cell florescence intensity measured by flow cytometry.

  20. Alternate Fuel Cell Membranes for Energy Independence

    SciTech Connect

    Storey, Robson, F.; Mauritz, Kenneth, A.; Patton, Derek, L.; Savin, Daniel, A.

    2012-12-18

    The overall objective of this project was the development and evaluation of novel hydrocarbon fuel cell (FC) membranes that possess high temperature performance and long term chemical/mechanical durability in proton exchange membrane (PEM) fuel cells (FC). The major research theme was synthesis of aromatic hydrocarbon polymers of the poly(arylene ether sulfone) (PAES) type containing sulfonic acid groups tethered to the backbone via perfluorinated alkylene linkages and in some cases also directly attached to the phenylene groups along the backbone. Other research themes were the use of nitrogen-based heterocyclics instead of acid groups for proton conduction, which provides high temperature, low relative humidity membranes with high mechanical/thermal/chemical stability and pendant moieties that exhibit high proton conductivities in the absence of water, and synthesis of block copolymers consisting of a proton conducting block coupled to poly(perfluorinated propylene oxide) (PFPO) blocks. Accomplishments of the project were as follows: 1) establishment of a vertically integrated program of synthesis, characterization, and evaluation of FC membranes, 2) establishment of benchmark membrane performance data based on Nafion for comparison to experimental membrane performance, 3) development of a new perfluoroalkyl sulfonate monomer, N,N-diisopropylethylammonium 2,2-bis(p-hydroxyphenyl) pentafluoropropanesulfonate (HPPS), 4) synthesis of random and block copolymer membranes from HPPS, 5) synthesis of block copolymer membranes containing high-acid-concentration hydrophilic blocks consisting of HPPS and 3,3'-disulfonate-4,4'-dichlorodiphenylsulfone (sDCDPS), 6) development of synthetic routes to aromatic polymer backbones containing pendent 1H-1,2,3-triazole moieties, 7) development of coupling strategies to create phase-separated block copolymers between hydrophilic sulfonated prepolymers and commodity polymers such as PFPO, 8) establishment of basic performance

  1. Modification and evaluation of fuel cell membranes

    NASA Astrophysics Data System (ADS)

    Nalawade, Amol Prataprao

    The primary goals of this study were modification of existing NafionRTM membranes and characterization of newly developed hydrocarbon-based membranes for high temperature fuel cell applications. Various NafionRTM/silicate nanocomposites were formulated via in situ sol-gel reactions for tetraethylorthosilicate. Different silicate composition profiles generated across membrane cross-sections were investigated by EDAX/ESEM. Composite water uptake, proton conductivity and fuel cell performance were comparable to that of unmodified Nafion RTM. Tafel analysis showed better electrode kinetics for composites having more silicate in the middle and less or no silicate at electrolyte-electrode interfaces. All composites showed reduced fuel cross-over and superior mechanical as well as chemical durability than unmodified NafionRTM. Poly(cyclohexadiene) (PCHD) materials were characterized in the interest of developing alternative low-cost proton exchange membranes. All cross-linked sulfonated (xsPCHD) membranes showed significantly higher water uptake at 80 °C and higher proton conductivity at 120 °C at all relative humidities (RH), compared to the current benchmark membrane, NafionRTM. A xsPCHD-poly(ethylene glycol) (PEG) copolymer and a xsPCHD-PEG blend surpassed the DOE target by exhibiting proton conductivities of 141.44 and 322.40 mS/cm, respectively, at 50 % RH. Although the PCHD-based PEMs exhibited thermal stability up to 150 °C, they showed poor mechanical properties which would cause poor membrane durability during fuel cell operation. Atomic force microscopy studies demonstrated nanophase separated morphology of xsPCHD having a higher degree of connectedness of hydrophilic domains in the copolymer and blends relative to the xsPCHD homopolymer. Broadband dielectric spectroscopy (BDS) was used to study sub-Tg relaxations in annealed poly(2,5-benzimidazole) (ABPBI) fuel cell precursor materials. A trend in degree of connectivity of charge migration pathways and

  2. Solubilization of pig lymphocyte plasma membrane and fractionation of some of the components

    PubMed Central

    Allan, D.; Crumpton, M. J.

    1971-01-01

    The degree of solubilization of pig lymphocyte plasma membrane by sodium deoxycholate was determined at a variety of temperatures and detergent concentrations. Approx. 95% of the membrane protein was soluble in 2% deoxycholate at 23°C. Some of the biological activities of the membrane survived this treatment. The leucine β-naphthylamidase activity was more readily soluble than the 5′-nucleotidase and these enzymes could be separated by extraction with 0.5% deoxycholate at 0°C. Membrane solubilized in 2% deoxycholate at 23°C was fractionated by sucrose-density-gradient centrifugation in 1% deoxycholate. The phospholipid was separated from the protein, which formed a fairly symmetrical peak that sedimented slightly slower than ovalbumin; the leucine naphthylamidase and 5′-nucleotidase activities were resolved from each other and from the main protein peak. Similar separations were achieved by elution from Sephadex G-200 and Sepharose 6B in 1% deoxycholate. The main proteins, however, appeared to possess much higher molecular weights than those indicated by sucrose-density-gradient centrifugation. This disparity suggests that many of the membrane proteins have a rod-like shape, especially since the results of experiments with [14C]deoxycholate revealed that the proteins did not bind significant amounts of deoxycholate. In contrast, 5′-nucleotidase and leucine naphthylamidase appeared to be globular. Polyacrylamide-gel electrophoresis of membrane solubilized in sodium dodecyl sulphate gave a similar distribution of protein to that achieved by sucrose-density-gradient centrifugation. Trace amounts only of polypeptides of molecular weight less than 10000 were detected. ImagesPLATE 1 PMID:4256533

  3. Membrane filtration of olive mill wastewater and exploitation of its fractions.

    PubMed

    Paraskeva, C A; Papadakis, V G; Kanellopoulou, D G; Koutsoukos, P G; Angelopoulos, K C

    2007-04-01

    Olive mill wastewater (OMW) produced from small units scattered in rural areas of Southern Europe is a major source of pollution of surface and subsurface water. In the present work, a treatment scheme based on physical separation methods is presented. The investigation was carried out using a pilot-plant unit equipped with ultrafiltration, nanofiltration, and reverse osmosis membranes. Approximately 80% of the total volume of wastewater treated by the membrane units was sufficiently cleaned to meet the standards for irrigation water. The concentrated fractions collected in the treatment concentrates were characterized by high organic load and high content of phenolic compounds. The concentrates were tested in hydroponic systems to examine their toxicity towards undesired herbs. The calculations of the cost of the overall process showed that fixed and operational costs could be recovered from the exploitation of OMW byproducts as water for irrigation and/or as bioherbicides.

  4. Hereditary spherocytosis, elliptocytosis, and other red cell membrane disorders.

    PubMed

    Da Costa, Lydie; Galimand, Julie; Fenneteau, Odile; Mohandas, Narla

    2013-07-01

    Hereditary spherocytosis and elliptocytosis are the two most common inherited red cell membrane disorders resulting from mutations in genes encoding various red cell membrane and skeletal proteins. Red cell membrane, a composite structure composed of lipid bilayer linked to spectrin-based membrane skeleton is responsible for the unique features of flexibility and mechanical stability of the cell. Defects in various proteins involved in linking the lipid bilayer to membrane skeleton result in loss in membrane cohesion leading to surface area loss and hereditary spherocytosis while defects in proteins involved in lateral interactions of the spectrin-based skeleton lead to decreased mechanical stability, membrane fragmentation and hereditary elliptocytosis. The disease severity is primarily dependent on the extent of membrane surface area loss. Both these diseases can be readily diagnosed by various laboratory approaches that include red blood cell cytology, flow cytometry, ektacytometry, electrophoresis of the red cell membrane proteins, and mutational analysis of gene encoding red cell membrane proteins.

  5. Membrane fluidity sensoring microbial fuel cell.

    PubMed

    Choi, Youngjin; Jung, Eunkyoung; Kim, Sunghyun; Jung, Seunho

    2003-04-01

    A study has been performed to examine the effect of temperature and ethanolic stresses on the coulombic efficiency of a microbial fuel cell. The conventional-type fuel cell containing Gram-negative bacteria, Proteus vulgaris, was investigated as a model system. From current output measurements, it was found that the coulombic yields were altered by environmental stresses such as temperature shock or ethanol treatment to the bacteria. While high-temperature or ethanolic shock led to a remarkable decrement in coulombic output, the low-temperature shock induced a slight increase in microbial fuel cell efficiency. These results indicate that the membrane fluidity is affected considerably by environmental stress, which in turn affects the electron transfer process through the bacterial cell membrane to and from the electrode. This interpretation was confirmed by the cyclic voltammetric study of a mediator on an electrode surface modified with the lipids extracted from the membrane of P. vulgaris under the given stress. Markedly different electrochemical behaviors were observed depending on the environmental stress. A reciprocal relationship between coulomb output and the ratio of saturation/unsaturation of fatty acids has been observed. This is the first report, to our knowledge, that the structural adaptation of membrane fatty acids in response to the environmental shock can regulate the coulombic efficiency of a microbial fuel cell.

  6. Pattern formation in cell membrane adhesion

    NASA Astrophysics Data System (ADS)

    Discher, Dennis; Hategan, A.; Sengupta, K.; Sackmann, E.

    2004-03-01

    Strong adhesion of highly active cells often nucleates focal adhesions or related structures that are, over time, reinforced by cytoskeleton (actin, etc.). Red cells lack such complex adhesion systems, but they are shown here to also exhibit complex spatial patterns within an adhesive contact zone. While strong adhesion and spreading of the red cell to a dense poly-L-lysine surface appears complete in < 1 s by reflective interference microscopy, over longer times of 10-15 min or more distinct patterns in fluorescently labeled membrane components emerge. The fluorescent lipid Fl-PE (fluorescein phosphoethanolamine), in particular, is seen to diffuse and reorganize (eg. worm-like domains of <500 nm) within the contact zone, independent of whether the cell is intact or ruptured. Lipid patterns are accompanied by visible perturbations in band 3 distribution and weaker perturbations in membrane skeleton actin. Although fluorescent poly-L-lysine is shown to be uniform under cells, pressing down on the membrane quenches the lipid patterns and reveals the topographical basis for pattern formation. Regions of strong contact are thus separated by regions where the membrane is more distant from the surface.

  7. Cell or Cell Membrane-Based Drug Delivery Systems

    PubMed Central

    Tan, Songwei; Wu, Tingting; Zhang, Dan; Zhang, Zhiping

    2015-01-01

    Natural cells have been explored as drug carriers for a long period. They have received growing interest as a promising drug delivery system (DDS) until recently along with the development of biology and medical science. The synthetic materials, either organic or inorganic, are found to be with more or less immunogenicity and/or toxicity. The cells and extracellular vesicles (EVs), are endogenous and thought to be much safer and friendlier. Furthermore, in view of their host attributes, they may achieve different biological effects and/or targeting specificity, which can meet the needs of personalized medicine as the next generation of DDS. In this review, we summarized the recent progress in cell or cell membrane-based DDS and their fabrication processes, unique properties and applications, including the whole cells, EVs and cell membrane coated nanoparticles. We expect the continuing development of this cell or cell membrane-based DDS will promote their clinic applications. PMID:26000058

  8. Osteogenic and Adipogenic Cell Fractions Isolated from Postnatal Mouse Calvaria

    PubMed Central

    Steenhuis, P.; Carr, K.M.; Pettway, G.J.; Ignelzi, M.A.

    2009-01-01

    The use of stem/progenitor cells represents a promising approach to treat craniofacial bone defects, but successful treatments will rely on the availability of cells that can be expanded in vitroand which will differentiate appropriately in vivo. The calvaria may represent a source of autologous cells for such purposes. We demonstrate expression of stem cell antigen-1 (Sca-1) in mouse calvaria. We isolated Sca-1+ and Sca-1– cells at high purity and tested the ability of these cells to differentiate into adipose and bone. We show that the Sca-1+ cell fraction has adipogenic differentiation potential and that the cell Sca-1– fraction has osteogenic differentiation potential. The Sca-1+ cell fraction partially retains its adipogenic differentiation potential and the Sca-1– cell fraction partially retains its osteogenic differentiation potential after in vitroexpansion. These data suggest that the calvaria may be used as a source of stem/progenitor cells that can be expanded in vitroand transplanted in vivofor craniofacial tissue regeneration. PMID:19088466

  9. Proteomic analysis of the cell envelope fraction of Escherichia coli.

    PubMed

    Fountoulakis, M; Gasser, R

    2003-01-01

    We applied proteomics technologies to analyze a membrane preparation of Escherichia coli, wild type strain and of transformants expressing human cytochrome P450s. The proteins were analyzed by two-dimensional electrophoresis and identified by matrix-assisted laser desorption ionization mass spectrometry. The membrane proteins were solubilized with both mild detergents such as CHAPS and strong detergents, such as sodium and lithium dodecyl sulfate, sodium cholate and sodium deoxycholate. In the E. colimembrane fraction, 394 different gene products were identified. Approximately 28% of them were predicted to be integral membrane proteins, of which 100 proteins have been predicted to carry one transmembrane region, ten proteins to carry two, and two proteins to include three transmembrane domains. The remaining are probably membrane-associated and cytosolic proteins. Cytochrome P450s did not enter the immobilized pH gradient strips but were efficiently analyzed in a two-dimensional, two-detergent system. Use of strong solubilizing agents resulted in the detection of about 20 membrane proteins, which were not detected following extraction with mild detergents and chaotropes. The present database is one of the largest for membrane proteins.

  10. Phosphatidic acid phosphatase and phospholipdase A activities in plasma membranes from fusing muscle cells.

    PubMed

    Kent, C; Vagelos, P R

    1976-06-17

    Plasma membrane from fusing embryonic muscle cells were assayed for phospholipase A activity to determine if this enzyme plays a role in cell fusion. The membranes were assayed under a variety of conditions with phosphatidylcholine as the substrate and no phospholipase A activity was found. The plasma membranes did contain a phosphatidic acid phosphatase which was optimally active in the presence of Triton X-100 and glycerol. The enzyme activity was constant from pH 5.2 to 7.0, and did not require divalent cations. Over 97% of the phosphatidic acid phosphatase activity was in the particulate fraction. The subcellular distribution of the phosphatidic acid phosphatase was the same as the distributions of the plasma membrane markers, (Na+ + k+)-ATPase and the acetylcholine receptor, which indicates that this phosphatase is located exclusively in the plasma membranes. There was no detectable difference in the phosphatidic acid phosphatase activities of plasma membranes from fusing and non-fusing cells.

  11. Integrated stereological and biochemical studies on hepatocytic membranes. I.V. Heterogeneous distribution of marker enzymes on endoplasmic reticulum membranes in fractions

    PubMed Central

    1980-01-01

    The purpose of the study was to consider quantitatively the relationships between the surface area of the endoplasmic reticulum (ER) and constituent marker enzyme activities, as they occur in fractions collected from rat liver homogenates. The ER surface area was estimated in five membrane-containing fractions by use of a combined cytochemical-stereological technique (5), while, at the same time, ER marker enzymes were assayed biochemically. Fraction/homogenate recoveries for the ER enzymes averaged 100%, total membrane surface area 98%, and ER surface area 96%. Relative specific activities, which compare the relative amounts of ER marker enzyme activities to the relative ER surface area in the membrane-containing fractions, indicate variable distributions for glucose-6-phosphatase and NADPH cytochrome c reductase, but not for esterase. PMID:6248565

  12. Influence of hydrophobic/hydrophilic fractions of extracellular organic matters of Microcystis aeruginosa on ultrafiltration membrane fouling.

    PubMed

    Zhou, Shiqing; Shao, Yisheng; Gao, Naiyun; Li, Lei; Deng, Jing; Tan, Chaoqun; Zhu, Mingqiu

    2014-02-01

    Fouling is a major obstacle to maintain the efficiency of ultrafiltration-based drinking water treatment process. Algal extracellular organic matters (EOMs) are currently considered as one of the major sources of membrane fouling. The objective of this study was to investigate the influence of different hydrophobic/hydrophilic fractions of EOM extracted from Microcystis aeruginosa on ultrafiltration membrane fouling at lab scale. The experimental data indicated that EOM exhibited similar flux decline trends on polyethersulfone (PES) and regenerated cellulose (RC) membranes but caused greater irreversible fouling on PES membrane than RC membrane due to its hydrophobic property. It was also observed that charged hydrophilic (CHPI) and neutral hydrophilic (NHPI) fractions caused greater flux decline over hydrophobic (HPO) and transphilic (TPI) fractions. For PES membrane, the order of the irreversible fouling potentials for the four fractions was HPO>TPI>CHPI>NHPI, while the irreversible fouling potentials of RC membrane were tiny and could be ignored. Fluorescence excitation-emission matrix (EEM) spectra and Fourier transform infrared (FTIR) spectra suggested that protein-like, polysaccharide-like and humic-like substances were the major components responsible for membrane fouling. The results also indicated that the irreversible fouling increased as the pH decreased. The addition of calcium to feed solutions led to more severe flux decline and irreversible fouling.

  13. Polyphosphoinositides are present in plasma membranes isolated from fusogenic carrot cells

    SciTech Connect

    Wheeler, J.J.; Boss, W.F.

    1987-10-01

    Fusogenic carrot cells grown in suspension culture were labeled 12 hours with myo-(2-/sup 3/H)inositol. Plasma membranes were isolated from the prelabeled fusogenic carrot cells by both aqueous polymer two-phase partitioning and Renografin density gradients. With both methods, the plasma membrane-enriched fractions, as identified by marker enzymes, were enriched in (/sup 3/H)inositol-labeled phosphatidylinositol monophosphate (PIP) and phosphatidylinositol bisphosphate (PIP/sub 2/). An additional (/sup 3/H)inositol-labeled lipid, lysophosphatidylinositol monophosphate, which migrated between PIP and PIP/sub 2/ on thin layer plates, was found primarily in the plasma membrane-rich fraction of the fusogenic cells. This was in contrast to lysophosphatidylinositol which is found primarily in the lower phase, microsomal/mitchrondrial-rich fraction.

  14. Membrane electrode assembly for a fuel cell

    NASA Technical Reports Server (NTRS)

    Prakash, Surya (Inventor); Narayanan, Sekharipuram R. (Inventor); Atti, Anthony (Inventor); Olah, George (Inventor); Smart, Marshall C. (Inventor)

    2006-01-01

    A catalyst ink for a fuel cell including a catalytic material and poly(vinylidene fluoride). The ink may be applied to a substrate to form an electrode, or bonded with other electrode layers to form a membrane electrode assembly (MEA).

  15. Artificial Red Cells with Polyhemoglobin Membranes.

    DTIC Science & Technology

    1981-09-01

    preparing emulsions and ejecting cells from the oil phase. IX. REFERENCES 1. Wallace, H. W., Asher, W. J., and Li, N. N. Liquid - liquid oxygenation: a...1S. KEY WORDS (Continue, an reverse side if naceoay mnd identify by block number) Artificial Blood, Hemoglobin, Polyhemoglobin, Biotonometry Liquid ...cell-size microdroplets containing 30% of hemoglobin were held in liquid membrane capsules and treated with glutaralddhyde that cross linked the

  16. Proteomic Analysis of Lipid Raft-Like Detergent-Resistant Membranes of Lens Fiber Cells

    PubMed Central

    Wang, Zhen; Schey, Kevin L.

    2015-01-01

    Purpose Plasma membranes of lens fiber cells have high levels of long-chain saturated fatty acids, cholesterol, and sphingolipids—key components of lipid rafts. Thus, lipid rafts are expected to constitute a significant portion of fiber cell membranes and play important roles in lens biology. The purpose of this study was to characterize the lens lipid raft proteome. Methods Quantitative proteomics, both label-free and iTRAQ methods, were used to characterize lens fiber cell lipid raft proteins. Detergent-resistant, lipid raft membrane (DRM) fractions were isolated by sucrose gradient centrifugation. To confirm protein localization to lipid rafts, protein sensitivity to cholesterol removal by methyl-β-cyclodextrin was quantified by iTRAQ analysis. Results A total of 506 proteins were identified in raft-like detergent-resistant membranes. Proteins identified support important functions of raft domains in fiber cells, including trafficking, signal transduction, and cytoskeletal organization. In cholesterol-sensitivity studies, 200 proteins were quantified and 71 proteins were strongly affected by cholesterol removal. Lipid raft markers flotillin-1 and flotillin-2 and a significant fraction of AQP0, MP20, and AQP5 were found in the DRM fraction and were highly sensitive to cholesterol removal. Connexins 46 and 50 were more abundant in nonraft fractions, but a small fraction of each was found in the DRM fraction and was strongly affected by cholesterol removal. Quantification of modified AQP0 confirmed that fatty acylation targeted this protein to membrane raft domains. Conclusions These data represent the first comprehensive profile of the lipid raft proteome of lens fiber cells and provide information on membrane protein organization in these cells. PMID:26747763

  17. The mechanism of facilitated cell membrane resealing.

    PubMed

    Togo, T; Alderton, J M; Bi, G Q; Steinhardt, R A

    1999-03-01

    Disruption of the plasma membrane evokes an exocytotic response that is required for rapid membrane resealing. We show here in Swiss 3T3 fibroblasts that a second disruption at the same site reseals more rapidly than the initial wound. This facilitated response of resealing was inhibited by both low external Ca2+ concentration and specific protein kinase C (PKC) inhibitors, bisindolylmaleimide I (BIS) and Gö-6976. In addition, activation of PKC by phorbol ester facilitated the resealing of a first wound. BIS and Gö-6976 suppressed the effect of phorbol ester on resealing rate. Fluorescent dye loss from a FM1-43 pre-labeled endocytotic compartment was used to investigate the relationship between exocytosis, resealing and the facilitation of resealing. Exocytosis of endocytotic compartments near the wounding site was correlated with successful resealing. The destaining did not occur when exocytosis and resealing were inhibited by low external Ca2+ concentration or by injected tetanus toxin. When the dye loaded cells were wounded twice, FM1-43 destaining at the second wound was less than at the first wound. Less destaining was also observed in cells pre-treated with phorbol ester, suggesting newly formed vesicles, which were FM1-43 unlabeled, were exocytosed in the resealing at repeated woundings. Facilitation was also blocked by brefeldin A (BFA), a fungal metabolite that inhibits vesicle formation at the Golgi apparatus. Lowering the temperature below 20 degrees C also blocked facilitation as expected from a block of Golgi function. BFA had no effect on the resealing rate of an initial wound. The facilitation of the resealing by phorbol ester was blocked by pre-treatment with BFA. These results suggest that at first wounding the cell used the endocytotic compartment to add membrane necessary for resealing. At a second wounding, PKC, activated by Ca2+ entry at the first wound, stimulated vesicle formation from the Golgi apparatus, resulting in more rapid resealing

  18. Selectivity of Direct Methanol Fuel Cell Membranes.

    PubMed

    Aricò, Antonino S; Sebastian, David; Schuster, Michael; Bauer, Bernd; D'Urso, Claudia; Lufrano, Francesco; Baglio, Vincenzo

    2015-11-24

    Sulfonic acid-functionalized polymer electrolyte membranes alternative to Nafion(®) were developed. These were hydrocarbon systems, such as blend sulfonated polyetheretherketone (s-PEEK), new generation perfluorosulfonic acid (PFSA) systems, and composite zirconium phosphate-PFSA polymers. The membranes varied in terms of composition, equivalent weight, thickness, and filler and were investigated with regard to their methanol permeation characteristics and proton conductivity for application in direct methanol fuel cells. The behavior of the membrane electrode assemblies (MEA) was investigated in fuel cell with the aim to individuate a correlation between membrane characteristics and their performance in a direct methanol fuel cell (DMFC). The power density of the DMFC at 60 °C increased according to a square root-like function of the membrane selectivity. This was defined as the reciprocal of the product between area specific resistance and crossover. The power density achieved at 60 °C for the most promising s-PEEK-based membrane-electrode assembly (MEA) was higher than the benchmark Nafion(®) 115-based MEA (77 mW·cm(-2) vs. 64 mW·cm(-2)). This result was due to a lower methanol crossover (47 mA·cm(-2) equivalent current density for s-PEEK vs. 120 mA·cm(-2) for Nafion(®) 115 at 60 °C as recorded at OCV with 2 M methanol) and a suitable area specific resistance (0.15 Ohm cm² for s-PEEK vs. 0.22 Ohm cm² for Nafion(®) 115).

  19. Selectivity of Direct Methanol Fuel Cell Membranes

    PubMed Central

    Aricò, Antonino S.; Sebastian, David; Schuster, Michael; Bauer, Bernd; D’Urso, Claudia; Lufrano, Francesco; Baglio, Vincenzo

    2015-01-01

    Sulfonic acid-functionalized polymer electrolyte membranes alternative to Nafion® were developed. These were hydrocarbon systems, such as blend sulfonated polyetheretherketone (s-PEEK), new generation perfluorosulfonic acid (PFSA) systems, and composite zirconium phosphate–PFSA polymers. The membranes varied in terms of composition, equivalent weight, thickness, and filler and were investigated with regard to their methanol permeation characteristics and proton conductivity for application in direct methanol fuel cells. The behavior of the membrane electrode assemblies (MEA) was investigated in fuel cell with the aim to individuate a correlation between membrane characteristics and their performance in a direct methanol fuel cell (DMFC). The power density of the DMFC at 60 °C increased according to a square root-like function of the membrane selectivity. This was defined as the reciprocal of the product between area specific resistance and crossover. The power density achieved at 60 °C for the most promising s-PEEK-based membrane-electrode assembly (MEA) was higher than the benchmark Nafion® 115-based MEA (77 mW·cm−2 vs. 64 mW·cm−2). This result was due to a lower methanol crossover (47 mA·cm−2 equivalent current density for s-PEEK vs. 120 mA·cm−2 for Nafion® 115 at 60 °C as recorded at OCV with 2 M methanol) and a suitable area specific resistance (0.15 Ohm cm2 for s-PEEK vs. 0.22 Ohm cm2 for Nafion® 115). PMID:26610582

  20. Integrin-like proteins are localized to plasma membrane fractions, not plastids, in Arabidopsis

    NASA Technical Reports Server (NTRS)

    Swatzell, L. J.; Edelmann, R. E.; Makaroff, C. A.; Kiss, J. Z.

    1999-01-01

    Integrins are a large family of integral membrane proteins that function in signal transduction in animal systems. These proteins are conserved in vertebrates, invertebrates, and fungi. Evidence from previous research suggests that integrin-like proteins may be present in plants as well, and that these proteins may function in signal transduction during gravitropism. In past studies, researchers have used monoclonal and polyclonal antibodies to localize beta 1 integrin-like proteins in plants. However, there is a disparity between data collected from these studies, especially since molecular weights obtained from these investigations range from 55-120 kDa for integrin-like proteins. To date, a complete investigation which employs all three basic immunolabeling procedures, immunoblotting, immunofluorescence microscopy, and immunogold labeling, in addition to extensive fractionation and exhaustive controls, has been lacking. In this paper, we demonstrate that use of a polyclonal antibody against the cytoplasmic domain of avian beta 1-integrin can produce potential artifacts in immunolocalization studies. However, these problems can be eliminated through use of starchless mutants or proper specimen preparation prior to electrophoresis. We also show that this antibody, when applied within the described parameters and with careful controls, identifies a large (100 kDa) integrin-like protein that is localized to plasma membrane fractions in Arabidopsis.

  1. Catalytic membranes for fuel cells

    SciTech Connect

    Liu, Di-Jia; Yang, Junbing; Wang, Xiaoping

    2011-04-19

    A fuel cell of the present invention comprises a cathode and an anode, one or both of the anode and the cathode including a catalyst comprising a bundle of longitudinally aligned graphitic carbon nanotubes including a catalytically active transition metal incorporated longitudinally and atomically distributed throughout the graphitic carbon walls of said nanotubes. The nanotubes also include nitrogen atoms and/or ions chemically bonded to the graphitic carbon and to the transition metal. Preferably, the transition metal comprises at least one metal selected from the group consisting of Fe, Co, Ni, Mn, and Cr.

  2. Microfluidic microbial fuel cells: from membrane to membrane free

    NASA Astrophysics Data System (ADS)

    Yang, Yang; Ye, Dingding; Li, Jun; Zhu, Xun; Liao, Qiang; Zhang, Biao

    2016-08-01

    Microfluidic microbial fuel cells (MMFCs) are small carbon-neutral devices that use self-organized bacteria to degrade organic substrates and harness energy from the waste water. Conventional MMFCs have made great strides in the past decade and have overcome some limitations, such as high capital costs and low energy output. A co-laminar flow MFC has been first proposed in 2011 with the potential to be an attractively power source to niche applications. Co-laminar MFCs typically operate without any physical membranes separating the reactants, and bacterial ecosystems can be easily manipulated by regulating the inlet conditions. This paper highlights recent accomplishments in the development of co-laminar MFCs, emphasizing basic principles, mass transport and fluid dynamics including boundary layer theory, entrance conditions and mixing zone issues. Furthermore, the development of current techniques, major challenges and the potential research directions are discussed.

  3. Size Control and Fractionation of Ionic Liquid Filled Polymersomes with Glassy and Rubbery Bilayer Membranes.

    PubMed

    So, Soonyong; Lodge, Timothy P

    2016-05-17

    We demonstrate control over the size of ionic liquid (IL) filled polymeric vesicles (polymersomes) by three distinct methods: mechanical extrusion, cosolvent-based processing in an IL, and fractionation of polymersomes in a biphasic system of IL and water. For the representative ionic liquid (1-ethyl-3-methylimidazolium bis(trifluoromethylsulfonyl) imide ([EMIM][TFSI])), the size and dispersity of polymersomes formed from 1,2-polybutadiene-b-poly(ethylene oxide) (PB-PEO) and polystyrene-b-poly(ethylene oxide) (PS-PEO) diblock copolymers were shown to be sensitive to assembly conditions. During mechanical extrusion through a polycarbonate membrane, the relatively larger polymersomes were broken up and reorganized into vesicles with mean size comparable to the membrane pore (100 nm radius); the distribution width also decreased significantly after only a few passes. Other routes were studied using the solvent-switch or cosolvent (CS) method, whereby the initial content of the cosolvent and the PEO block length of PS-PEO were systemically changed. The nonvolatility of the ionic liquid directly led to the desired concentration of polymersomes in the ionic liquid using a single step, without the dialysis conventionally used in aqueous systems, and the mean vesicle size depended on the amount of cosolvent employed. Finally, selective phase transfer of PS-PEO polymersomes based on size was used to extract larger polymersomes from the IL to the aqueous phase via interfacial tension controlled phase transfer. The interfacial tension between the PS membrane and the aqueous phase was varied with the concentration of sodium chloride (NaCl) in the aqueous phase; then the larger polymersomes were selectively separated to the aqueous phase due to differences in shielding of the hydrophobic core (PS) coverage by the hydrophilic corona brush (PEO). This novel fractionation is a simple separation process without any special apparatus and can help to prepare monodisperse polymersomes

  4. Biochemical characterization of a V-ATPase of tracheal smooth muscle plasma membrane fraction.

    PubMed

    Pacheco, G; Lippo de Bécemberg, I; Gonzalez de Alfonzo, R; Alfonzo, M J

    1996-07-25

    A biochemical characterization of a Mg(2+)-ATPase activity associated with a plasma membrane fraction isolated from airway (tracheal) smooth muscle was performed. This enzyme is an integral part of the membrane remaining tightly bound after 0.6 M KCl extraction. This enzyme activity showed a cold inactivation in the presence of ATP and Mg2+. Also, this Mg(2+)-ATPase was stimulated by monovalent anions being Cl-, the best anion for such stimulation, even though Br- and I- were good substitutes and F- was ineffective. This Cl--stimulated activity showed a powerful nucleosidetriphosphatase activity having the following divalent cation specificity: Mg2+ > Mn2+ > Ca2+, where Zn2+ and Fe2+ were ineffective. This ATPase activity was not inhibited by ouabain oligomycin C and vanadate indicating that neither P- or F-ATPases were associated with this enzyme activity. However, the existence of a V-ATPase was shown by the significant inhibition causes by bafilomycin A1. Additionally, this V-ATPase seems to be coupled to Cl- conductor because duramycin inhibited this ATPase activity. The presence of a H+ pump associated to this V-ATPase was shown indirectly, through the stimulatory effect produced by uncouplers such as FCCP and 1799, which were able to produce significant stimulation of this V-ATPase indicating the existence of a H(+)-ATPase. Finally, the immunodetection of a 72 kDa polypeptide using a specific antibody against the A subunit (72 kDa) of V-ATPase from chromaffin granule demonstrated the presence of a V-ATPase in this plasma membrane fraction.

  5. Interaction of peptides with cell membranes: insights from molecular modeling

    NASA Astrophysics Data System (ADS)

    Li, Zhen-lu; Ding, Hong-ming; Ma, Yu-qiang

    2016-03-01

    The investigation of the interaction of peptides with cell membranes is the focus of active research. It can enhance the understanding of basic membrane functions such as membrane transport, fusion, and signaling processes, and it may shed light on potential applications of peptides in biomedicine. In this review, we will present current advances in computational studies on the interaction of different types of peptides with the cell membrane. Depending on the properties of the peptide, membrane, and external environment, the peptide-membrane interaction shows a variety of different forms. Here, on the basis of recent computational progress, we will discuss how different peptides could initiate membrane pores, translocate across the membrane, induce membrane endocytosis, produce membrane curvature, form fibrils on the membrane surface, as well as interact with functional membrane proteins. Finally, we will present a conclusion summarizing recent progress and providing some specific insights into future developments in this field.

  6. Durability aspects of polymer electrolyte membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Sethuraman, Vijay Anand

    In order for the successful adoption of proton exchange membrane (PEM) fuel cell technology, it is imperative that durability is understood, quantified and improved. A number of mechanisms are known to contribute to PEMFC membrane electrode assembly (MEA) performance degradation. In this dissertation, we show, via experiments, some of the various processes that degrade the proton exchange membrane in a PEM fuel cell; and catalyst poisoning due to hydrogen sulfide (H2S) and siloxane. The effect of humidity on the chemical stability of two types of membranes, [i.e., perfluorosulfonic acid type (PFSA, NafionRTM 112) and biphenyl sulfone hydrocarbon type, (BPSH-35)] was studied by subjecting the MEAs to open-circuit voltage (OCV) decay and potential cycling tests at elevated temperatures and low inlet gas relative humidities. The BPSH-35 membranes showed poor chemical stability in ex situ Fenton tests compared to that of NafionRTM membranes. However, under fuel cell conditions, BPSH-35 MEAs outperformed NafionRTM 112 MEAs in both the OCV decay and potential cycling tests. For both membranes, (i) at a given temperature, membrane degradation was more pronounced at lower humidities and (ii) at a given relative humidity operation, increasing the cell temperature accelerated membrane degradation. Mechanical stability of these two types of membranes was also studied using relative humidity (RH) cycling. Hydrogen peroxide (H2O2) formation rates in a proton exchange membrane (PEM) fuel cell were estimated by studying the oxygen reduction reaction (ORR) on a rotating ring disc electrode (RRDE). Fuel cell conditions were replicated by depositing a film of Pt/Vulcan XC-72 catalyst onto the disk and by varying the temperature, dissolved O2 concentration and the acidity levels in HClO4. The HClO4 acidity was correlated to ionomer water activity and hence fuel cell humidity. H 2O2 formation rates showed a linear dependence on oxygen concentration and square dependence on water

  7. Sputter-deposited fuel cell membranes and electrodes

    NASA Technical Reports Server (NTRS)

    Narayanan, Sekharipuram R. (Inventor); Jeffries-Nakamura, Barbara (Inventor); Chun, William (Inventor); Ruiz, Ron P. (Inventor); Valdez, Thomas I. (Inventor)

    2001-01-01

    A method for preparing a membrane for use in a fuel cell membrane electrode assembly includes the steps of providing an electrolyte membrane, and sputter-depositing a catalyst onto the electrolyte membrane. The sputter-deposited catalyst may be applied to multiple sides of the electrolyte membrane. A method for forming an electrode for use in a fuel cell membrane electrode assembly includes the steps of obtaining a catalyst, obtaining a backing, and sputter-depositing the catalyst onto the backing. The membranes and electrodes are useful for assembling fuel cells that include an anode electrode, a cathode electrode, a fuel supply, and an electrolyte membrane, wherein the electrolyte membrane includes a sputter-deposited catalyst, and the sputter-deposited catalyst is effective for sustaining a voltage across a membrane electrode assembly in the fuel cell.

  8. Dynamic physical properties of dissociated tumor cells revealed by dielectrophoretic field-flow fractionation

    PubMed Central

    Shim, Sangjo; Gascoyne, Peter; Noshari, Jamileh; Stemke Hale, Katherine

    2013-01-01

    Metastatic disease results from the shedding of cancer cells from a solid primary tumor, their transport through the cardiovascular system as circulating tumor cells (CTCs) and their engraftment and growth at distant sites. Little is known about the properties and fate of tumor cells as they leave their growth site and travel as single cells. We applied analytical dielectrophoretic field-flow fractionation (dFFF) to study the membrane capacitance, density and hydrodynamic properties together with the size and morphology of cultured tumor cells after they were harvested and placed into single cell suspensions. After detachment, the tumor cells exhibited biophysical properties that changed with time through a process of cytoplasmic shedding whereby membrane and cytoplasm were lost. This process appeared to be distinct from the cell death mechanisms of apoptosis, anoikis and necrosis and it may explain why multiple phenotypes are seen among CTCs isolated from patients and among the tumor cells obtained from ascitic fluid of patients. The implications of dynamic biophysical properties and cytoplasmic loss for CTC migration into small blood vessels in the circulatory system, survival and gene expression are discussed. Because the total capacitance of tumor cells remained higher than blood cells even after they had shed cytoplasm, dFFF offers a compelling, antibody-independent technology for isolating viable CTCs from blood even when they are no larger than peripheral blood mononuclear cells. PMID:21691666

  9. The plasma membrane-enriched fraction proteome response during adaptation to hydrogen peroxide in Saccharomyces cerevisiae.

    PubMed

    Pedroso, Nuno; Gomes-Alves, Patrícia; Marinho, H Susana; Brito, Verônica B; Boada, Cristina; Antunes, Fernando; Herrero, Enrique; Penque, Deborah; Cyrne, Luísa

    2012-10-01

    In Saccharomyces cerevisiae, adaptation to hydrogen peroxide (H₂O₂) decreases plasma membrane permeability to H₂O₂, changes its lipid composition and reorganizes ergosterol-rich microdomains by a still unknown mechanism. Here we show, by a quantitative analysis of the H₂O₂-induced adaptation effect on the S. cerevisiae plasma membrane-enriched fraction proteome, using two-dimensional gel electrophoresis, that 44 proteins are differentially expressed. Most of these proteins were regulated at a post-transcriptional level. Fourteen of these proteins contain redox-sensitive cysteine residues and nine proteins are associated with lipid and vesicle traffic. In particular, three proteins found in eisosomes and in the eisosome-associated membrane compartment occupied by Can1p were up-regulated (Pil1p, Rfs1p and Pst2p) during adaptation to H₂O₂. Survival studies after exposure to lethal H₂O₂ doses using yeast strains bearing a gene deletion corresponding to proteins associated to lipid and vesicle traffic demonstrated for the first time that down-regulation of Kes1p, Vps4p and Ynl010wp and up-regulation of Atp1 and Atp2 increases resistance to H₂O₂. Moreover, for the pil1Δ strain, H₂O₂ at low levels produces a hormetic effect by increasing proliferation. In conclusion, these data further confirms the plasma membrane as an active cellular site during adaptation to H₂O₂ and shows that proteins involved in lipid and vesicle traffic are important mediators of H₂O₂ adaptation.

  10. Molecular dynamics study of lipid bilayers modeling the plasma membranes of normal murine thymocytes and leukemic GRSL cells.

    PubMed

    Andoh, Yoshimichi; Okazaki, Susumu; Ueoka, Ryuichi

    2013-04-01

    Molecular dynamics (MD) calculations for the plasma membranes of normal murine thymocytes and thymus-derived leukemic GRSL cells in water have been performed under physiological isothermal-isobaric conditions (310.15K and 1 atm) to investigate changes in membrane properties induced by canceration. The model membranes used in our calculations for normal and leukemic thymocytes comprised 23 and 25 kinds of lipids, respectively, including phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, sphingomyelin, lysophospholipids, and cholesterol. The mole fractions of the lipids adopted here were based on previously published experimental values. Our calculations clearly showed that the membrane area was increased in leukemic cells, and that the isothermal area compressibility of the leukemic plasma membranes was double that of normal cells. The calculated membranes of leukemic cells were thus considerably bulkier and softer in the lateral direction compared with those of normal cells. The tilt angle of the cholesterol and the conformation of the phospholipid fatty acid tails both showed a lower level of order in leukemic cell membranes compared with normal cell membranes. The lateral radial distribution function of the lipids also showed a more disordered structure in leukemic cell membranes than in normal cell membranes. These observations all show that, for the present thymocytes, the lateral structure of the membrane is considerably disordered by canceration. Furthermore, the calculated lateral self-diffusion coefficient of the lipid molecules in leukemic cell membranes was almost double that in normal cell membranes. The calculated rotational and wobbling autocorrelation functions also indicated that the molecular motion of the lipids was enhanced in leukemic cell membranes. Thus, here we have demonstrated that the membranes of thymocyte leukemic cells are more disordered and more fluid than normal cell membranes.

  11. Copper transporters are responsible for copper isotopic fractionation in eukaryotic cells

    PubMed Central

    Cadiou, Jean-Loup; Pichat, Sylvain; Bondanese, Victor P.; Soulard, Alexandre; Fujii, Toshiyuki; Albarède, Francis; Oger, Philippe

    2017-01-01

    Copper isotopic composition is altered in cancerous compared to healthy tissues. However, the rationale for this difference is yet unknown. As a model of Cu isotopic fractionation, we monitored Cu uptake in Saccharomyces cerevisiae, whose Cu import is similar to human. Wild type cells are enriched in 63Cu relative to 65Cu. Likewise, 63Cu isotope enrichment in cells without high-affinity Cu transporters is of slightly lower magnitude. In cells with compromised Cu reductase activity, however, no isotope fractionation is observed and when Cu is provided solely in reduced form for this strain, copper is enriched in 63Cu like in the case of the wild type. Our results demonstrate that Cu isotope fractionation is generated by membrane importers and that its amplitude is modulated by Cu reduction. Based on ab initio calculations, we propose that the fractionation may be due to Cu binding with sulfur-rich amino acids: methionine and cysteine. In hepatocellular carcinoma (HCC), lower expression of the STEAP3 copper reductase and heavy Cu isotope enrichment have been reported for the tumor mass, relative to the surrounding tissue. Our study suggests that copper isotope fractionation observed in HCC could be due to lower reductase activity in the tumor. PMID:28303916

  12. Possible association of Neu2 with plasma membrane fraction from mouse thymus exhibited sialidase activity with fetuin at pH 7.0 but not at pH 4.5.

    PubMed

    Kijimoto-Ochiai, Shigeko; Doi, Naoko; Fujii, Miwako; Go, Shinji; Kabayama, Kazuya; Moriya, Setsuko; Miyagi, Taeko; Koda, Toshiaki

    2013-08-01

    Compared to other organs, the mouse thymus exhibits a high level of sialidase activity in both the soluble and crude membrane fractions, as measured at neutral pH using 4MU-Neu5Ac as a substrate. The main purpose of the present study was to identify the sialidase with a high level of the activity at neutral pH in the crude membrane. Several parameters were analyzed using the soluble (S) fraction, N and D fractions that were obtained by NP-40 or DOC/NP-40 solubilization from the thymus crude membrane. The main sialidase activity in the N fraction exhibited almost the same pI as that of soluble Neu2 and 60% of the activity was removed from the membrane by three washes with 10 mM Tris-buffer, at pH 7.0. The N fraction preferentially hydrolyzed the sialic acid bond of glycoprotein and exhibited sialidase activity with fetuin at pH 7.0 but not at pH 4.5. The same activity was observed in a plasma membrane-rich fraction. To date, the removal of sialic acid from fetuin at pH 7.0 was reported only with soluble Neu2 and the membrane fraction from Neu2-transfected COS cells. We analyzed the gene that controls the sialidase activity in the crude membrane fraction at pH 7.0 using SMXA recombinant mice and found that compared with other three genes, Neu2 presented the best correlation with the activity level. We suggest that Neu2 is most likely responsible for the main activity in the N fraction, due to its association with the membrane by an unknown mechanism.

  13. Fuel cell membranes and crossover prevention

    DOEpatents

    Masel, Richard I.; York, Cynthia A.; Waszczuk, Piotr; Wieckowski, Andrzej

    2009-08-04

    A membrane electrode assembly for use with a direct organic fuel cell containing a formic acid fuel includes a solid polymer electrolyte having first and second surfaces, an anode on the first surface and a cathode on the second surface and electrically linked to the anode. The solid polymer electrolyte has a thickness t:.gtoreq..times..times..times..times. ##EQU00001## where C.sub.f is the formic acid fuel concentration over the anode, D.sub.f is the effective diffusivity of the fuel in the solid polymer electrolyte, K.sub.f is the equilibrium constant for partition coefficient for the fuel into the solid polymer electrolyte membrane, I is Faraday's constant n.sub.f is the number of electrons released when 1 molecule of the fuel is oxidized, and j.sub.f.sup.c is an empirically determined crossover rate of fuel above which the fuel cell does not operate.

  14. Embryonal cell surface recognition. Extraction of an active plasma membrane component.

    PubMed

    Merrell, R; Gottlieb, D I; Glaser, L

    1975-07-25

    Plasma membranes obtained from different neural regions of the chicken embryo have previously been shown to specifically bind to homotypic cells and prevent cell aggregation (Merrell, R., and Glaser, L. (1973) Proc. Natl. Acad. Sci. U. S. A. 70, 2794-2798). Proteins responsible for the specific inhibition of cell aggregation have been solubilized from the plasma membrane of neural retina and optic tectum by delipidation with acetone followed by extraction with lithium diiodosalicylate. The extracts show the same regional and temporal specificity as previously shown for plasma membrane recognition by the same cells (Gottlieb, D. I., Merrell, R., and Glaser, L. (1974) Proc. Natl. Acad. Sci. U. S. A. 71, 1800-1802). Two micrograms of the most purified protein fraction inhibits the aggregation of 2.5 times 10(-4) cells under standard assay conditions. This represents a 20-fold increase in specific activity compared to whole membranes.

  15. Interaction of Inorganic Nanoparticles With Cell Membranes

    DTIC Science & Technology

    2008-10-20

    explain the change in the Zeta-potential of the beads we studied the adsorption of protein on Chitosan coated SPIONs. The particles were incubated in...protein adsorption which enables us understand better the pathway of our particles through the membrane and inside the cell. Combined with...investigation regarding the protein adsorption and their influence on the colloidal stability we have now the tools to investigate and perhaps to understand

  16. Virus-Mimetic Fusogenic Exosomes for Direct Delivery of Integral Membrane Proteins to Target Cell Membranes.

    PubMed

    Yang, Yoosoo; Hong, Yeonsun; Nam, Gi-Hoon; Chung, Jin Hwa; Koh, Eunee; Kim, In-San

    2017-02-06

    An efficient system for direct delivery of integral membrane proteins is successfully developed using a new biocompatible exosome-based platform. Fusogenic exosomes harboring viral fusogen, vascular stomatitis virus (VSV)-G protein, can fuse with and modify plasma membranes in a process called "membrane editing." This can facilitate the transfer of biologically active membrane proteins into the target cell membranes both in vitro and in vivo.

  17. Membrane catalyst layer for fuel cells

    DOEpatents

    Wilson, Mahlon S.

    1993-01-01

    A gas reaction fuel cell incorporates a thin catalyst layer between a solid polymer electrolyte (SPE) membrane and a porous electrode backing. The catalyst layer is preferably less than about 10 .mu.m in thickness with a carbon supported platinum catalyst loading less than about 0.35 mgPt/cm.sup.2. The film is formed as an ink that is spread and cured on a film release blank. The cured film is then transferred to the SPE membrane and hot pressed into the surface to form a catalyst layer having a controlled thickness and catalyst distribution. Alternatively, the catalyst layer is formed by applying a Na.sup.+ form of a perfluorosulfonate ionomer directly to the membrane, drying the film at a high temperature, and then converting the film back to the protonated form of the ionomer. The layer has adequate gas permeability so that cell performance is not affected and has a density and particle distribution effective to optimize proton access to the catalyst and electronic continuity for electron flow from the half-cell reaction occurring at the catalyst.

  18. A review of radiation-grafted polymer electrolyte membranes for alkaline polymer electrolyte membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Zhou, Tianchi; Shao, Rong; Chen, Song; He, Xuemei; Qiao, Jinli; Zhang, Jiujun

    2015-10-01

    The past two decades have witnessed many efforts to develop radiation-grafted alkaline membranes for alkaline PEM fuel cell applications, as such membranes have certain advantages over other kinds of alkaline membranes, including well-controlled composition, functionality, and other promising properties. To facilitate research and development in this area, the present paper reviews radiation-grafted alkaline membranes. We examine their synthesis/fabrication/characterization, membrane material selection, and theoretical approaches for fundamental understanding. We also present detailed examinations of their application in fuel cell in terms of the working principles of the radiation grafting process, the fabrication of MEAs using radiation-grafted membranes, the membranes' corresponding performance in alkaline PEM fuel cells, as well as performance optimization. The paper also summarizes the challenges and mitigation strategies for radiation-grafted alkaline membranes and their application in PEM fuel cells, presenting an overall picture of the technology as it presently stands.

  19. Biodegradation characteristics and size fractionation of landfill leachate for integrated membrane treatment.

    PubMed

    Insel, Güçlü; Dagdar, Mina; Dogruel, Serdar; Dizge, Nadir; Ubay Cokgor, Emine; Keskinler, Bülent

    2013-09-15

    The fate of organics and nitrogen during the biological treatment with MBR and subsequent membrane filtration processes (nano filtration, NF; reverse osmosis, RO) were investigated for a landfill leachate. The chemical oxygen demand (COD) and total Kjeldahl nitrogen (TKN) removal performances of membrane bioreactor (MBR) were obtained to be around 89% and 85%, respectively. The effluent COD of MBR was measured to be 1935 mg/L (30 kDa) which is much lower than experimentally determined soluble inert COD of 3200 mg/L using 0.45 μm filter. The readily and slowly biodegradable COD fractions were estimated to be 17% and 52% of raw influent COD, respectively. The respirometry based modeling test performed on raw leachate exhibited much slower degradation kinetics compared to municipal wastewater. A unique subset of model parameters was extracted from batch respirometry by using acclimated MBR sludge. The sequential ultrafiltration (UF) experiments (particle size distribution, PSD) revealed that most of the organics was below 2 nm filter mesh size. In addition, NF/RO post treatment after MBR system was required to increase COD and total nitrogen (TN) removal performances up to 99%. Relatively lower salt rejection rates around 94% was obtained for RO system as a post treatment of MBR system.

  20. In vitro antioxidant properties of chicken skin enzymatic protein hydrolysates and membrane fractions.

    PubMed

    Onuh, John O; Girgih, Abraham T; Aluko, Rotimi E; Aliani, Michel

    2014-05-01

    Chicken thigh and breast skin proteins were hydrolysed using alcalase or a combination of pepsin and pancreatin (PP), each at concentrations of 1-4%. The chicken skin protein hydrolysates (CSPHs) were then fractionated by membrane ultrafiltration into different molecular weight peptides (<1, 1-3, 3-5 and 5-10kDa) and analysed for antioxidant properties. Results showed that the CSPHs had a significantly (p<0.05) lower scavenging activity against DPPH radicals when compared to reduced glutathione. The chicken breast skin hydrolysates had significantly higher DPPH scavenging activity than the chicken thigh skin hydrolysates. DPPH scavenging and metal ion chelation increased significantly (p<0.05) from 29-40% to 86-89%, respectively with increasing proteolytic enzyme concentration. In contrast, the antioxidant properties decreased as peptide size increased. We conclude that CSPHs and their peptide fractions may be used as ingredients in the formulation of functional foods and nutraceuticals for the control and management of oxidative stress-related diseases.

  1. Fuel cell membrane hydration and fluid metering

    DOEpatents

    Jones, Daniel O.; Walsh, Michael M.

    2003-01-01

    A hydration system includes fuel cell fluid flow plate(s) and injection port(s). Each plate has flow channel(s) with respective inlet(s) for receiving respective portion(s) of a given stream of reactant fluid for a fuel cell. Each injection port injects a portion of liquid water directly into its respective flow channel. This serves to hydrate at least corresponding part(s) of a given membrane of the corresponding fuel cell(s). The hydration system may be augmented by a metering system including flow regulator(s). Each flow regulator meters an injecting at inlet(s) of each plate of respective portions of liquid into respective portion(s) of a given stream of fluid by corresponding injection port(s).

  2. Fuel cell membrane hydration and fluid metering

    DOEpatents

    Jones, Daniel O.; Walsh, Michael M.

    1999-01-01

    A hydration system includes fuel cell fluid flow plate(s) and injection port(s). Each plate has flow channel(s) with respective inlet(s) for receiving respective portion(s) of a given stream of reactant fluid for a fuel cell. Each injection port injects a portion of liquid water directly into its respective flow channel in order to mix its respective portion of liquid water with the corresponding portion of the stream. This serves to hydrate at least corresponding part(s) of a given membrane of the corresponding fuel cell(s). The hydration system may be augmented by a metering system including flow regulator(s). Each flow regulator meters an injecting at inlet(s) of each plate of respective portions of liquid into respective portion(s) of a given stream of fluid by corresponding injection port(s).

  3. Tunable Microfluidic Devices for Hydrodynamic Fractionation of Cells and Beads: A Review

    PubMed Central

    Alvankarian, Jafar; Majlis, Burhanuddin Yeop

    2015-01-01

    The adjustable microfluidic devices that have been developed for hydrodynamic-based fractionation of beads and cells are important for fast performance tunability through interaction of mechanical properties of particles in fluid flow and mechanically flexible microstructures. In this review, the research works reported on fabrication and testing of the tunable elastomeric microfluidic devices for applications such as separation, filtration, isolation, and trapping of single or bulk of microbeads or cells are discussed. Such microfluidic systems for rapid performance alteration are classified in two groups of bulk deformation of microdevices using external mechanical forces, and local deformation of microstructures using flexible membrane by pneumatic pressure. The main advantage of membrane-based tunable systems has been addressed to be the high capability of integration with other microdevice components. The stretchable devices based on bulk deformation of microstructures have in common advantage of simplicity in design and fabrication process. PMID:26610519

  4. Membrane Purification Cell for Aluminum Recycling

    SciTech Connect

    David DeYoung; James Wiswall; Cong Wang

    2011-11-29

    Recycling mixed aluminum scrap usually requires adding primary aluminum to the scrap stream as a diluent to reduce the concentration of non-aluminum constituents used in aluminum alloys. Since primary aluminum production requires approximately 10 times more energy than melting scrap, the bulk of the energy and carbon dioxide emissions for recycling are associated with using primary aluminum as a diluent. Eliminating the need for using primary aluminum as a diluent would dramatically reduce energy requirements, decrease carbon dioxide emissions, and increase scrap utilization in recycling. Electrorefining can be used to extract pure aluminum from mixed scrap. Some example applications include producing primary grade aluminum from specific scrap streams such as consumer packaging and mixed alloy saw chips, and recycling multi-alloy products such as brazing sheet. Electrorefining can also be used to extract valuable alloying elements such as Li from Al-Li mixed scrap. This project was aimed at developing an electrorefining process for purifying aluminum to reduce energy consumption and emissions by 75% compared to conventional technology. An electrolytic molten aluminum purification process, utilizing a horizontal membrane cell anode, was designed, constructed, operated and validated. The electrorefining technology could also be used to produce ultra-high purity aluminum for advanced materials applications. The technical objectives for this project were to: - Validate the membrane cell concept with a lab-scale electrorefining cell; - Determine if previously identified voltage increase issue for chloride electrolytes holds for a fluoride-based electrolyte system; - Assess the probability that voltage change issues can be solved; and - Conduct a market and economic analysis to assess commercial feasibility. The process was tested using three different binary alloy compositions (Al-2.0 wt.% Cu, Al-4.7 wt.% Si, Al-0.6 wt.% Fe) and a brazing sheet scrap composition (Al-2

  5. Polymer synthesis toward fuel cell membrane materials

    NASA Astrophysics Data System (ADS)

    Rebeck, Nathaniel T.

    Fuel cells are a promising technology that will be part of the future energy landscape. New membranes for alkaline and proton exchange membrane fuel cells are needed to improve the performance, simplify the system, and reduce cost. Polymer chemistry can be applied to develop new polymers and to assemble polymers into improved membranes that need less water, have increased performance and are less expensive, thereby removing the deficiencies of current membranes. Nucleophilic aromatic substitution polymerization typically produces thermally stable engineering polymers that can be easily functionalized. New functional monomers were developed to explore new routes to novel functional polymers. Sulfonamides were discovered as new activating groups for polymerization of high molecular weight thermooxidatively stable materials with sulfonic acid latent functionality. While the sulfonamide functional polymers could be produced, the sulfonamide group proved to be too stable to convert into a sulfonic acid after reaction. The reactivity of 2-aminophenol was investigated to search for a new class of ion conducting polymer materials. Both the amine and the phenol groups are found to be reactive in a nucleophilic aromatic substitution, however not to the extent to allow the formation of high molecular weight polymer materials. Layer-by-layer films were assembled from aqueous solutions of poly(styrene sulfonate) and trimethylammonium functionalized poly(phenylene oxide). The deposition conditions were adjusted to increase the free charge carrier content, and chloride conductivites reached almost 30 mS/cm for the best films. Block and random poly(phenylene oxide) copolymers were produced from 2,6-dimethylphenol and 2,6-diphenylphenol and the methyl substituted repeat units were functionalized with trimethylammonium bromide. The block copolymers displayed bromide conductivities up to 26 mS/cm and outperformed the random copolymers, indicating that morphology has an effect on ion

  6. Probing red cell membrane cholesterol movement with cyclodextrin.

    PubMed

    Steck, Theodore L; Ye, Jin; Lange, Yvonne

    2002-10-01

    We probed the kinetics with which cholesterol moves across the human red cell bilayer and exits the membrane using methyl-beta-cyclodextrin as an acceptor. The fractional rate of cholesterol transfer (% s(-1)) was unprecedented, the half-time at 37 degrees C being ~1 s. The kinetics observed under typical conditions were independent of donor concentration and directly proportional to acceptor concentration. The rate of exit of membrane cholesterol fell hyperbolically to zero with increasing dilution. The energy of activation for cholesterol transfer was the same at high and low dilution; namely, 27-28 Kcal/mol. This behavior is not consistent with an exit pathway involving desorption followed by aqueous diffusion to acceptors nor with a simple one-step collision mechanism. Rather, it is that predicted for an activation-collision mechanism in which the reversible partial projection of cholesterol molecules out of the bilayer precedes their collisional capture by cyclodextrin. Because the entire membrane pool was transferred in a single first-order process under all conditions, we infer that the transbilayer diffusion (flip-flop) of cholesterol must have proceeded faster than its exit, i.e., with a half-time of <1 s at 37 degrees C.

  7. Recruitment of activating NK-cell receptors 2B4 and NKG2D to membrane microdomains in mammalian cells is dependent on their transmembrane regions.

    PubMed

    Gütgemann, Stephan A; Sandusky, Mina M; Wingert, Sabine; Claus, Maren; Watzl, Carsten

    2015-04-01

    Membrane microdomains play an important role in the regulation of natural killer (NK) cell activities. These cholesterol-rich membrane domains are enriched at the activating immunological synapse and several activating NK-cell receptors are known to localize to membrane microdomains upon receptor engagement. In contrast, inhibitory receptors do not localize in these specialized membrane domains. In addition, the functional competence of educated NK cells correlates with a confinement of activating receptors in membrane microdomains. However, the molecular basis for this confinement is unknown. Here, we investigate the structural requirements for the recruitment of the human-activating NK-cell receptors NKG2D and 2B4 to detergent-resistant membrane fractions in the murine BA/F3 cell line and in the human NK-cell line NKL. This stimulation-dependent recruitment occurred independently of the intracellular domains of the receptors. However, either interfering with the association between NKG2D and DAP10, or mutating the transmembrane region of 2B4 impacted the recruitment of the receptors to detergent-resistant membrane fractions and modulated the function of 2B4 in NK cells. Our data suggest a potential interaction between the transmembrane region of NK-cell receptors and membrane lipids as a molecular mechanism involved in determining the membrane confinement of activating NK-cell receptors.

  8. Microfabrication of High-Resolution Porous Membranes for Cell Culture

    PubMed Central

    Kim, Monica Y.; Li, David Jiang; Pham, Long K.; Wong, Brandon G.

    2014-01-01

    Microporous membranes are widely utilized in cell biology to study cell-cell signaling and cell migration. However, the thickness and low porosity of commercial track-etched membranes limit the quality of cell imaging and the degree of cell-cell contact that can be achieved on such devices. We employ photolithography-based microfabrication to achieve porous membranes with pore diameter as small as 0.9 μm, up to 40% porosity, and less than 5% variation in pore size. Through the use of a soap release layer, membranes as thin as 1 μm can be achieved. The thin membranes minimally disrupt contrast enhancement optics, thus allowing good quality imaging of unlabeled cells under white light, unlike commercial membranes. In addition, the polymer membrane materials display low autofluorescence even after patterning, facilitating high quality fluorescence microscopy. Finally, confocal imaging suggests that substantial cell-cell contact is possible through the pores of these thin membranes. This membrane technology can enhance existing uses of porous membranes in cell biology as well as enable new types of experiments. PMID:24567663

  9. Membrane tension feedback on shape and motility of eukaryotic cells

    NASA Astrophysics Data System (ADS)

    Winkler, Benjamin; Aranson, Igor S.; Ziebert, Falko

    2016-04-01

    In the framework of a phase field model of a single cell crawling on a substrate, we investigate how the properties of the cell membrane affect the shape and motility of the cell. Since the membrane influences the cell dynamics on multiple levels and provides a nontrivial feedback, we consider the following fundamental interactions: (i) the reduction of the actin polymerization rate by membrane tension; (ii) area conservation of the cell's two-dimensional cross-section vs. conservation of the circumference (i.e. membrane inextensibility); and (iii) the contribution from the membrane's bending energy to the shape and integrity of the cell. As in experiments, we investigate two pertinent observables - the cell's velocity and its aspect ratio. We find that the most important effect is the feedback of membrane tension on the actin polymerization. Bending rigidity has only minor effects, visible mostly in dynamic reshaping events, as exemplified by collisions of the cell with an obstacle.

  10. Protective effects of fractions from Artemisia biennis hydro-ethanolic extract against doxorubicin-induced oxidative stress and apoptosis in PC12 cells

    PubMed Central

    Mojarrab, Mahdi; Mehrabi, Mehran; Ahmadi, Farahnaz; Hosseinzadeh, Leila

    2016-01-01

    Objective(s): This study was designed to indicate whether different fractions from Artemisia biennis hydroethanolic extract could provide cytoprotection against oxidative stress and apoptosis induced by doxorubicin (DOX) in rat pheochromocytoma cell line (PC12). Material and Methods: Cell viability was determined by MTT assay. Also, activation of caspase-3 and superoxide dismutase were evaluated by spectrophotometry. Detection of reactive oxygen species (ROS) and measurement of mitochondrial membrane potential (MMP) were performed by flowcytometry. Results: Treatment of PC12 cells with DOX reduced viability dose dependently. For evaluation of the effect of fractions (A-G) on DOX-induced cytotoxicity, PC12 cells were pretreated for 24 hr with the A. biennis fractions and then cells were treated with DOX. The fractions C and D increased PC12 cells viability significantly compared to DOX treated cells. Moreover, pretreatment with fractions C and D for 24 hr attenuated DOX-mediated apoptosis and the anti-apoptotic action of A. biennis fractions was partially dependent on inhibition of caspase 3 activity and also increasing the mitochondrial membrane potential (MMP). Selected A. biennis fractions also suppressed the generation of ROS and increased superoxide dismutase (SOD) activity. Conclusion: Taken together our observation indicated that subtoxic concentration of aforementioned fractions of A. biennis hydroetanolic extract has protective effect against apoptosis induced by DOX in PC12 cell. The results highlighted that fractions C and D may exert cytoprotective effects through their antioxidant actions. PMID:27403257

  11. Polybenzimidazole-multiwall carbon nanotubes composite membranes for polymer electrolyte membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Guerrero Moreno, Nayibe; Gervasio, Dominic; Godínez García, Andrés; Pérez Robles, Juan Francisco

    2015-12-01

    Polymer membranes are prepared as a composite of polybenzimidazole and non-functionalized multiwall carbon nanotubes (PBI-CNT) and polybenzimidazole (PBI) only. Each is doped with H3PO4 (PA) and used as a proton exchange membrane (PEM) as the electrolyte in a fuel cell. The proton conductivities at 180 °C for the doped PBI membrane (PBIPA) and the doped PBI-CNT membrane (PBICNTPA) are 6.3 × 10-2 and 7.4 × 10-2 Scm-1 respectively. A single fuel cell having these membranes as electrolyte has a Pt catalyzed hydrogen gas fed anode and a similar oxygen cathode without humidification of feed gases; the cell with the PBICNTPA membrane has higher open circuit voltage (0.96 V) than that with a PBIPA membrane (0.8 V) at 180 °C. The mechanical stability of the membrane improves with CNTs addition. The tensile strength of the composite PBI-CNT membrane with 1 wt.% CNTs loading is 32% higher and the Young's Modulus is 147% higher than the values for a membrane of PBI alone. The improvement in conductivity and mechanical properties in the composite membrane due to the CNT addition indicates that a PBI-CNT membrane is a good alternative as a membrane electrolyte in a PEMFC.

  12. Characterization of antibody binding to cell surface antigens using a plasma membrane-bound plate assay.

    PubMed

    Vater, C A; Reid, K; Bartle, L M; Goldmacher, V S

    1995-01-01

    A procedure has been developed for measuring antibody binding to cell surface antigens using an immobilized plasma membrane fraction. In this method, isolated plasma membranes are dried onto wells of a 96-well microtiter plate and incubated with antibodies that recognize a cell surface protein. Bound antibody is detected indirectly using an enzyme-linked or fluorescently tagged second antibody. Alternatively, the primary antibody itself can be labeled and its binding can be detected directly. The assay is simple and fast and provides several advantages over whole cell binding assays currently in widespread use.

  13. Isolation and Characterization of Glycophorin from Carp Red Blood Cell Membranes

    PubMed Central

    Aoki, Takahiko; Chimura, Kenji; Nakao, Nobuhiro; Mizuno, Yasuko

    2014-01-01

    We isolated a high-purity carp glycophorin from carp erythrocyte membranes following extraction using the lithium diiodosalicylate (LIS)-phenol method and streptomycin treatment. The main carp glycophorin was observed to locate at the position of the carp and human band-3 proteins on an SDS-polyacrylamide gel. Only the N-glycolylneuraminic acid (NeuGc) form of sialic acid was detected in the carp glycophorin. The oligosaccharide fraction was separated into two components (P-1 and P-2) using a Glyco-Pak DEAE column. We observed bacteriostatic activity against five strains of bacteria, including two known fish pathogens. Fractions from the carp erythrocyte membrane, the glycophorin oligosaccharide and the P-1 also exhibited bacteriostatic activity; whereas the glycolipid fraction and the glycophorin fraction without sialic acid did not show the activity. The carp glycophorin molecules attach to the flagellum of V. anguillarum or the cell surface of M. luteus and inhibited bacterial growth. PMID:25110961

  14. A simple method for discriminating between cell membrane and cytosolic proteins.

    PubMed

    Serna, Laura

    2005-03-01

    * Transgenic plants expressing either green fluorescent protein (GFP)-genomic DNA or GFP-cDNA fusions have been used as powerful tools to define the subcellular localization of many proteins. Because most plant cells are highly vacuolated, the cytosol is confined to a thin layer at the periphery of the cells, making it very difficult to distinguish among cell wall, cell membrane and cytosolic GFP-fusion proteins. * Plasmolysis tests inform about cell-wall localization of GFP-tagged proteins, but they do not discriminate between its cell membrane and/or cytoplasmic localization. By observing the GFP signal in transgenic protoplasts placed at a hypotonic solution, it was possible to distinguish between cell membrane and cytosolic GFP-tagged proteins. * The osmotic disruption of the protoplast vacuole in the hypotonic solution allows the diffusion of the GFP signal from the cell periphery to the central part of the cell volume when the GFP is fused to a soluble protein. By contrast, such diffusion does not occur when the protein under study is attached to the cell membrane. * The present method is easier, faster and cheaper than subcellular fractionating studies and/or immunoelectron microscopy, which have been traditionally used to discern between cell membrane and cytosolic proteins.

  15. Membrane-Filtered Olive Mill Wastewater: Quality Assessment of the Dried Phenolic-Rich Fraction.

    PubMed

    Sedej, Ivana; Milczarek, Rebecca; Wang, Selina C; Sheng, Runqi; de Jesús Avena-Bustillos, Roberto; Dao, Lan; Takeoka, Gary

    2016-04-01

    A current trend in olive mill wastewater (OMWW) management is to not only decrease environmental pollution but also to extract and utilize valuable by-products. Therefore, the objectives of this study were to explore different techniques for drying a phenolic-rich membrane filtration fraction of OMWW and compare the techniques in terms of the dried product quality and feasibility of the process. The OMWW from 2 (3-phase and 2-phase) California mills was subjected to a 2-step membrane filtration process using a novel vibratory system. The reverse osmosis retentate (RO-R) is a phenolic-rich coproduct stream, and the reverse osmosis permeate is a near-pure water stream that could be recycled into the milling process. Spray-, freeze-, and infrared-drying were applied to obtain solid material from the RO-R. Drying of the RO-R was made possible only with addition of 10% maltodextrin as a carrier. The total soluble phenolics in dried RO-R were in the range 0.15 to 0.58 mg gallic acid equivalents/g of dry weight for 2-phase RO-R, and 1.38 to 2.17 mg gallic acid equivalents/g of dry weight for the 3-phase RO-R. Spray-dried RO-R from 3-phase OMWW showed remarkable antioxidant activity. Protocatechuic acid, tyrosol, vanillic acid, and p-coumaric acid were quantified in all dried RO-R, whereas 3-hydroxytyrosol was found in 3-phase dried RO-R. This combination of separation and drying technologies helps to add value and shelf-stability to an olive oil by-product and increase environmental sustainability of its production.

  16. Definition of glomerular antigens by monoclonal antibodies produced against a human glomerular membrane fraction.

    PubMed

    Neale, T J; Callus, M S; Donovan, L C; Baird, H

    1990-10-01

    Experimental animal models of glomerulonephritis (GN) produced by direct antibody binding to non-basement membrane glomerular capillary wall antigens do not to date have human parallels. To examine the potential for this form of humoral glomerular injury in man, we sought to define discrete human non-GBM glomerular antigenic targets using hybridoma technology. Mice were immunised intraperitoneally with 20-100 micrograms of a human glomerular membrane fraction (HGMF). Six fusions have yielded 12 stable reagents defined by positive glomerular indirect immunofluorescence (IF) and microELISA using HGMF as the screening antigen. Subclass analysis of ascitic McAbs indicated several IgG1, one IgG2b, and three IgM reagents. Distinctive IF patterns of reactivity with epithelial, endothelial or mesangial structures have been observed, with or without peritubular capillary, tubular basement membrane and vessel wall reactivity. Seven normal non-renal human organs and the kidneys of rat, rabbit and sheep have shown patterns characteristic of each individual McAb, restricted to human or with species cross reactivity. To partially characterise McAb-reactive antigens, detergent-solubilised renal cortex and collagenase-solubilised GBM (CS-GBM) extracts have been probed by immunoblot. A unique McAb 7-5Q, reactive with glomerular and tubular epithelial structures, binds major bands of approximately 107 KD and 93 KD in detergent solubilised cortex and a single band of similar size by immunoprecipitation (110 KD). 5-3A (a human-restricted linear-reacting McAb) binds bands of 20-200 KD (major band 58 KD) in CS-GBM. In conclusion, distinct species-restricted and more broadly disposed glomerular epitopes are definable in man by McAbs and are potential targets for humoral injury. Purification of these antigens will allow assay for circulating putative nephritogenic auto-antibody and potentially, McAbs may be useful in screening urine for evidence of occult structural renal disease.

  17. Fractionation of the Hypericum perforatum L. extract: PMF, and PDT effects of the fractions against HL-60 leukemic cells

    NASA Astrophysics Data System (ADS)

    Tsontou, M.; Dimitriou, H.; Filippidis, G.; Tsimaris, I.; Kalmanti, M.; Skalkos, D.

    2007-02-01

    In the last three years we have prepared and studied the polar methanolic extract PMF, of the herb Hypericum perforatum L, and studied as a new, alternative photosensitizing substance for PDT. Hypericum perforatum L., as well as PMF, contains a number of naphthodianthrone derivatives (hypericins), such as hypericin and pseudohypericin, as its main photosensitizing constituents. PMF has been tested as a PDT agent in vitro in bladder cancer cells, leukemia cells, and in vivo in rat tumor bearing urinary bladder. In order to evaluate the contribution of the hypericins in the overall PDT action, and prepare a better photosensitizing extract than PMF, we have separated the extract in four main fractions (1,2,3,4), and tested their PDT effects against the HL-60 leukemic cells. The concentration of hypericins in the extracts was found 0.08% for fraction 1, 0.09% for fraction 2, 0.8% for fraction 3, and 2,8% for fraction 4. The PDT activity observed among the fractions was proportional to their hypericins concentration, thus increasing in the order of increasing number: fraction 4 > fraction 3 > fraction 2 > fraction 1. Fraction 4 proved to be the most powerful fraction. However, despite its relatively high hypericins concentration (2.8%), compared with the total extract PMF (1.37%), fraction 4 proved to be less active in the cell line tested. This result indicates that there are other photosensitizing constituents within the PMF extract which contribute significantly in the overall PDT action, and therefore the extract should be used as it is for further PDT studies, without any further purification.

  18. Intravacuolar Membranes Regulate CD8 T Cell Recognition of Membrane-Bound Toxoplasma gondii Protective Antigen.

    PubMed

    Lopez, Jodie; Bittame, Amina; Massera, Céline; Vasseur, Virginie; Effantin, Grégory; Valat, Anne; Buaillon, Célia; Allart, Sophie; Fox, Barbara A; Rommereim, Leah M; Bzik, David J; Schoehn, Guy; Weissenhorn, Winfried; Dubremetz, Jean-François; Gagnon, Jean; Mercier, Corinne; Cesbron-Delauw, Marie-France; Blanchard, Nicolas

    2015-12-15

    Apicomplexa parasites such as Toxoplasma gondii target effectors to and across the boundary of their parasitophorous vacuole (PV), resulting in host cell subversion and potential presentation by MHC class I molecules for CD8 T cell recognition. The host-parasite interface comprises the PV limiting membrane and a highly curved, membranous intravacuolar network (IVN) of uncertain function. Here, using a cell-free minimal system, we dissect how membrane tubules are shaped by the parasite effectors GRA2 and GRA6. We show that membrane association regulates access of the GRA6 protective antigen to the MHC I pathway in infected cells. Although insertion of GRA6 in the PV membrane is key for immunogenicity, association of GRA6 with the IVN limits presentation and curtails GRA6-specific CD8 responses in mice. Thus, membrane deformations of the PV regulate access of antigens to the MHC class I pathway, and the IVN may play a role in immune modulation.

  19. The three dimensionality of cell membranes: lamellar to cubic membrane transition as investigated by electron microscopy.

    PubMed

    Chong, Ketpin; Deng, Yuru

    2012-01-01

    Biological membranes are generally perceived as phospholipid bilayer structures that delineate in a lamellar form the cell surface and intracellular organelles. However, much more complex and highly convoluted membrane organizations are ubiquitously present in many cell types under certain types of stress, states of disease, or in the course of viral infections. Their occurrence under pathological conditions make such three-dimensionally (3D) folded and highly ordered membranes attractive biomarkers. They have also stimulated great biomedical interest in understanding the molecular basis of their formation. Currently, the analysis of such membrane arrangements, which include tubulo-reticular structures (TRS) or cubic membranes of various subtypes, is restricted to electron microscopic methods, including tomography. Preservation of membrane structures during sample preparation is the key to understand their true 3D nature. This chapter discusses methods for appropriate sample preparations to successfully examine and analyze well-preserved highly ordered membranes by electron microscopy. Processing methods and analysis conditions for green algae (Zygnema sp.) and amoeba (Chaos carolinense), mammalian cells in culture and primary tissue cells are described. We also discuss methods to identify cubic membranes by transmission electron microscopy (TEM) with the aid of a direct template matching method and by computer simulation. A 3D analysis of cubic cell membrane topology by electron tomography is described as well as scanning electron microscopy (SEM) to investigate surface contours of isolated mitochondria with cubic membrane arrangement.

  20. Houttuynia cordata Thunb fraction induces human leukemic Molt-4 cell apoptosis through the endoplasmic reticulum stress pathway.

    PubMed

    Prommaban, Adchara; Kodchakorn, Kanchanok; Kongtawelert, Prachya; Banjerdpongchai, Ratana

    2012-01-01

    Houttuynia cordata Thunb (HCT) is a native herb found in Southeast Asia which features various pharmacological activities against allergy, inflammation, viral and bacterial infection, and cancer. The aims of this study were to determine the cytotoxic effect of 6 fractions obtained from silica gel column chromatography of alcoholic HCT extract on human leukemic Molt-4 cells and demonstrate mechanisms of cell death. Six HCT fractions were cytotoxic to human lymphoblastic leukemic Molt-4 cells in a dose-dependent manner by MTT assay, fraction 4 exerting the greatest effects. Treatment with IC50 of HCT fraction 4 significantly induced Molt-4 apoptosis detected by annexinV-FITC/propidium iodide for externalization of phosphatidylserine to the outer layer of cell membrane. The mitochondrial transmembrane potential was reduced in HCT fraction 4-treated Molt-4 cells. Moreover, decreased expression of Bcl-xl and increased levels of Smac/Diablo, Bax and GRP78 proteins were noted on immunoblotting. In conclusion, HCT fraction 4 induces Molt-4 apoptosis cell through an endoplasmic reticulum stress pathway.

  1. Synthesis of Nanogels via Cell Membrane-Templated Polymerization

    PubMed Central

    Zhang, Jianhua; Gao, Weiwei; Fang, Ronnie H.; Dong, Anjie

    2015-01-01

    The synthesis of biomimetic hydrogel nanoparticles coated with natural cell membrane is described. Compared to existing strategy of wrapping cell membrane onto pre-formed nanoparticle substrates, this new approach forms the cell membrane-derived vesicles first, followed by growing nanoparticle cores in situ. It adds significant controllability over the nanoparticle properties and opens unique opportunities for a broad range of biomedical applications. PMID:26044721

  2. Cell Adhesion and Growth on the Anodized Aluminum Oxide Membrane.

    PubMed

    Park, Jeong Su; Moon, Dalnim; Kim, Jin-Seok; Lee, Jin Seok

    2016-03-01

    Nanotopological cues are popular tools for in vivo investigation of the extracellular matrix (ECM) and cellular microenvironments. The ECM is composed of multiple components and generates a complex microenvironment. The development of accurate in vivo methods for the investigation of ECM are important for disease diagnosis and therapy, as well as for studies on cell behavior. Here, we fabricated anodized aluminum oxide (AAO) membranes using sulfuric and oxalic acid under controlled voltage and temperature. The membranes were designed to possess three different pore and interpore sizes, AAO-1, AAO-2, and AAO-3 membranes, respectively. These membranes were used as tools to investigate nanotopology-signal induced cell behavior. Cancerous cells, specifically, the OVCAR-8 cell-line, were cultured on porous AAO membranes and the effects of these membranes on cell shape, proliferation, and viability were studied. AAO-1 membranes bearing small sized pores were found to maintain the spreading shape of the cultured cells. Cells cultured on AAO-2 and AAO-3 membranes, bearing large pore-sized AAO membranes, changed shape from spreading to rounding. Furthermore, cellular area decreased when cells were cultured on all three AAO membranes that confirmed decreased levels of focal adhesion kinase (FAK). Additionally, OVCAR-8 cells exhibited increased proliferation on AAO membranes possessing various pore sizes, indicating the importance of the nanosurface structure in regulating cell behaviors, such as cell proliferation. Our results suggest that porous-AAO membranes induced nanosurface regulated cell behavior as focal adhesion altered the intracellular organization of the cytoskeleton. Our results may find potential applications as tools in in vivo cancer research studies.

  3. Cell membrane modulation as adjuvant in cancer therapy.

    PubMed

    Zalba, Sara; Ten Hagen, Timo L M

    2017-01-01

    Cancer is a complex disease involving numerous biological processes, which can exist in parallel, can be complementary, or are engaged when needed and as such can replace each other. This redundancy in possibilities cancer cells have, are fundamental to failure of therapy. However, intrinsic features of tumor cells and tumors as a whole provide also opportunities for therapy. Here we discuss the unique and specific makeup and arrangement of cell membranes of tumor cells and how these may help treatment. Interestingly, knowledge on cell membranes and associated structures is present already for decades, while application of membrane modification and manipulation as part of cancer therapy is lagging. Recent developments of scientific tools concerning lipids and lipid metabolism, opened new and previously unknown aspects of tumor cells and indicate possible differences in lipid composition and membrane function of tumor cells compared to healthy cells. This field, coined Lipidomics, demonstrates the importance of lipid components in cell membrane in several illnesses. Important alterations in cancer, and specially in resistant cancer cells compared to normal cells, opened the door to new therapeutic strategies. Moreover, the ability to modulate membrane components and/or properties has become a reality. Here, developments in cancer-related Lipidomics and strategies to interfere specifically with cancer cell membranes and how these affect cancer treatment are discussed. We hypothesize that combination of lipid or membrane targeted strategies with available care to improve chemotherapy, radiotherapy and immunotherapy will bring the much needed change in treatment in the years to come.

  4. Hydrocarbon-based polymer electrolyte cerium composite membranes for improved proton exchange membrane fuel cell durability

    NASA Astrophysics Data System (ADS)

    Lee, Hyejin; Han, Myungseong; Choi, Young-Woo; Bae, Byungchan

    2015-11-01

    Hydrocarbon-based cerium composite membranes were prepared for proton exchange membrane fuel cell applications to increase oxidative stability. Different amounts of cerium ions were impregnated in sulfonated poly(arylene ether sulfone) (SPES) membranes and their physicochemical properties were investigated according to the cerium content. Field-emission scanning electron microscopy and inductively coupled plasma analyses confirmed the presence of cerium ions in the composite membranes and 1H NMR indicated the successful coordination of sulfonic acid groups with the metal ions. Increasing amounts of cerium ions resulted in decreases in the proton conductivity and water uptake, but enhanced oxidative stability. The oxidative stability of the composite membranes was proven via a hydrogen peroxide exposure experiment which mimicked fuel cell operating conditions. In addition, more than 2200 h was achieved with the composite membrane under in situ accelerated open circuit voltage (OCV) durability testing (DOE protocol), whereas the corresponding pristine SPES membrane attained only 670 h.

  5. Impedance study of membrane dehydration and compression in proton exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Le Canut, Jean-Marc; Latham, Ruth; Mérida, Walter; Harrington, David A.

    Electrochemical impedance spectroscopy (EIS) is used to measure drying and rehydration in proton exchange membrane fuel cells running under load. The hysteresis between forward and backward acquisition of polarization curves is shown to be largely due to changes in the membrane resistance. Drying tests are carried out with hydrogen and simulated reformate (hydrogen and carbon dioxide), and quasi-periodic drying and rehydration conditions are studied. The membrane hydration state is clearly linked to the high-frequency arc in the impedance spectrum, which increases in size for dry conditions indicating an increase in membrane resistance. Changes in impedance spectra as external compression is applied to the cell assembly show that EIS can separate membrane and interfacial effects, and that changes in membrane resistance dominate. Reasons for the presence of a capacitance in parallel with the membrane resistance are discussed.

  6. Development of composite membranes of PVA-TEOS doped KOH for alkaline membrane fuel cell

    SciTech Connect

    Haryadi, Sugianto, D.; Ristopan, E.

    2015-12-29

    Anion exchange membranes (AEMs) play an important role in separating fuel and oxygen (or air) in the Alkaline Membrane Fuel Cells. Preparation of hybrid organic inorganic materials of Polyvinylalcohol (PVA) - Tetraethylorthosilicate (TEOS) composite membrane doped KOH for direct alcohol alkaline fuel cell application has been investigated. The sol-gel method has been used to prepare the composite membrane of PVA-TEOS through crosslinking step and catalyzed by concentrated of hydrochloric acid. The gel solution was cast on the membrane plastic plate to obtain membrane sheets. The dry membranes were then doped by immersing in various concentrations of KOH solutions for about 4 hours. Investigations of the cross-linking process and the presence of hydroxyl group were conducted by FTIR as shown for frequency at about 1600 cm{sup −1} and 3300 cm{sup −1} respectively. The degree of swelling in ethanol decreased as the KOH concentration for membrane soaking process increased. The ion exchange capacity (IEC) of the membrane was 0.25meq/g. This composite membranes display significant ionic conductivity of 3.23 x 10{sup −2} S/cm in deionized water at room temperature. In addition, the morphology observation by scanning electron microscope (SEM) of the membrane indicates that soaking process of membrane in KOH increased thermal resistant.

  7. Development of composite membranes of PVA-TEOS doped KOH for alkaline membrane fuel cell

    NASA Astrophysics Data System (ADS)

    Haryadi, Sugianto, D.; Ristopan, E.

    2015-12-01

    Anion exchange membranes (AEMs) play an important role in separating fuel and oxygen (or air) in the Alkaline Membrane Fuel Cells. Preparation of hybrid organic inorganic materials of Polyvinylalcohol (PVA) - Tetraethylorthosilicate (TEOS) composite membrane doped KOH for direct alcohol alkaline fuel cell application has been investigated. The sol-gel method has been used to prepare the composite membrane of PVA-TEOS through crosslinking step and catalyzed by concentrated of hydrochloric acid. The gel solution was cast on the membrane plastic plate to obtain membrane sheets. The dry membranes were then doped by immersing in various concentrations of KOH solutions for about 4 hours. Investigations of the cross-linking process and the presence of hydroxyl group were conducted by FTIR as shown for frequency at about 1600 cm-1 and 3300 cm-1 respectively. The degree of swelling in ethanol decreased as the KOH concentration for membrane soaking process increased. The ion exchange capacity (IEC) of the membrane was 0.25meq/g. This composite membranes display significant ionic conductivity of 3.23 x 10-2 S/cm in deionized water at room temperature. In addition, the morphology observation by scanning electron microscope (SEM) of the membrane indicates that soaking process of membrane in KOH increased thermal resistant.

  8. Xyloglucan biosynthesis by Golgi membranes from suspension-cultured sycamore (Acer pseudoplatanus) cells

    SciTech Connect

    White, A.R.; Xin, Yi )

    1990-05-01

    Xyloglucan is a major hemicellulose polysaccharide in plant cell walls. Biosynthesis of such cell wall polysaccharides is closely linked to the process of plant cell growth and development. Xyloglucan polysaccharides consist of a {beta}-1,4 glucan backbone synthesized by xyloglucan synthase and sidechains of xylose, galactose, and fucose added by other transferase enzymes. Most plant Golgi and plasma membranes also contain glucan synthases I II, which make {beta}-1,4 and {beta}-1,3 glucans, respectively. All of these enzymes have very similar activities. Cell walls on suspension-cultured cells from Acer pseudoplatanus (sycamore maple) were enzymatically softened prior to cell disruption by passing through a 30 {mu}m nylon screen. Cell membranes from homogenates were separated by ultracentrifugation on top-loaded or flotation sucrose density gradients. Samples were collected by gradient fractionation and assayed for membrane markers and xyloglucan and glucan synthase activities. Standard marker assays (cyt. c reductase for eR, IDPase UDPase for Golgi, and eosin 5{prime}-malelmide binding for plasma membrane) showed partial separation of these three membrane types. Golgi and plasma membrane markers overlapped in most gradients. Incorporation of {sup 14}C-labeled sugars from UDP-glucose and UDP-xylose was used to detect xyloglucan synthase, glucan synthases I II, and xylosyl transferase in Golgi membrane fractions. These activities overlapped, although distinct peaks of xyloglucan synthase and xylosyl transferase were found. Ca{sup ++} had a stimulatory effect on glucan synthases I II, while Mn{sup ++} had an inhibitory effect on glucan synthase I in the presence of Ca{sup ++}. The similarity of these various synthase activities demonstrates the need for careful structural characterization of newly synthesized polysaccharides.

  9. Variety of RNAs in Peripheral Blood Cells, Plasma, and Plasma Fractions.

    PubMed

    Savelyeva, Anna V; Kuligina, Elena V; Bariakin, Dmitry N; Kozlov, Vadim V; Ryabchikova, Elena I; Richter, Vladimir A; Semenov, Dmitry V

    2017-01-01

    Human peripheral blood contains RNA in cells and in extracellular membrane vesicles, microvesicles and exosomes, as well as in cell-free ribonucleoproteins. Circulating mRNAs and noncoding RNAs, being internalized, possess the ability to modulate vital processes in recipient cells. In this study, with SOLiD sequencing technology, we performed identification, classification, and quantification of RNAs from blood fractions: cells, plasma, plasma vesicles pelleted at 16,000g and 160,000g, and vesicle-depleted plasma supernatant of healthy donors and non-small cell lung cancer (NSCLC) patients. It was determined that 16,000g blood plasma vesicles were enriched with cell-free mitochondria and with a set of mitochondrial RNAs. The variable RNA set of blood plasma 160,000g pellets reflected the prominent contribution of U1, U5, and U6 small nuclear RNAs' fragments and at the same time was characterized by a remarkable depletion of small nucleolar RNAs. Besides microRNAs, the variety of fragments of mRNAs and snoRNAs dominated in the set of circulating RNAs differentially expressed in blood fractions of NSCLC patients. Taken together, our data emphasize that not only extracellular microRNAs but also circulating fragments of messenger and small nuclear/nucleolar RNAs represent prominent classes of circulating regulatory ncRNAs as well as promising circulating biomarkers for the development of disease diagnostic approaches.

  10. Variety of RNAs in Peripheral Blood Cells, Plasma, and Plasma Fractions

    PubMed Central

    Kuligina, Elena V.; Bariakin, Dmitry N.; Kozlov, Vadim V.; Richter, Vladimir A.; Semenov, Dmitry V.

    2017-01-01

    Human peripheral blood contains RNA in cells and in extracellular membrane vesicles, microvesicles and exosomes, as well as in cell-free ribonucleoproteins. Circulating mRNAs and noncoding RNAs, being internalized, possess the ability to modulate vital processes in recipient cells. In this study, with SOLiD sequencing technology, we performed identification, classification, and quantification of RNAs from blood fractions: cells, plasma, plasma vesicles pelleted at 16,000g and 160,000g, and vesicle-depleted plasma supernatant of healthy donors and non-small cell lung cancer (NSCLC) patients. It was determined that 16,000g blood plasma vesicles were enriched with cell-free mitochondria and with a set of mitochondrial RNAs. The variable RNA set of blood plasma 160,000g pellets reflected the prominent contribution of U1, U5, and U6 small nuclear RNAs' fragments and at the same time was characterized by a remarkable depletion of small nucleolar RNAs. Besides microRNAs, the variety of fragments of mRNAs and snoRNAs dominated in the set of circulating RNAs differentially expressed in blood fractions of NSCLC patients. Taken together, our data emphasize that not only extracellular microRNAs but also circulating fragments of messenger and small nuclear/nucleolar RNAs represent prominent classes of circulating regulatory ncRNAs as well as promising circulating biomarkers for the development of disease diagnostic approaches. PMID:28127559

  11. Nonhumidified High-Temperature Membranes Developed for Proton Exchange Membrane Fuel Cells

    NASA Technical Reports Server (NTRS)

    Kinder, James D.

    2005-01-01

    Fuel cells are being considered for a wide variety of aerospace applications. One of the most versatile types of fuel cells is the proton-exchange-membrane (PEM) fuel cell. PEM fuel cells can be easily scaled to meet the power and space requirements of a specific application. For example, small 100-W PEM fuel cells are being considered for personal power for extravehicular activity suit applications, whereas larger PEM fuel cells are being designed for primary power in airplanes and in uninhabited air vehicles. Typically, PEM fuel cells operate at temperatures up to 80 C. To increase the efficiency and power density of the fuel cell system, researchers are pursuing methods to extend the operating temperature of the PEM fuel cell to 180 C. The most widely used membranes in PEM fuel cells are Nafion 112 and Nafion 117--sulfonated perfluorinated polyethers that were developed by DuPont. In addition to their relatively high cost, the properties of these membranes limit their use in a PEM fuel cell to around 80 C. The proton conductivity of Nafion membranes significantly decreases above 80 C because the membrane dehydrates. The useful operating range of Nafion-based PEM fuel cells can be extended to over 100 C if ancillary equipment, such as compressors and humidifiers, is added to maintain moisture levels within the membrane. However, the addition of these components reduces the power density and increases the complexity of the fuel cell system.

  12. DYNAMICS OF ANTIGENIC MEMBRANE SITES RELATING TO CELL AGGREGATION IN DICTYOSTELIUM DISCOIDEUM

    PubMed Central

    Beug, H.; Katz, F. E.; Gerisch, G.

    1973-01-01

    Membrane interaction in aggregating cells of Dictyostelium discoideum can be blocked by univalent antibodies directed against specific membrane sites. Using a quantitative technique for measuring cell association, two classes of target sites for blocking antibodies were distinguished and their developmental dynamics studied. One class of these sites is specific for aggregation-competent cells, their quantity rising from virtually 0-level during growth, with a steep increase shortly before cell aggregation. The serological activity of these structures is species specific; they are not detectable in a nonaggregating mutant, but present in a revertant undergoing normal morphogenesis. Patterns of cell assembly in the presence of antibodies show that selective blockage of these membrane sites abolishes the preference for end-to-end association which is typical for aggregating cells. A second class of target sites is present in comparable quantities in particle fractions from both growth-phase and aggregation-competent cells. Blockage of these sites leads to aggregation patterns in which the side-by-side contacts of aggregating cells are abolished. The target sites of aggregation-inhibiting antibodies are suggested to be identical or associated with the molecular units of the cell membrane that mediate cell-to-cell contacts during aggregation. The results indicate that in one cell, two independent classes of contact sites can be simultaneously active. PMID:4631665

  13. Graphene-Induced Pore Formation on Cell Membranes

    NASA Astrophysics Data System (ADS)

    Duan, Guangxin; Zhang, Yuanzhao; Luan, Binquan; Weber, Jeffrey K.; Zhou, Royce W.; Yang, Zaixing; Zhao, Lin; Xu, Jiaying; Luo, Judong; Zhou, Ruhong

    2017-02-01

    Examining interactions between nanomaterials and cell membranes can expose underlying mechanisms of nanomaterial cytotoxicity and guide the design of safer nanomedical technologies. Recently, graphene has been shown to exhibit potential toxicity to cells; however, the molecular processes driving its lethal properties have yet to be fully characterized. We here demonstrate that graphene nanosheets (both pristine and oxidized) can produce holes (pores) in the membranes of A549 and Raw264.7 cells, substantially reducing cell viability. Electron micrographs offer clear evidence of pores created on cell membranes. Our molecular dynamics simulations reveal that multiple graphene nanosheets can cooperate to extract large numbers of phospholipids from the membrane bilayer. Strong dispersion interactions between graphene and lipid-tail carbons result in greatly depleted lipid density within confined regions of the membrane, ultimately leading to the formation of water-permeable pores. This cooperative lipid extraction mechanism for membrane perforation represents another distinct process that contributes to the molecular basis of graphene cytotoxicity.

  14. Selective effect of cell membrane on synaptic neurotransmission

    NASA Astrophysics Data System (ADS)

    Postila, Pekka A.; Vattulainen, Ilpo; Róg, Tomasz

    2016-01-01

    Atomistic molecular dynamics simulations were performed with 13 non-peptidic neurotransmitters (NTs) in three different membrane environments. The results provide compelling evidence that NTs are divided into membrane-binding and membrane-nonbinding molecules. NTs adhere to the postsynaptic membrane surface whenever the ligand-binding sites of their synaptic receptors are buried in the lipid bilayer. In contrast, NTs that have extracellular ligand-binding sites do not have a similar tendency to adhere to the membrane surface. This finding is a seemingly simple yet important addition to the paradigm of neurotransmission, essentially dividing it into membrane-independent and membrane-dependent mechanisms. Moreover, the simulations also indicate that the lipid composition especially in terms of charged lipids can affect the membrane partitioning of NTs. The revised paradigm, highlighting the importance of cell membrane and specific lipids for neurotransmission, should to be of interest to neuroscientists, drug industry and the general public alike.

  15. The application of Dow Chemical's perfluorinated membranes in proton-exchange membrane fuel cells

    NASA Technical Reports Server (NTRS)

    Eisman, G. A.

    1989-01-01

    Dow Chemical's research activities in fuel cells revolve around the development of perfluorosulfonic acid membranes useful as the proton transport medium and separator. Some of the performance characteristics which are typical for such membranes are outlined. The results of tests utilizing a new experimental membrane useful in proton-exchange membrane fuel cells are presented. The high voltage at low current densities can lead to higher system efficiencies while, at the same time, not sacrificing other critical properties pertinent to membrane fuel cell operation. A series of tests to determine response times indicated that on-off cycles are on the order of 80 milliseconds to reach 90 percent of full power. The IR free voltage at 100 amps/sq ft was determined and the results indicating a membrane/electrode package resistance to be .15 ohm-sq cm at 100 amps/sq ft.

  16. New application of a subcellular fractionation method to kidney and testis for the determination of conjugated linoleic acid in selected cell organelles of healthy and cancerous human tissues.

    PubMed

    Hoffmann, Kristina; Blaudszun, Jörg; Brunken, Claus; Höpker, Wilhelm-Wolfgang; Tauber, Roland; Steinhart, Hans

    2005-03-01

    To clarify the mechanism of the anticarcinogenic effect of conjugated linoleic acid (CLA), its intracellular distribution needs to be determined. Subcellular fractionation using centrifugation techniques is a method that is frequently used for isolation of cell organelles from different tissues. But as the size and density of the organelles differ, the method needs to be optimised for every type of tissue. The novelty of this study is the application of a subcellular fractionation method to human healthy and cancerous renal and testicular tissue. Separation of total tissue homogenate into nuclei, cytosol, and a mixture of mitochondria and plasma membranes was achieved by differential centrifugation. As mitochondria and plasma membranes seemed to be too similar in size and weight to be separated by differential centrifugation, discontinuous density-gradient centrifugation was carried out successfully. The purity of the subcellular fractions was checked by measuring the activity of marker enzymes. All fractions were highly enriched in their corresponding marker enzyme. However, the nuclear fractions of kidney and renal cell carcinoma were slightly contaminated with mitochondria and plasma membrane fractions of all tissues with lysosomes. The fraction designated the cytosolic fraction contained not only cytosol, but also microsomes and lysosomes. The CLA contents of the subcellular fractions were in the range 0.13-0.37% of total fatty acids and were lowest in the plasma membrane fractions of all types of tissue studied. C16:0, C18:0, C18:1 c9, C18:2 n-6, and C20:4 n-6 were found to be the major fatty acids in all the subcellular fractions studied. However, marked variations in fatty acid content between subcellular fractions and between types of tissue were detectable. Because of these differences between tissues, no general statement on characteristic fatty acid profiles of single subcellular fractions is possible.

  17. Valorisation of tuna processing waste biomass for recovery of functional and antioxidant peptides using enzymatic hydrolysis and membrane fractionation process.

    PubMed

    Saidi, Sami; Ben Amar, Raja

    2016-10-01

    The enzymatic hydrolysis using Prolyve BS coupled to membrane process (Ultrafiltration (UF) and nanofiltration (NF)) is a means of biotransformation of tuna protein waste to Tuna protein hydrolysate (TPH) with higher added values. This method could be an effective solution for the production of bioactive compounds used in various biotechnological applications and minimizing the pollution problems generated by the seafood processing industries. The amino acid composition, functional and antioxidant properties of produced TPH were evaluated. The results show that the glutamic acid, aspartic acid, glycine, alaline, valine and leucine were the major amino acids detected in the TPH profile. After membrane fractionation process, those major amino acids were concentrated in the NF retentate (NFR). The NFR and NF permeate (NFP) have a higher protein solubility (>95 %) when compared to TPH (80 %). Higher oil and water binding capacity were observed in TPH and higher emulsifying and foam stability was found in UF retentate. The NFP showed the highest DPPH radical scavenging activity (65 %). The NFR contained antioxidant amino acid (30.3 %) showed the highest superoxide radical and reducing power activities. The TPH showed the highest iron chelating activity (75 %) compared to other peptide fractions. The effect of the membrane fractionation on the molecular weight distribution of the peptide and their bioactivities was underlined. We concluded that the TPH is a valuable source of bioactive peptides and their peptide fractions may serve as useful ingredients for application in food industry and formulation of nutritional products.

  18. Membrane fouling in microfiltration used for cell harvesting

    NASA Astrophysics Data System (ADS)

    Kaghazchi, Tahereh; Zokaee, Farzin; Zare, Abbas

    2001-03-01

    In the present study the membrane fouling in microfiltration used for cell harvesting in a deadend system has been investigated. Experimental results were analysed in terms of existing membrane filtration models and membrane resistances. The cake filtration model (CFM) and standard blocking model (SBM) have been considered in this study. Various membrane resistances were determined at different processing time, feed concentration and stirring speed. Resistances to permeation in this system include filter medium, pore blocking, adsorption, cake layer and concentration polarization.

  19. Asparagine-linked sugar chains of glycoproteins in calf thymocyte plasma membrane. Isolation and fractionation of oligosaccharides liberated by hydrazinolysis.

    PubMed

    Yoshima, H; Takasaki, S; Kobata, A

    1980-07-01

    The plasma membrane glycoproteins of calf thymocytes were converted to glycopeptides by exhaustive pronase digestion. Glycopeptides with asparagine-linked sugar chains were separated from those with mucine-type sugar chains by Bio-Gel P-10 column chromatography. The asparagine-linked sugar chains were released as oligosaccharides from the peptide moiety by hydrazinolysis and labeled by reduction with NaB[3H]4. The radioactive oligosaccharides were fractionated into fifteen acidic components and ten neutral components by combination of paper electrophoresis and Bio-Gel P-4 column chromatography. The acidic nature of all fifteen acidic components can be ascribed to their N-acetylneuraminic acid residues. The Bio-Gel P-4 column chromatographic patterns of the neutral oligosaccharide fraction and of the neutral fraction obtained on sialidase treatment of the pooled acidic oligosaccharide fraction were totally different, indicating that the acidic oligosaccharides are not simple sialyl derivatives of the neutral oligosaccharides.

  20. Interaction of gentamicin polycation with model and cell membranes.

    PubMed

    Kovács, Eugenia; Savopol, Tudor; Iordache, Maria-Minodora; Săplăcan, Lavinia; Sobaru, Iuliana; Istrate, Claudia; Mingeot-Leclercq, Marie-Paule; Moisescu, Mihaela-Georgeta

    2012-10-01

    The interaction of positively-charged antibiotic gentamicin with cell membranes was studied to determine if any changes in membrane organization were induced by the drug. Opossum kidney epithelia (OK) cells were used as models of eukaryotic cells. Two methods were used: laurdan fluorescence spectroscopy and fluorescence anisotropy recordings on 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene p-toluenesulfonate (TMA-DPH) labeled cell suspensions. Both methods showed an altered membrane hydration and fluidity of gentamicin treated cells. Liposomes prepared from dimyristoyl-phosphatidylcholine (DMPC) mixed with cardiolipin, which mimics the heterogeneous charge composition of the natural cell membrane, were used to determine the effect of gentamicin on artificial bilayers. The membrane lipid packing as revealed by generalized polarization (GP) and fluorescence anizotropy variation with increasing temperature was studied. It was found that the generalized polarization of liposomal membranes containing a negatively charged lipid (cardiolipin) is higher in the presence of gentamicin; in the membrane of living cell (OK), gentamicin induces, on the contrary, a decrease of general polarization. Considering the role of membrane organization in the function of transmembrane channels and receptors, our findings suggest hypotheses that may explain the permeation of gentamicin through the living cell membrane by using these channels.

  1. Cholesterol modulates CFTR confinement in the plasma membrane of primary epithelial cells.

    PubMed

    Abu-Arish, Asmahan; Pandzic, Elvis; Goepp, Julie; Matthes, Elizabeth; Hanrahan, John W; Wiseman, Paul W

    2015-07-07

    The cystic fibrosis transmembrane conductance regulator (CFTR) is a plasma-membrane anion channel that, when mutated, causes the disease cystic fibrosis. Although CFTR has been detected in a detergent-resistant membrane fraction prepared from airway epithelial cells, suggesting that it may partition into cholesterol-rich membrane microdomains (lipid rafts), its compartmentalization has not been demonstrated in intact cells and the influence of microdomains on CFTR lateral mobility is unknown. We used live-cell imaging, spatial image correlation spectroscopy, and k-space image correlation spectroscopy to examine the aggregation state of CFTR and its dynamics both within and outside microdomains in the plasma membrane of primary human bronchial epithelial cells. These studies were also performed during treatments that augment or deplete membrane cholesterol. We found two populations of CFTR molecules that were distinguishable based on their dynamics at the cell surface. One population showed confinement and had slow dynamics that were highly cholesterol dependent. The other, more abundant population was less confined and diffused more rapidly. Treatments that deplete the membrane of cholesterol caused the confined fraction and average number of CFTR molecules per cluster to decrease. Elevating cholesterol had the opposite effect, increasing channel aggregation and the fraction of channels displaying confinement, consistent with CFTR recruitment into cholesterol-rich microdomains with dimensions below the optical resolution limit. Viral infection caused the nanoscale microdomains to fuse into large platforms and reduced CFTR mobility. To our knowledge, these results provide the first biophysical evidence for multiple CFTR populations and have implications for regulation of their surface expression and channel function.

  2. Fluorescence and polarization imaging of membrane dynamics in living cells

    NASA Astrophysics Data System (ADS)

    Wagner, M.; Weber, P.; Bruns, T.; Strauss, W. S. L.; Schneckenburger, H.

    2009-02-01

    Methods of wide field fluorescence microscopy for measuring membrane dynamics in living cells are described. These methods are based on laser pulse excitation of the membrane marker 6-dodecanoyl-2-dimethylamino naphthalene (laurdan) whose emission spectra, fluorescence decay kinetics and anisotropies are sensitive to membrane stiffness and fluidity. Plasma membranes are selected by illumination with an evanescent electromagnetic field and distinguished from intracellular membranes assessed by whole cell illumination. While fluorescence spectra of laurdan appeared red-shifted with decreasing membrane stiffness, fluorescence anisotropy and rotational relaxation times were reduced with increasing membrane fluidity. Membrane stiffness was found to increase with decreasing temperature and increasing amounts of cholesterol. In addition, membrane stiffness of the plasma membrane was always higher than that of intracellular membranes. These effects may have some influence on pathogenesis of certain diseases, uptake of pharmaceutical agents or cell aging. Present experiments are limited to fluorescence microscopy with total internal reflection (TIR) or epi-illumination, but corresponding methods can also be used for screening of larger cell collectives, e.g. in microtiter plates.

  3. Conductivity Measurements of Synthesized Heteropoly Acid Membranes for Proton Exchange Membrane Fuel Cells

    SciTech Connect

    Record, K.A.; Haley, B.T.; Turner, J.

    2006-01-01

    Fuel cell technology is receiving attention due to its potential to be a pollution free method of electricity production when using renewably produced hydrogen as fuel. In a Proton Exchange Membrane (PEM) fuel cell H2 and O2 react at separate electrodes, producing electricity, thermal energy, and water. A key component of the PEM fuel cell is the membrane that separates the electrodes. DuPont’s Nafion® is the most commonly used membrane in PEM fuel cells; however, fuel cell dehydration at temperatures near 100°C, resulting in poor conductivity, is a major hindrance to fuel cell performance. Recent studies incorporating heteropoly acids (HPAs) into membranes have shown an increase in conductivity and thus improvement in performance. HPAs are inorganic materials with known high proton conductivities. The primary objective of this work is to measure the conductivity of Nafion, X-Ionomer membranes, and National Renewable Energy Laboratory (NREL) Developed Membranes that are doped with different HPAs at different concentrations. Four-point conductivity measurements using a third generation BekkTech conductivity test cell are used to determine membrane conductivity. The effect of multiple temperature and humidification levels is also examined. While the classic commercial membrane, Nafion, has a conductivity of approximately 0.10 S/cm, measurements for membranes in this study range from 0.0030 – 0.58 S/cm, depending on membrane type, structure of the HPA, and the relative humidity. In general, the X-ionomer with H6P2W21O71 HPA gave the highest conductivity and the Nafion with the 12-phosphotungstic (PW12) HPA gave the lowest. The NREL composite membranes had conductivities on the order of 0.0013 – 0.025 S/cm.

  4. Preparation and performance of nano silica/Nafion composite membrane for proton exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Wang, Keping; McDermid, Scott; Li, Jing; Kremliakova, Natalia; Kozak, Paul; Song, Chaojie; Tang, Yanghua; Zhang, Jianlu; Zhang, Jiujun

    Composite membranes made from Nafion ionomer with nano phosphonic acid-functionalised silica and colloidal silica were prepared and evaluated for proton exchange membrane fuel cells (PEMFCs) operating at elevated temperature and low relative humidity (RH). The phosphonic acid-functionalised silica additive obtained from a sol-gel process was well incorporated into Nafion membrane. The particle size determined using transmission electron microscope (TEM) had a narrow distribution with an average value of approximately 11 nm and a standard deviation of ±4 nm. The phosphonic acid-functionalised silica additive enhanced proton conductivity and water retention by introducing both acidic groups and porous silica. The proton conductivity of the composite membrane with the acid-functionalised silica was 0.026 S cm -1, 24% higher than that of the unmodified Nafion membrane at 85 °C and 50% RH. Compared with the Nafion membrane, the phosphonic acid-functionalised silica (10% loading level) composite membrane exhibited 60 mV higher fuel cell performance at 1 A cm -2, 95 °C and 35% RH, and 80 mV higher at 0.8 A cm -2, 120 °C and 35% RH. The fuel cell performance of composite membrane made with 6% colloidal silica without acidic group was also higher than unmodified Nafion membrane, however, its performance was lower than the acid-functionalised silica additive composite membrane.

  5. Roles of membrane trafficking in plant cell wall dynamics

    PubMed Central

    Ebine, Kazuo; Ueda, Takashi

    2015-01-01

    The cell wall is one of the characteristic components of plant cells. The cell wall composition differs among cell types and is modified in response to various environmental conditions. To properly generate and modify the cell wall, many proteins are transported to the plasma membrane or extracellular space through membrane trafficking, which is one of the key protein transport mechanisms in eukaryotic cells. Given the diverse composition and functions of the cell wall in plants, the transport of the cell wall components and proteins that are involved in cell wall-related events could be specialized for each cell type, i.e., the machinery for cell wall biogenesis, modification, and maintenance could be transported via different trafficking pathways. In this review, we summarize the recent progress in the current understanding of the roles and mechanisms of membrane trafficking in plant cells and focus on the biogenesis and regulation of the cell wall. PMID:26539200

  6. Using NK Cell Lipid Raft Fractionation to Understand the Role of Lipid Rafts in NK Cell Receptor Signaling.

    PubMed

    Serrano-Pertierra, Esther; López-Larrea, Carlos

    2016-01-01

    Lipid rafts were first defined as detergent-resistant membranes (DRMs) due to their relative insolubility in non-ionic detergents. Although they should not be confused with lipid rafts, DRMs are a valuable starting point for the study of these membrane domains and the interactions of proteins with rafts.Here we describe the isolation of DRMs by ultracentrifugation on a sucrose gradient, a method we have used to study the role of lipid rafts in NKG2D-mediated signaling. We also describe raft fractionation of NK cells involving the selective solubility of β-octylglucoside (β-OG). OG is a non-ionic detergent that efficiently dissolves DRMs but does not disrupt protein associations with the cytoskeleton. Using these two techniques may yield useful information about the proteins involved in receptor recruitment into lipid rafts and the interactions of the actin cytoskeleton with lipid rafts.

  7. Enantiomeric fraction determination of 2-arylpropionic acids in a package plant membrane bioreactor.

    PubMed

    Hashim, Nor H; Stuetz, Richard M; Khan, Stuart J

    2013-05-01

    Enantiomeric compositions of three 2-arylpropionic acid (2-APA) drugs, ibuprofen, naproxen, and ketoprofen, were monitored in a membrane bioreactor (MBR) treating municipal effluent in a small rural town in Australia. Specific enantiomers were determined as amide diastereomers using the chiral derivatizing reagent, (R)-1-phenylethylamine (PEA), followed by gas chromatography-tandem mass spectrometry (GC-MS/MS). The six individual enantiomers were quantified by isotope dilution and the enantiomeric fractions (EFs) were determined. Over four separate sampling events, ibuprofen EF ranged from 0.88 to 0.94 (median 0.93) in the influent and 0.38 to 0.40 (median 0.39) in the effluent. However, no significant change in ketoprofen EF was observed, with influent EFs of 0.56-0.60 (median 0.58) and effluent EFs 0.54-0.68 (median 0.56). This is the first report of enantiospecific analysis of ketoprofen in municipal wastewater and it is not yet clear why such different behavior was observed compared to ibuprofen. Naproxen EF was consistently measured at 0.99 in the influent and ranged from 0.86 to 0.94 (median 0.91) in the effluent. This study demonstrates that EF is a relatively stable parameter and does not fluctuate according to concentration or other short-term variables introduced by sampling limitations. The enantiospecific analysis of chiral chemicals presents a promising approach to elucidate a more thorough understanding of biological treatment processes and a potential tool for monitoring the performance of key biological pathways.

  8. Anatomy of the red cell membrane skeleton: unanswered questions.

    PubMed

    Lux, Samuel E

    2016-01-14

    The red cell membrane skeleton is a pseudohexagonal meshwork of spectrin, actin, protein 4.1R, ankyrin, and actin-associated proteins that laminates the inner membrane surface and attaches to the overlying lipid bilayer via band 3-containing multiprotein complexes at the ankyrin- and actin-binding ends of spectrin. The membrane skeleton strengthens the lipid bilayer and endows the membrane with the durability and flexibility to survive in the circulation. In the 36 years since the first primitive model of the red cell skeleton was proposed, many additional proteins have been discovered, and their structures and interactions have been defined. However, almost nothing is known of the skeleton's physiology, and myriad questions about its structure remain, including questions concerning the structure of spectrin in situ, the way spectrin and other proteins bind to actin, how the membrane is assembled, the dynamics of the skeleton when the membrane is deformed or perturbed by parasites, the role lipids play, and variations in membrane structure in unique regions like lipid rafts. This knowledge is important because the red cell membrane skeleton is the model for spectrin-based membrane skeletons in all cells, and because defects in the red cell membrane skeleton underlie multiple hemolytic anemias.

  9. Acetylcholine Receptor Organization in Membrane Domains in Muscle Cells

    PubMed Central

    Piguet, Joachim; Schreiter, Christoph; Segura, Jean-Manuel; Vogel, Horst; Hovius, Ruud

    2011-01-01

    Nicotinic acetylcholine receptors (nAChR) in muscle fibers are densely packed in the postsynaptic region at the neuromuscular junction. Rapsyn plays a central role in directing and clustering nAChR during cellular differentiation and neuromuscular junction formation; however, it has not been demonstrated whether rapsyn is the only cause of receptor immobilization. Here, we used single-molecule tracking methods to investigate nAChR mobility in plasma membranes of myoblast cells during their differentiation to myotubes in the presence and absence of rapsyn. We found that in myoblasts the majority of nAChR were immobile and that ∼20% of the receptors showed restricted diffusion in small domains of ∼50 nm. In myoblasts devoid of rapsyn, the fraction of mobile nAChR was considerably increased, accompanied by a 3-fold decrease in the immobile population of nAChR with respect to rapsyn-expressing cells. Half of the mobile receptors were confined to domains of ∼120 nm. Measurements performed in heterologously transfected HEK cells confirmed the direct immobilization of nAChR by rapsyn. However, irrespective of the presence of rapsyn, about one-third of nAChR were confined in 300-nm domains. Our results show (i) that rapsyn efficiently immobilizes nAChR independently of other postsynaptic scaffold components; (ii) nAChR is constrained in confined membrane domains independently of rapsyn; and (iii) in the presence of rapsyn, the size of these domains is strongly reduced. PMID:20978122

  10. Electron-beam direct processing on living cell membrane

    SciTech Connect

    Hoshino, Takayuki; Morishima, Keisuke

    2011-10-24

    We demonstrated a direct processing on a living Hep G2 cell membrane in conventional cultivation conditions using an electron beam. Electron beam-induced deposition from liquid precursor 3,4-ethylenedioxythiophene and ablation was performed on the living cells. The 2.5-10 keV electron beam which was irradiated through a 100-nm-thick SiN nanomembrane could induce a deposition pattern and a ablation on a living cell membrane. This electron beam direct processing can provide simple in-situ cell surface modification for an analytical method of living cell membrane dynamic.

  11. Accurate control of oxygen level in cells during culture on silicone rubber membranes with application to stem cell differentiation.

    PubMed

    Powers, Daryl E; Millman, Jeffrey R; Bonner-Weir, Susan; Rappel, Michael J; Colton, Clark K

    2010-01-01

    Oxygen level in mammalian cell culture is often controlled by placing culture vessels in humidified incubators with a defined gas phase partial pressure of oxygen (pO(2gas)). Because the cells are consuming oxygen supplied by diffusion, a difference between pO(2gas) and that experienced by the cells (pO(2cell)) arises, which is maximal when cells are cultured in vessels with little or no oxygen permeability. Here, we demonstrate theoretically that highly oxygen-permeable silicone rubber membranes can be used to control pO(2cell) during culture of cells in monolayers and aggregates much more accurately and can achieve more rapid transient response following a disturbance than on polystyrene and fluorinated ethylene-propylene copolymer membranes. Cell attachment on silicone rubber was achieved by physical adsorption of fibronectin or Matrigel. We use these membranes for the differentiation of mouse embryonic stem cells to cardiomyocytes and compare the results with culture on polystyrene or on silicone rubber on top of polystyrene. The fraction of cells that are cardiomyocyte-like increases with decreasing pO(2) only when using oxygen-permeable silicone membrane-based dishs, which contract on silicone rubber but not polystyrene. The high permeability of silicone rubber results in pO(2cell) being equal to pO(2gas) at the tissue-membrane interface. This, together with geometric information from histological sections, facilitates development of a model from which the pO(2) distribution within the resulting aggregates is computed. Silicone rubber membranes have significant advantages over polystyrene in controlling pO(2cell), and these results suggest they are a valuable tool for investigating pO(2) effects in many applications, such as stem cell differentiation.

  12. Effect of EMP fields on cell membrane potentials

    SciTech Connect

    Gailey, P.C.; Easterly, C.E.

    1993-06-01

    A simple model is presented for cell membrane potentials induced during exposure to electromagnetic pulse (EMP). Using calculated values of internal electric field strength induced during EMP exposure, the model predicts that cell membrane potentials of about 100 mV may be induced for time frames on the order of 10 ns. Possible biological effects of these potentials including electroporation area discussed.

  13. A label-free proteome analysis strategy for identifying quantitative changes in erythrocyte membranes induced by red cell disorders.

    PubMed

    Pesciotta, Esther N; Sriswasdi, Sira; Tang, Hsin-Yao; Mason, Philip J; Bessler, Monica; Speicher, David W

    2012-12-05

    Red blood cells have been extensively studied but many questions regarding membrane properties and pathophysiology remain unanswered. Proteome analysis of red cell membranes is complicated by a very wide dynamic range of protein concentrations as well as the presence of proteins that are very large, very hydrophobic, or heterogeneously glycosylated. This study investigated the removal of other blood cell types, red cell membrane extraction, differing degrees of fractionation using 1-D SDS gels, and label-free quantitative methods to determine optimized conditions for proteomic comparisons of clinical blood samples. The results showed that fractionation of red cell membranes on 1-D SDS gels was more efficient than low-ionic-strength extractions followed by 1-D gel fractionation. When gel lanes were sliced into 30 uniform slices, a good depth of analysis that included the identification of most well-characterized, low-abundance red cell membrane proteins including those present at 500 to 10,000 copies per cell was obtained. Furthermore, the size separation enabled detection of changes due to proteolysis or in vivo protein crosslinking. A combination of Rosetta Elucidator quantitation and subsequent statistical analysis enabled the robust detection of protein differences that could be used to address unresolved questions in red cell disorders. This article is part of a Special Issue entitled: Integrated omics.

  14. Fractionated stem cell infusions for patients with plasma cell myeloma undergoing autologous hematopoietic cell transplantation.

    PubMed

    Landau, Heather; Wood, Kevin; Chung, David J; Koehne, Guenther; Lendvai, Nikoletta; Hassoun, Hani; Lesokhin, Alexander; Hoover, Elizabeth; Zheng, Junting; Devlin, Sean M; Giralt, Sergio

    2016-08-01

    We conducted a phase II trial investigating the impact of fractionated hematopoietic cell infusions on engraftment kinetics and symptom burden in patients with plasma cell myeloma (PCM) undergoing autologous hematopoietic cell transplant (AHCT). We hypothesized that multiple hematopoietic cell infusions would reduce duration of neutropenia and enhance immune recovery resulting in a better tolerated procedure. Twenty-six patients received high-dose melphalan followed by multiple cell infusions (Days 0, +2, +4, +6) and were compared to PCM patients (N = 77) who received high-dose melphalan and a single infusion (Day 0) (concurrent control group). The primary endpoint was number of days with ANC <500K/mcL. Symptom burden was assessed using the MSK-modified MD Anderson Symptom Inventory. Median duration of neutropenia was similar in study (4 days, range 3-5) and control patients (4 days, range 3-9) (p = 0.654). There was no significant difference in the number of red cell or platelet transfusions, days of fever, diarrhea, antibiotics, number of documented infections, or length of admission. Symptom burden surveys showed that AHCT was well-tolerated in both study and control patients. We conclude that fractionated stem cell infusions following high-dose melphalan do not enhance engraftment kinetics or significantly alter patients' clinical course following AHCT in PCM.

  15. Favorable effect of in-situ generated platinum in the membrane on fuel cell membrane durability

    NASA Astrophysics Data System (ADS)

    Macauley, Natalia; Wong, Ka Hung; Watson, Mark; Kjeang, Erik

    2015-12-01

    The overall lifetime of polymer electrolyte fuel cells is often determined by the membrane durability. Platinum, which may dissolve from the catalyst layers during fuel cell operation and deposit in the membrane, has been shown to have both positive and negative effects on membrane stability. In the present work, we analyze what specific conditions are required in order to reach a favorable, membrane stabilizing effect with the controlled use of platinum in the membrane. Using accelerated membrane durability testing, field operated membrane samples, and electron microscopy, we demonstrate that a high platinum concentration with specific particle shapes and sizes is essential for enhanced membrane stability. Specifically, star shaped and dendritic particles with high particle density and high surface area are shown to be preferable. These particles contain high levels of Pt(111) and are expected to have high catalytic activity toward peroxide quenching and crossover gas consumption, thereby mitigating chemical membrane degradation. On the other hand, small, dispersed cubic particles are found to have no effect or the opposite, negative effect on membrane stability.

  16. Cell wall and membrane changes associated with ethambutol resistance in Mycobacterium tuberculosis H37Ra.

    PubMed Central

    Sareen, M; Khuller, G K

    1990-01-01

    Biochemical variations accompanying the acquisition of ethambutol (EMB) resistance in a single-step mutant of Mycobacterium tuberculosis H37Ra were analyzed. Comparative analysis of phospholipids revealed a reduced content in the EMB-resistant strain, particularly in the cell membrane fraction. Significant alterations were observed in the individual phospholipid content and phospholipid fatty acyl group composition of whole cells and subcellular fractions. Quantitative changes were seen in the chemical constituents of the cell walls of resistant cultures in comparison with those of EMB-susceptible cultures of M. tuberculosis. Alterations in the binding of 1-anilinonaphthalene-8-sulfonate to whole cells of an EMB-resistant strain indicated structural changes on the cell surface. Structural changes in the cell wall may play an important role in the resistance of M. tuberculosis H37Ra to EMB. PMID:2126690

  17. Radiation-Grafted Polymer Electrolyte Membranes for Water Electrolysis Cells: Evaluation of Key Membrane Properties.

    PubMed

    Albert, Albert; Barnett, Alejandro O; Thomassen, Magnus S; Schmidt, Thomas J; Gubler, Lorenz

    2015-10-14

    Radiation-grafted membranes can be considered an alternative to perfluorosulfonic acid (PFSA) membranes, such as Nafion, in a solid polymer electrolyte electrolyzer. Styrene, acrylonitrile, and 1,3-diisopropenylbenzene monomers are cografted into preirradiated 50 μm ethylene tetrafluoroethylene (ETFE) base film, followed by sulfonation to introduce proton exchange sites to the obtained grafted films. The incorporation of grafts throughout the thickness is demonstrated by scanning electron microscopy/energy-dispersive X-ray spectroscopy (SEM/EDX) analysis of the membrane cross-sections. The membranes are analyzed in terms of grafting kinetics, ion-exchange capacity (IEC), and water uptake. The key properties of radiation-grafted membranes and Nafion, such as gas crossover, area resistance, and mechanical properties, are evaluated and compared. The plot of hydrogen crossover versus area resistance of the membranes results in a property map that indicates the target areas for membrane development for electrolyzer applications. Tensile tests are performed to assess the mechanical properties of the membranes. Finally, these three properties are combined to establish a figure of merit, which indicates that radiation-grafted membranes obtained in the present study are promising candidates with properties superior to those of Nafion membranes. A water electrolysis cell test is performed as proof of principle, including a comparison to a commercial membrane electrode assembly (MEA).

  18. Durability of symmetrically and asymmetrically porous polybenzimidazole membranes for high temperature proton exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Jheng, Li-Cheng; Chang, Wesley Jen-Yang; Hsu, Steve Lien-Chung; Cheng, Po-Yang

    2016-08-01

    Two types of porous polybenzimidazole (PBI) membranes with symmetric and asymmetric morphologies were fabricated by the template-leaching method and characterized by scanning electron microscope (SEM). Their physicochemical properties were compared in terms of acid-doping level, proton conductivity, mechanical strength, and oxidative stability. The durability of fuel cell operation is one of the most challenging for the PBI based membrane electrode assembly (MEA) used in high-temperature proton exchange membrane fuel cells (HT-PEMFCs). In the present work, we carried out a long-term steady-state fuel cell test to compare the effect of membrane structure on the cell voltage degradation. It has also been demonstrated that the asymmetrically porous PBI could bring some notable improvements on the durability of fuel cell operation, the fuel crossover problem, and the phosphoric acid leakage.

  19. Studying the Nucleated Mammalian Cell Membrane by Single Molecule Approaches

    PubMed Central

    Wang, Feng; Wu, Jiazhen; Gao, Jing; Liu, Shuheng; Jiang, Junguang; Jiang, Shibo; Wang, Hongda

    2014-01-01

    The cell membrane plays a key role in compartmentalization, nutrient transportation and signal transduction, while the pattern of protein distribution at both cytoplasmic and ectoplasmic sides of the cell membrane remains elusive. Using a combination of single-molecule techniques, including atomic force microscopy (AFM), single molecule force spectroscopy (SMFS) and stochastic optical reconstruction microscopy (STORM), to study the structure of nucleated cell membranes, we found that (1) proteins at the ectoplasmic side of the cell membrane form a dense protein layer (4 nm) on top of a lipid bilayer; (2) proteins aggregate to form islands evenly dispersed at the cytoplasmic side of the cell membrane with a height of about 10–12 nm; (3) cholesterol-enriched domains exist within the cell membrane; (4) carbohydrates stay in microdomains at the ectoplasmic side; and (5) exposed amino groups are asymmetrically distributed on both sides. Based on these observations, we proposed a Protein Layer-Lipid-Protein Island (PLLPI) model, to provide a better understanding of cell membrane structure, membrane trafficking and viral fusion mechanisms. PMID:24806512

  20. In-situ membrane hydration measurement of proton exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Lai, Yeh-Hung; Fly, Gerald W.; Clapham, Shawn

    2015-01-01

    Achieving proper membrane hydration control is one of the most critical aspects of PEM fuel cell development. This article describes the development and application of a novel 50 cm2 fuel cell device to study the in-situ membrane hydration by measuring the through-thickness membrane swelling via an array of linear variable differential transducers. Using this setup either as an air/air (dummy) cell or as a hydrogen/air (operating) cell, we performed a series of hydration and dehydration experiments by cycling the RH of the inlet gas streams at 80 °C. From the linear relationship between the under-the-land swelling and the over-the-channel water content, the mechanical constraint within the fuel cell assembly can suppress the membrane water uptake by 11%-18%. The results from the air/air humidity cycling test show that the membrane can equilibrate within 120 s for all RH conditions and that membrane can reach full hydration at a RH higher than 140% in spite of the use of a liquid water impermeable Carbel MP30Z microporous layer. This result confirms that the U.S. DOE's humidity cycling mechanical durability protocol induces sufficient humidity swings to maximize hygrothermal mechanical stresses. This study shows that the novel experimental technique can provide a robust and accurate means to study the in-situ hydration of thin membranes subject to a wide range of fuel cell conditions.

  1. Axially resolved polarisation microscopy of membrane dynamics in living cells

    NASA Astrophysics Data System (ADS)

    Wagner, Michael; Weber, Petra; Schneckenburger, Herbert

    2007-07-01

    Membrane dynamics has a large impact on cellular uptake and release of various metabolites or pharmaceutical agents. For a deeper understanding of the cellular processes involved, we used U373-MG human glioblastoma cells as a model system. As conventional microscopy does not permit to investigate individual layers in living cells, we used structured illumination techniques and total internal reflection fluorescence microscopy (TIRFM) to analyse the plasma membrane and intracellular membranes of living cells selectively. Optical image sections provide a high resolution and the possibility of 3D reconstruction. Membranes of living cells were characterized by the membrane marker 6-dodecanoyl-2-dimethylamino naphthalene (laurdan). Due to its spectral and kinetic properties this fluorescence marker appears appropriate for measuring membrane stiffness and fluidity. After excitation with linearly polarized laser pulses, membrane fluidity of human glioblastoma cells was determined by measurements of steady-state and time-resolved fluorescence anisotropy r(t), since with increasing viscosity of the environment, the rotation of an excited molecule is impeded. The corresponding time constant τ r of molecular relaxation decreased with temperature and increased with the amount of cholesterol. In addition, fluorescence anisotropy r(t) values of the plasma membrane were larger than the values of intracellular membranes for all temperatures in the range of 16°C<=T<=41°C.

  2. Empirical membrane lifetime model for heavy duty fuel cell systems

    NASA Astrophysics Data System (ADS)

    Macauley, Natalia; Watson, Mark; Lauritzen, Michael; Knights, Shanna; Wang, G. Gary; Kjeang, Erik

    2016-12-01

    Heavy duty fuel cells used in transportation system applications such as transit buses expose the fuel cell membranes to conditions that can lead to lifetime-limiting membrane failure via combined chemical and mechanical degradation. Highly durable membranes and reliable predictive models are therefore needed in order to achieve the ultimate heavy duty fuel cell lifetime target of 25,000 h. In the present work, an empirical membrane lifetime model was developed based on laboratory data from a suite of accelerated membrane durability tests. The model considers the effects of cell voltage, temperature, oxygen concentration, humidity cycling, humidity level, and platinum in the membrane using inverse power law and exponential relationships within the framework of a general log-linear Weibull life-stress statistical distribution. The obtained model is capable of extrapolating the membrane lifetime from accelerated test conditions to use level conditions during field operation. Based on typical conditions for the Whistler, British Columbia fuel cell transit bus fleet, the model predicts a stack lifetime of 17,500 h and a membrane leak initiation time of 9200 h. Validation performed with the aid of a field operated stack confirmed the initial goal of the model to predict membrane lifetime within 20% of the actual operating time.

  3. Ultrafiltration by a compacted clay membrane. I - Oxygen and hydrogen isotopic fractionation. II - Sodium ion exclusion at various ionic strengths.

    NASA Technical Reports Server (NTRS)

    Coplen, T. B.; Hanshaw, B. B.

    1973-01-01

    Laboratory experiments were carried out to determine the magnitude of the isotopic fractionation of distilled water and of 0.01N NaCl forced to flow at ambient temperature under a hydraulic pressure drop of 100 bars across a montmorillonite disk compacted to a porosity of 35% by a pressure of 330 bars. The ultrafiltrates in both experiments were depleted in D by 2.5% and in O-18 by 0.8% relative to the residual solution. No additional isotopic fractionation due to a salt-filtering mechanism was observed at NaCl concentrations up to 0.01N. Adsorption is most likely the principal mechanism which produces isotopic fractionation, but molecular diffusion may play a minor role. The results suggest that oxygen and hydrogen isotopic fractionation of ground water during passage through compacted clayey sediments should be a common occurrence, in accord with published interpretations of isotopic data from the Illinois and Alberta basins. It is shown how it is possible to proceed from the ion exchange capacity of clay minerals and, by means of the Donnan membrane equilibrium concept and the Teorell-Meyer-Siever theory, develop a theory to explain why and to what extent ultrafiltration occurs when solutions of known concentration are forced to flow through a clay membrane.

  4. Revealing the Dynamics of Thylakoid Membranes in Living Cyanobacterial Cells

    DOE PAGES

    Stingaciu, Laura-Roxana; O’Neill, Hugh; Liberton, Michelle; ...

    2016-01-21

    Cyanobacteria are photosynthetic prokaryotes that make major contributions to the production of the oxygen in the Earth atmosphere. The photosynthetic machinery in cyanobacterial cells is housed in flattened membrane structures called thylakoids. The structural organization of cyanobacterial cells and the arrangement of the thylakoid membranes in response to environmental conditions have been widely investigated. However, there is limited knowledge about the internal dynamics of these membranes in terms of their flexibility and motion during the photosynthetic process. We present a direct observation of thylakoid membrane undulatory motion in vivo and show a connection between membrane mobility and photosynthetic activity. High-resolutionmore » inelastic neutron scattering experiments on the cyanobacterium Synechocystis sp. PCC 6803 assessed the flexibility of cyanobacterial thylakoid membrane sheets and the dependence of the membranes on illumination conditions. We observed softer thylakoid membranes in the dark that have three-to four fold excess mobility compared to membranes under high light conditions. We find our analysis indicates that electron transfer between photosynthetic reaction centers and the associated electrochemical proton gradient across the thylakoid membrane result in a significant driving force for excess membrane dynamics. Lastly, these observations provide a deeper understanding of the relationship between photosynthesis and cellular architecture.« less

  5. Revealing the Dynamics of Thylakoid Membranes in Living Cyanobacterial Cells

    SciTech Connect

    Stingaciu, Laura-Roxana; O’Neill, Hugh; Urban, Volker S.; Ohl, Michael

    2016-01-21

    Cyanobacteria are photosynthetic prokaryotes that make major contributions to the production of the oxygen in the Earth atmosphere. The photosynthetic machinery in cyanobacterial cells is housed in flattened membrane structures called thylakoids. The structural organization of cyanobacterial cells and the arrangement of the thylakoid membranes in response to environmental conditions have been widely investigated. However, there is limited knowledge about the internal dynamics of these membranes in terms of their flexibility and motion during the photosynthetic process. We present a direct observation of thylakoid membrane undulatory motion in vivo and show a connection between membrane mobility and photosynthetic activity. High-resolution inelastic neutron scattering experiments on the cyanobacterium Synechocystis sp. PCC 6803 assessed the flexibility of cyanobacterial thylakoid membrane sheets and the dependence of the membranes on illumination conditions. We observed softer thylakoid membranes in the dark that have three-to four fold excess mobility compared to membranes under high light conditions. We find our analysis indicates that electron transfer between photosynthetic reaction centers and the associated electrochemical proton gradient across the thylakoid membrane result in a significant driving force for excess membrane dynamics. Lastly, these observations provide a deeper understanding of the relationship between photosynthesis and cellular architecture.

  6. Revealing the Dynamics of Thylakoid Membranes in Living Cyanobacterial Cells

    NASA Astrophysics Data System (ADS)

    Stingaciu, Laura-Roxana; O’Neill, Hugh; Liberton, Michelle; Urban, Volker S.; Pakrasi, Himadri B.; Ohl, Michael

    2016-01-01

    Cyanobacteria are photosynthetic prokaryotes that make major contributions to the production of the oxygen in the Earth atmosphere. The photosynthetic machinery in cyanobacterial cells is housed in flattened membrane structures called thylakoids. The structural organization of cyanobacterial cells and the arrangement of the thylakoid membranes in response to environmental conditions have been widely investigated. However, there is limited knowledge about the internal dynamics of these membranes in terms of their flexibility and motion during the photosynthetic process. We present a direct observation of thylakoid membrane undulatory motion in vivo and show a connection between membrane mobility and photosynthetic activity. High-resolution inelastic neutron scattering experiments on the cyanobacterium Synechocystis sp. PCC 6803 assessed the flexibility of cyanobacterial thylakoid membrane sheets and the dependence of the membranes on illumination conditions. We observed softer thylakoid membranes in the dark that have three-to four fold excess mobility compared to membranes under high light conditions. Our analysis indicates that electron transfer between photosynthetic reaction centers and the associated electrochemical proton gradient across the thylakoid membrane result in a significant driving force for excess membrane dynamics. These observations provide a deeper understanding of the relationship between photosynthesis and cellular architecture.

  7. Detecting Nanodomains in Living Cell Membrane by Fluorescence Correlation Spectroscopy

    NASA Astrophysics Data System (ADS)

    He, Hai-Tao; Marguet, Didier

    2011-05-01

    Cell membranes actively participate in numerous cellular functions. Inasmuch as bioactivities of cell membranes are known to depend crucially on their lateral organization, much effort has been focused on deciphering this organization on different length scales. Within this context, the concept of lipid rafts has been intensively discussed over recent years. In line with its ability to measure diffusion parameters with great precision, fluorescence correlation spectroscopy (FCS) measurements have been made in association with innovative experimental strategies to monitor modes of molecular lateral diffusion within the plasma membrane of living cells. These investigations have allowed significant progress in the characterization of the cell membrane lateral organization at the suboptical level and have provided compelling evidence for the in vivo existence of raft nanodomains. We review these FCS-based studies and the characteristic structural features of raft nanodomains. We also discuss the findings in regards to the current view of lipid rafts as a general membrane-organizing principle.

  8. Photocatalytic Degradation of Cell Membrane Coatings for Controlled Drug Release.

    PubMed

    Rao, Lang; Meng, Qian-Fang; Huang, Qinqin; Liu, Pei; Bu, Lin-Lin; Kondamareddy, Kiran Kumar; Guo, Shi-Shang; Liu, Wei; Zhao, Xing-Zhong

    2016-06-01

    Biomimetic cell-membrane-camouflaged particles with desirable features have been widely used for various biomedical applications. However, there are few reports on employing these particles for cancer drug delivery due to the failure of the membrane coatings to be efficiently degraded in the tumor microenvironment which hampers the drug release. In this work, core-shell SiO2 @TiO2 nanoparticles with enhanced photocatalytic activity are used for controlled degradation of surface erythrocyte membrane coatings. The antitumor drug docetaxel is encapsulated into nanocarriers to demonstrate the controlled drug release under ultraviolet irradiation, and the drug-loaded nanoparticles are further used for enhanced cancer cell therapy. Here, a simple but practical method for degradation of cell membrane coatings is presented, and a good feasibility of using cell membrane-coated nanocarriers for controlled drug delivery is demonstrated.

  9. Cell Membranes Under Hydrostatic Pressure Subjected to Micro-Injection

    NASA Astrophysics Data System (ADS)

    Vassilev, Vassil M.; Kostadinov, Kostadin G.; Mladenov, Ivaïlo M.; Shulev, Assen A.; Stoilov, Georgi I.; Djondjorov, Peter A.

    2011-04-01

    The work is concerned with the determination of the mechanical behaviour of cell membranes under uniform hydrostatic pressure subject to micro-injections. For that purpose, assuming that the shape of the deformed cell membrane is axisymmetric a variational statement of the problem is developed on the ground of the so-called spontaneous curvature model. In this setting, the cell membrane is regarded as an axisymmetric surface in the three-dimensional Euclidean space providing a stationary value of the shape energy functional under the constraint of fixed total area and fixed enclosed volume. The corresponding Euler-Lagrange equations and natural boundary conditions are derived, analyzed and used to express the forces and moments in the membrane. Several examples of such surfaces representing possible shapes of cell membranes under pressure subjected to micro injection are determined numerically.

  10. The enriched fraction of Elephantopus scaber Triggers apoptosis and inhibits multi-drug resistance transporters in human epithelial cancer cells

    PubMed Central

    Beeran, Asmy Appadath; Maliyakkal, Naseer; Rao, Chamallamudi Mallikarjuna; Udupa, Nayanabhirama

    2015-01-01

    Background: Medicinal plants have played an important role in the development of clinically useful anticancer agents. Elephantopus scaber (Asteraceae) (ES) is widely used in Indian traditional system of medicine for the treatment of various ailments including cancer. Objective: To investigate anticancer effects of ES in human epithelial cancer cells. Materials and Methods: Cytotoxicity of ethanolic extract of ES (ES-ET) and its fractions, such as ES Petroleum ether fraction (ES-PET), ES Dichloromethane fraction (ES DCM), n Butyl alcohol fraction (ES-BT), and ES-Rest (ES-R) were assessed in human epithelial cancer cell lines using sulforhodamine B (SRB) assay. Acridine orange/ethidium bromide assay and Hoechst 33342 assays were used to gauge induction of apoptosis. Cell cycle analysis and micronuclei assay were used to assess cell cycle specific pharmacological effects and drug induced genotoxicty. Further, the ability of ES to inhibit multi drug resistant (MDR) transporters (ABC-B1 and ABC-G2) was determined by Rhodamine (Rho) and Mitoxantrone (MXR) efflux assays. Results: The enriched fraction of ES (ES DCM) possessed dose-dependent potent cytotoxicity in human epithelial cancer cells. Further, treatment of cancer cells (HeLa, A549, MCF-7, and Caco-2) with ES DCM showed hall mark properties of apoptosis (membrane blebbing, nuclear condensation etc.). Similarly, ES DCM caused enhanced sub G0 content and micronuclei formation indicating the induction of apoptosis and drug induced genotoxicity in cancer cells, respectively. Interestingly, ES DCM inhibited MDR transporters (ABC B1 and ABC G2) in cancer cells. Conclusion: The enriched fraction of ES imparted cytotoxic effects, triggered apoptosis, induced genotoxicity, and inhibited MDR transporters in human epithelial cancer cells. Thus, ES appears to be potential anticancer agent. PMID:25829763

  11. Antiproliferative activity of buttermilk lipid fractions isolated using food grade and non-food grade solvents on human cancer cell lines.

    PubMed

    Castro-Gómez, Pilar; Rodríguez-Alcalá, Luis M; Monteiro, Karin M; Ruiz, Ana L T G; Carvalho, João E; Fontecha, Javier

    2016-12-01

    Buttermilk is a dairy by-product with a high content of milk fat globule membranes (MFGMs), whose protein constituents are reported to be antiproliferative. Lipids represent about half of the composition of MFGM. The aim of this study was to isolate buttermilk lipid fractions and evaluate their potential antiproliferative effect. Selective extraction with food grade or non-food grade solvents was performed. Antiproliferative effectiveness of lipid extracts and their neutral and polar fractions was evaluated on nine human cancer cell lines. Fractions obtained using food grade ethanol gave a higher yield than those obtained using non-food grade solvents, and they effectively inhibited cell viability of the cancer cell lines investigated. These fractions, rich in phospho- and sphingolipids, were strongly antiproliferative against human ovary and colon cancer cells. This observation allowed us to hypothesize further analyses aimed at promoting the use of buttermilk polar lipid fractions as functional food additives.

  12. How the antimicrobial peptides destroy bacteria cell membrane: Translocations vs. membrane buckling

    NASA Astrophysics Data System (ADS)

    Golubovic, Leonardo; Gao, Lianghui; Chen, Licui; Fang, Weihai

    2012-02-01

    In this study, coarse grained Dissipative Particle Dynamics simulation with implementation of electrostatic interactions is developed in constant pressure and surface tension ensemble to elucidate how the antimicrobial peptide molecules affect bilayer cell membrane structure and kill bacteria. We find that peptides with different chemical-physical properties exhibit different membrane obstructing mechanisms. Peptide molecules can destroy vital functions of the affected bacteria by translocating across their membranes via worm-holes, or by associating with membrane lipids to form hydrophilic cores trapped inside the hydrophobic domain of the membranes. In the latter scenario, the affected membranes are strongly corrugated (buckled) in accord with very recent experimental observations [G. E. Fantner et al., Nat. Nanotech., 5 (2010), pp. 280-285].

  13. Band 3 and glycophorin are progressively aggregated in density-fractionated sickle and normal red blood cells. Evidence from rotational and lateral mobility studies.

    PubMed Central

    Corbett, J D; Golan, D E

    1993-01-01

    Band 3 aggregation in the plane of the red blood cell (RBC) membrane is postulated to be important in the pathophysiology of hemolysis of dense sickle and normal RBCs. We used the fluorescence photobleaching recovery and polarized fluorescence depletion techniques to measure the lateral and rotational mobility of band 3, glycophorins, and phospholipid analogues in membranes of density-separated intact RBCs from seven patients with sickle cell disease and eight normal controls. The fractions of laterally mobile band 3 and glycophorin decreased progressively as sickle RBC density increased. Normal RBCs also showed a progressive decrease in band 3 fractional mobility with increasing buoyant density. Rapidly rotating, slowly rotating, and rotationally immobile forms of band 3 were observed in both sickle and normal RBC membranes. The fraction of rapidly rotating band 3 progressively decreased and the fraction of rotationally immobile band 3 progressively increased with increasing sickle RBC density. Changes in the fraction of rotationally immobile band 3 were not reversible upon hypotonic swelling of dense sickle RBCs, and normal RBCs osmotically shrunken in sucrose buffers failed to manifest band 3 immobilization at median cell hemoglobin concentration values characteristic of dense sickle RBCs. We conclude that dense sickle and normal RBCs acquire irreversible membrane abnormalities that cause transmembrane protein immobilization and band 3 aggregation. Band 3 aggregates could serve as cell surface sites of autologous antibody binding and thereby lead to removal of dense sickle and normal (senescent) RBCs from the circulation. PMID:8423219

  14. Exploring the inhibitory effect of membrane tension on cell polarization

    PubMed Central

    Wang, Jing; Yang, Gen; Ouyang, Qi; Wang, Yugang; Zhang, Lei

    2017-01-01

    Cell polarization toward an attractant is influenced by both physical and chemical factors. Most existing mathematical models are based on reaction-diffusion systems and only focus on the chemical process occurring during cell polarization. However, membrane tension has been shown to act as a long-range inhibitor of cell polarization. Here, we present a cell polarization model incorporating the interplay between Rac GTPase, filamentous actin (F-actin), and cell membrane tension. We further test the predictions of this model by performing single cell measurements of the spontaneous polarization of cancer stem cells (CSCs) and non-stem cancer cells (NSCCs), as the former have lower cell membrane tension. Based on both our model and the experimental results, cell polarization is more sensitive to stimuli under low membrane tension, and high membrane tension improves the robustness and stability of cell polarization such that polarization persists under random perturbations. Furthermore, our simulations are the first to recapitulate the experimental results described by Houk et al., revealing that aspiration (elevation of tension) and release (reduction of tension) result in a decrease in and recovery of the activity of Rac-GTP, respectively, and that the relaxation of tension induces new polarity of the cell body when a cell with the pseudopod-neck-body morphology is severed. PMID:28135277

  15. The Flocculating Cationic Polypetide from Moringa oleifera Seeds Damages Bacterial Cell Membranes by Causing Membrane Fusion.

    PubMed

    Shebek, Kevin; Schantz, Allen B; Sines, Ian; Lauser, Kathleen; Velegol, Stephanie; Kumar, Manish

    2015-04-21

    A cationic protein isolated from the seeds of the Moringa oleifera tree has been extensively studied for use in water treatment in developing countries and has been proposed for use in antimicrobial and therapeutic applications. However, the molecular basis for the antimicrobial action of this peptide, Moringa oleifera cationic protein (MOCP), has not been previously elucidated. We demonstrate here that a dominant mechanism of MOCP antimicrobial activity is membrane fusion. We used a combination of cryogenic electron microscopy (cryo-EM) and fluorescence assays to observe and study the kinetics of fusion of membranes in liposomes representing model microbial cells. We also conducted cryo-EM experiments on E. coli cells where MOCP was seen to fuse the inner and outer membranes. Coarse-grained molecular dynamics simulations of membrane vesicles with MOCP molecules were used to elucidate steps in peptide adsorption, stalk formation, and fusion between membranes.

  16. Membrane curvature in cell biology: An integration of molecular mechanisms.

    PubMed

    Jarsch, Iris K; Daste, Frederic; Gallop, Jennifer L

    2016-08-15

    Curving biological membranes establishes the complex architecture of the cell and mediates membrane traffic to control flux through subcellular compartments. Common molecular mechanisms for bending membranes are evident in different cell biological contexts across eukaryotic phyla. These mechanisms can be intrinsic to the membrane bilayer (either the lipid or protein components) or can be brought about by extrinsic factors, including the cytoskeleton. Here, we review examples of membrane curvature generation in animals, fungi, and plants. We showcase the molecular mechanisms involved and how they collaborate and go on to highlight contexts of curvature that are exciting areas of future research. Lessons from how membranes are bent in yeast and mammals give hints as to the molecular mechanisms we expect to see used by plants and protists.

  17. Membrane curvature in cell biology: An integration of molecular mechanisms

    PubMed Central

    Daste, Frederic

    2016-01-01

    Curving biological membranes establishes the complex architecture of the cell and mediates membrane traffic to control flux through subcellular compartments. Common molecular mechanisms for bending membranes are evident in different cell biological contexts across eukaryotic phyla. These mechanisms can be intrinsic to the membrane bilayer (either the lipid or protein components) or can be brought about by extrinsic factors, including the cytoskeleton. Here, we review examples of membrane curvature generation in animals, fungi, and plants. We showcase the molecular mechanisms involved and how they collaborate and go on to highlight contexts of curvature that are exciting areas of future research. Lessons from how membranes are bent in yeast and mammals give hints as to the molecular mechanisms we expect to see used by plants and protists. PMID:27528656

  18. Fuel cell using novel electrolyte membrane

    SciTech Connect

    Polak, A.J.; Beuhler, A.J.

    1986-06-10

    An apparatus is described for producing electricity from a fuel gas having a gaseous component which is capable, in the presence of a catalytic agent, of dissociating to yield hydrogen ions comprising: (a) a thin film organic-inorganic membrane which comprises a single phase blend from about 1% to about 70% by weight of a heteropoly acid and salts; (b) a membrane housing comprising a fuel gas chamber and an oxidant gas chamber separated by a substantially imporous partition comprising the membrane defined in element (a), the membrane having a first surface in communication with the fuel gas chamber and a second surface in communication with the oxidant gas chamber; (c) two separate portions of catalytic agent effective to promote dissociation and combination, one portion in contact with the first surface of the membrane and one portion in contact with the second surface of the membrane; and, (d) means for forming electrical connection in operative contact with the catalytic agent in contact with the first surface of the membrane and in operative contact with the catalytic agent in contact with the second surface of the membrane.

  19. Unbound fraction of fluconazole and linezolid in human plasma as determined by ultrafiltration: Impact of membrane type.

    PubMed

    Kratzer, Alexander; Kees, Frieder; Dorn, Christoph

    2016-12-15

    Ultrafiltration is a rapid and convenient method to determine the free concentrations of drugs in plasma. Several ultrafiltration devices based on Eppendorf cups are commercially available, but are not validated for such use by the manufacturer. Plasma pH, temperature and relative centrifugal force as well as membrane type can influence the results. In the present work, we developed an ultrafiltration method in order to determine the free concentrations of linezolid or fluconazole, both neutral and moderately lipophilic antiinfective drugs for parenteral as well as oral administration, in plasma of patients. Whereas both substances behaved relatively insensitive in human plasma regarding variations in pH (7.0-8.5), temperature (5-37°C) or relative centrifugal force (1000-10.000xg), losses of linezolid were observed with the Nanosep Omega device due to adsorption onto the polyethersulfone membrane (unbound fraction 75% at 100mg/L and 45% at 0.1mg/L, respectively). No losses were observed with Vivacon which is equipped with a membrane of regenerated cellulose. With fluconazole no differences between Nanosep and Vivacon were observed. Applying standard conditions (pH 7.4/37°C/1000xg/20min), the mean unbound fraction of linezolid in pooled plasma from healthy volunteers was 81.5±2.8% using Vivacon, that of fluconazole was 87.9±3.5% using Nanosep or 89.4±3.3% using Vivacon. The unbound fraction of linezolid was 85.4±3.7% in plasma samples from surgical patients and 92.1±6.2% in ICU patients, respectively. The unbound fraction of fluconazole was 93.9±3.3% in plasma samples from ICU patients.

  20. Fluorescence imaging of cholesterol and temperature dependent cell membrane dynamics

    NASA Astrophysics Data System (ADS)

    Weber, Petra; Wagner, Michael; Strauss, Wolfgang S. L.; Schneckenburger, Herbert

    2007-07-01

    Cholesterol content is an important factor for membrane dynamics of living cells. With well defined protocols of depletion and enrichment the impact of cholesterol on membrane dynamics was examined by fluorescence microscopy. In addition, the intracellular cholesterol content was determined with biochemical methods. Changes of cholesterol amounts in cell membranes have previously been related to specific disease and may have some influence on the uptake of pharmaceutical agents. A combination of conventional and total internal reflection fluorescence microscopy was applied to the fluorescence marker laurdan, a polarity-sensitive probe, whose electronic excitation energy is different in polar and non-polar environment. Once incorporated into cell membranes, the fluorescence of laurdan shows a spectral shift towards longer wavelength when its molecules get into contact with adjacent water molecules, e.g. when a phase transition from the tightly packed gel phase to the liquid crystalline phase of membrane lipids occurs. The generalized polarization (GP, characterizing this spectral shift) as well as the fluorescence lifetime (τ) of laurdan revealed to be appropriate measures for membrane stiffness and fluidity. GP generally decreased with increasing temperature and was always higher for the plasma membrane than for intracellular membranes. Enrichment of cholesterol caused a pronounced increase, whereas depletion of cholesterol caused a decrease of GP. In addition, pronounced changes of the fluorescence lifetime pattern occurred in the subnanosecond range. GP, and τ were determined as integral values of single cells or small cell collectives and were also displayed as microscopic images.

  1. Layer-by-layer cell membrane assembly

    NASA Astrophysics Data System (ADS)

    Matosevic, Sandro; Paegel, Brian M.

    2013-11-01

    Eukaryotic subcellular membrane systems, such as the nuclear envelope or endoplasmic reticulum, present a rich array of architecturally and compositionally complex supramolecular targets that are as yet inaccessible. Here we describe layer-by-layer phospholipid membrane assembly on microfluidic droplets, a route to structures with defined compositional asymmetry and lamellarity. Starting with phospholipid-stabilized water-in-oil droplets trapped in a static droplet array, lipid monolayer deposition proceeds as oil/water-phase boundaries pass over the droplets. Unilamellar vesicles assembled layer-by-layer support functional insertion both of purified and of in situ expressed membrane proteins. Synthesis and chemical probing of asymmetric unilamellar and double-bilayer vesicles demonstrate the programmability of both membrane lamellarity and lipid-leaflet composition during assembly. The immobilized vesicle arrays are a pragmatic experimental platform for biophysical studies of membranes and their associated proteins, particularly complexes that assemble and function in multilamellar contexts in vivo.

  2. The application of Dow Chemical's perfluorinated membranes in proton-exchange membrane fuel cells

    NASA Technical Reports Server (NTRS)

    Eisman, G. A.

    1989-01-01

    Dow Chemical's research activities in fuel cell devices revolves around the development and subsequent investigation of the perfluorinated inomeric membrane separator useful in proton-exchange membrane systems. Work is currently focusing on studying the effects of equivalent weight, thickness, water of hydration, pretreatment procedures, as well as the degree of water management required for a given membrane separator in the cell. The presentation will include details of certain aspects of the above as well as some of the requirements for high and low power generation.

  3. Improved Membrane Materials for PEM Fuel Cell Application

    SciTech Connect

    Kenneth A. Mauritz; Robert B. Moore

    2008-06-30

    The overall goal of this project is to collect and integrate critical structure/property information in order to develop methods that lead to significant improvements in the durability and performance of polymer electrolyte membrane fuel cell (PEMFC) materials. This project is focused on the fundamental improvement of PEMFC membrane materials with respect to chemical, mechanical and morphological durability as well as the development of new inorganically-modified membranes.

  4. Plasma membrane proteomics of human embryonic stem cells and human embryonal carcinoma cells.

    PubMed

    Dormeyer, Wilma; van Hoof, Dennis; Braam, Stefan R; Heck, Albert J R; Mummery, Christine L; Krijgsveld, Jeroen

    2008-07-01

    Human embryonic stem cells (hESCs) are of immense interest in regenerative medicine as they can self-renew indefinitely and can give rise to any adult cell type. Human embryonal carcinoma cells (hECCs) are the malignant counterparts of hESCs found in testis tumors. hESCs that have acquired chromosomal abnormalities in culture are essentially indistinguishable from hECC. Direct comparison of karyotypically normal hESCs with hECCs could lead to understanding differences between their mechanisms of growth control and contribute to implementing safe therapeutic use of stem cells without the development of germ cell cancer. While several comparisons of hECCs and hESCs have been reported, their cell surface proteomes are largely unknown, partly because plasma membrane proteomics is still a major challenge. Here, we present a strategy for the identification of plasma membrane proteins that has been optimized for application to the relatively small numbers of stem cells normally available, and that does not require tedious cell fractionation. The method led to the identification of 237 and 219 specific plasma membrane proteins in the hESC line HUES-7 and the hECC line NT2/D1, respectively. In addition to known stemness-associated cell surface markers like ALP, CD9, and CTNNB, a large number of receptors, transporters, signal transducers, and cell-cell adhesion proteins were identified. Our study revealed that several Hedgehog and Wnt pathway members are differentially expressed in hESCs and hECCs including NPC1, FZD2, FZD6, FZD7, LRP6, and SEMA4D, which play a pivotal role in stem cell self-renewal and cancer growth. Various proteins encoded on chromosome 12p, duplicated in testicular cancer, were uniquely identified in hECCs. These included GAPDH, LDHB, YARS2, CLSTN3, CSDA, LRP6, NDUFA9, and NOL1, which are known to be upregulated in testicular cancer. Distinct HLA molecules were revealed on the surface of hESCs and hECCs, despite their low abundance. Results were

  5. High lipid order of Arabidopsis cell-plate membranes mediated by sterol and DYNAMIN-RELATED PROTEIN1A function.

    PubMed

    Frescatada-Rosa, Márcia; Stanislas, Thomas; Backues, Steven K; Reichardt, Ilka; Men, Shuzhen; Boutté, Yohann; Jürgens, Gerd; Moritz, Thomas; Bednarek, Sebastian Y; Grebe, Markus

    2014-12-01

    Membranes of eukaryotic cells contain high lipid-order sterol-rich domains that are thought to mediate temporal and spatial organization of cellular processes. Sterols are crucial for execution of cytokinesis, the last stage of cell division, in diverse eukaryotes. The cell plate of higher-plant cells is the membrane structure that separates daughter cells during somatic cytokinesis. Cell-plate formation in Arabidopsis relies on sterol- and DYNAMIN-RELATED PROTEIN1A (DRP1A)-dependent endocytosis. However, functional relationships between lipid membrane order or lipid packing and endocytic machinery components during eukaryotic cytokinesis have not been elucidated. Using ratiometric live imaging of lipid order-sensitive fluorescent probes, we show that the cell plate of Arabidopsis thaliana represents a dynamic, high lipid-order membrane domain. The cell-plate lipid order was found to be sensitive to pharmacological and genetic alterations of sterol composition. Sterols co-localize with DRP1A at the cell plate, and DRP1A accumulates in detergent-resistant membrane fractions. Modifications of sterol concentration or composition reduce cell-plate membrane order and affect DRP1A localization. Strikingly, DRP1A function itself is essential for high lipid order at the cell plate. Our findings provide evidence that the cell plate represents a high lipid-order domain, and pave the way to explore potential feedback between lipid order and function of dynamin-related proteins during cytokinesis.

  6. Optimization of protein fractionation by skim milk microfiltration: Choice of ceramic membrane pore size and filtration temperature.

    PubMed

    Jørgensen, Camilla Elise; Abrahamsen, Roger K; Rukke, Elling-Olav; Johansen, Anne-Grethe; Schüller, Reidar B; Skeie, Siv B

    2016-08-01

    The objective of this study was to investigate how ceramic membrane pore size and filtration temperature influence the protein fractionation of skim milk by cross flow microfiltration (MF). Microfiltration was performed at a uniform transmembrane pressure with constant permeate flux to a volume concentration factor of 2.5. Three different membrane pore sizes, 0.05, 0.10, and 0.20µm, were used at a filtration temperature of 50°C. Furthermore, at pore size 0.10µm, 2 different filtration temperatures were investigated: 50 and 60°C. The transmission of proteins increased with increasing pore size, giving the permeate from MF with the 0.20-µm membrane a significantly higher concentration of native whey proteins compared with the permeates from the 0.05- and 0.10-µm membranes (0.50, 0.24, and 0.39%, respectively). Significant amounts of caseins permeated the 0.20-µm membrane (1.4%), giving a permeate with a whitish appearance and a casein distribution (αS2-CN: αS1-CN: κ-CN: β-CN) similar to that of skim milk. The 0.05- and 0.10-µm membranes were able to retain all caseins (only negligible amounts were detected). A permeate free from casein is beneficial in the production of native whey protein concentrates and in applications where transparency is an important functional characteristic. Microfiltration of skim milk at 50°C with the 0.10-µm membrane resulted in a permeate containing significantly more native whey proteins than the permeate from MF at 60°C. The more rapid increase in transmembrane pressure and the significantly lower concentration of caseins in the retentate at 60°C indicated that a higher concentration of caseins deposited on the membrane, and consequently reduced the native whey protein transmission. Optimal protein fractionation of skim milk into a casein-rich retentate and a permeate with native whey proteins were obtained by 0.10-µm MF at 50°C.

  7. Interaction of capsaicinoids with cell membrane models does not correlate with pungency of peppers

    NASA Astrophysics Data System (ADS)

    Geraldo, Vananélia P. N.; Ziglio, Analine C.; Gonçalves, Débora; Oliveira, Osvaldo N.

    2017-04-01

    Mixed monolayers were prepared using phospholipids in order to mimic cell membranes and fractions of capsaicinoids (extracted from Malagueta, Caps-M, and Bhut Jolokia, Caps-B, peppers). According to their surface-pressure isotherms and polarization-modulated infrared reflection absorption spectra (PM-IRRAS), weak molecular-level interactions were observed between Caps and phospholipids. Both Caps-M and Caps-B penetrated into the alkyl tail region of the monolayer, interacted with the phosphate group of the phospholipids and affected hydration of their Cdbnd O groups. Since the physiological activity of Caps is not governed solely by interaction with cell membranes, it should require participation of a neuronal membrane receptor, e.g. vanilloid receptor (TRPV1).

  8. Direct measurements of membrane potential and membrane resistance of human red cells

    PubMed Central

    Lassen, U. V.; Sten-Knudsen, O.

    1968-01-01

    1. In order to evaluate the membrane potentials calculated from the distribution of chloride ions in human red cells and plasma, it is desirable to have a direct measurement of the transmembrane potential of these cells. 2. A method has been devised for introducing a capillary micro-electrode into human red cells. The method allows simultaneous measurements of potential and membrane resistance with only one micro-electrode located in the cell. 3. Upon impalement of single cells in plasma, a scatter of membrane potentials and of resistance values was obtained. The potential drop never exceeded -14 mV and the maximum resistances were about 7 Ω. cm2. Positive potentials were obtained on impalement of red cell aggregates. 4. Arguments are given to support the view that it is in these cells which suffer least damage from the impalement that maximum values of membrane potentials and resistances are observed. The errors caused by the change in the liquid junction during the impalement have been estimated. 5. As judged from this study, it seems permissible under normal conditions to calculate the membrane potential of the red cell from the chloride concentrations in plasma and in intracellular water. PMID:5649641

  9. Radiation Interaction with Therapeutic Drugs and Cell Membranes

    SciTech Connect

    Martin, Diana I.; Manaila, Elena N.; Matei, Constantin I.; Iacob, Nicusor I.; Ighigeanu, Daniel I.; Craciun, Gabriela D.; Moisescu, Mihaela I.; Savopol, Tudor D.; Kovacs, Eugenia A.; Cinca, Sabin A.; Margaritescu, Irina D.

    2007-04-23

    This transient permeabilized state of the cell membrane, named the 'cell electroporation' (CE) can be used to increase cells uptake of drugs that do not readily pass cell membrane, thus enabling their cytotoxicity. The anticancer drugs, such as bleomycin (BL) and cisplatin, are the most candidates for the combined use with ionizing and non-ionizing radiation fields. The methods and installations for the cell electroporation by electron beam (EB) and microwave (MW) irradiation are presented. The viability tests of the human leukocytes under EB and MW exposure with/without the BL in the cell cultures are discussed.

  10. Heterogeneity of Arabinogalactan-Proteins on the Plasma Membrane of Rose Cells.

    PubMed Central

    Serpe, M. D.; Nothnagel, E. A.

    1996-01-01

    Arabinogalactan-proteins (AGPs) have been purified from the plasma membrane of suspension-cultured Paul's Scarlet rose (Rosa sp.) cells. The two most abundant and homogeneous plasma membrane AGP fractions were named plasma membrane AGP1 (PM-AGP1) and plasma membrane AGP2 (PM-AGP2) and had apparent molecular masses of 140 and 217 kD, respectively. Both PM-AGP1 and PM-AGP2 had [beta]-(1-3)-, [beta]-(1,6)-, and [beta]-(1,3,6)-galactopyranosyl residues, predominantly terminal [alpha]-arabinofuranosyl residues, and (1,4)- and terminal glucuronopyranosyl residues. The protein moieties of PM-AGP1 and PM-AGP2 were both rich in hydroxyproline, alanine, and serine, but differed in the abundance of hydroxyproline, which was 1.6 times higher in PM-AGP2 than in PM-AGP1. Another difference was the overall protein content, which was 3.7% (w/w) in PM-AGP1 and 15% in PM-AGP2. As judged by their behavior on reverse-phase chromatography, PM-AGP1 and PM-AGP2 were not more hydrophobic than AGPs from the cell wall or culture medium. In contrast, a minor plasma membrane AGP fraction eluted later on reverse-phase chromatography and was more negatively charged at pH 5 than either PM-AGP1 or PM-AGP2. The more negatively charged fraction contained molecules with a glycosyl composition characteristic of AGPs and included at least two different macromolecules. The results of this investigation indicate that Rosa plasma membrane contains at least four distinct AGPs or AGP-like molecules. These molecules differed from each other in size, charge, hydrophobicity, amino-acyl composition, and/or protein content. PMID:12226444

  11. Investigating cell membrane structure and dynamics with TCSPC-FLIM

    NASA Astrophysics Data System (ADS)

    Le Marois, Alix; Owen, Dylan M.; Suhling, Klaus

    2015-03-01

    We report the use of Time-Correlated Single Photon Counting (TCSPC) in a polarization-resolved Fluorescence Lifetime Imaging (FLIM) setup for the investigation of cell membrane structural and dynamic properties. This technique allows us to study the orientation and mobility of fluorescent membrane dyes, namely di-4-ANEPPDHQ and DiO, in model bilayers of different lipid compositions. Dipole alignment and extent of rotational motion can be linked to membrane order and fluidity. Comparison of the time-resolved anisotropy decays of the two fluorescent dyes suggests that rotational motion of membrane constituents is restricted in liquid-ordered phases, and appears to be limited to the region of aliphatic tails in liquid-disordered phases. In living cells, understanding the membrane structure provides crucial information on its functional properties, such as exo- and endocytosis, cell mobility and signal transduction.

  12. Automated membrane test cell apparatus and method for so using

    SciTech Connect

    Yeager, H.L.; Malinsky, J.D.

    1984-11-20

    An automated electrolytic membrane test cell apparatus adaptable for the purpose of accurately measuring cationic transport and water transport numbers for membranes used in chlor-alkali cells under operating conditions similar to those used in such cells is disclosed. The apparatus comprises a test cell, said test cell being adapted to hold a permselective membrane sealingly supported therein so as to create separate anode and cathode compartments, each of said compartments having a suitable electrode, and heating electrolyte inlet and outlet means attached thereto. The apparatus further comprises means to select one of a plurality of anolyte and catholyte test solutions and control means adapted to control the electrolysis, circulation and heating of said solutions and the generation of all test samples needed to perform the measurements necessary to calculate said transport numbers. When used in conjunction with radioactive tracer techniques, considerably improvements are possible in the accuracy and ease with which transport phenomena in said membrane can be studied.

  13. Single cell wound generates electric current circuit and cell membrane potential variations that requires calcium influx.

    PubMed

    Luxardi, Guillaume; Reid, Brian; Maillard, Pauline; Zhao, Min

    2014-07-24

    Breaching of the cell membrane is one of the earliest and most common causes of cell injury, tissue damage, and disease. If the compromise in cell membrane is not repaired quickly, irreversible cell damage, cell death and defective organ functions will result. It is therefore fundamentally important to efficiently repair damage to the cell membrane. While the molecular aspects of single cell wound healing are starting to be deciphered, its bio-physical counterpart has been poorly investigated. Using Xenopus laevis oocytes as a model for single cell wound healing, we describe the temporal and spatial dynamics of the wound electric current circuitry and the temporal dynamics of cell membrane potential variation. In addition, we show the role of calcium influx in controlling electric current circuitry and cell membrane potential variations. (i) Upon wounding a single cell: an inward electric current appears at the wound center while an outward electric current is observed at its sides, illustrating the wound electric current circuitry; the cell membrane is depolarized; calcium flows into the cell. (ii) During cell membrane re-sealing: the wound center current density is maintained for a few minutes before decreasing; the cell membrane gradually re-polarizes; calcium flow into the cell drops. (iii) In conclusion, calcium influx is required for the formation and maintenance of the wound electric current circuitry, for cell membrane re-polarization and for wound healing.

  14. Membrane-electrode assemblies for electrochemical cells

    DOEpatents

    Swathirajan, Sundararajan; Mikhail, Youssef M.

    1993-01-01

    A combination, unitary, membrane and electrode assembly with a solid polymer electrolyte membrane, and first and second electrodes at least partially embedded in opposed surfaces of the membrane. The electrodes each comprise a respective group of finely divided carbon particles, very finely divided catalytic particles supported on internal and external surfaces of the carbon particles and a proton conductive material intermingled with the catalytic and carbon particles. A first group of finely divided carbon particles forming the first electrode has greater water attraction and retention properties, and is more hydrophilic than a second group of carbon particles forming the second electrode. In a preferred method, the membrane electrode assembly of the invention is prepared by forming a slurry of proton conductive material and at least one group of the carbon and catalyst particles. The slurry is applied to the opposed surfaces of the membrane and heated while being pressed to the membrane for a time and at a temperature and compressive load sufficient to embed at least a portion of the particles into the membrane.

  15. Insulin receptor: Interaction with nonreceptor glycoprotein from liver cell membranes

    PubMed Central

    Maturo, Joseph M.; Hollenberg, Morley D.

    1978-01-01

    In crude receptor preparations (either particulate or soluble) of rat liver membranes, the insulin receptor exhibits complicated binding kinetics (two binding plateaus, half-saturated at approximately 60 pM and 700 pM insulin) and an apparent chromatographic heterogeneity, suggested by the presence of two detectable, soluble insulin-binding components with apparent Stokes radii of 72 Å and 38 Å. In contrast, the insulin receptor isolated by affinity chromatography exhibits a simple binding isotherm (half-maximal saturation of binding at 700 pM insulin) without evidence for negative cooperativity and behaves as a single component (apparent Stokes radius of 38 Å) upon chromatography on Sepharose 6B. The apparent discrepancies between the properties of the unpurified insulin receptor and the affinity-purified receptor can be attributed to the presence in crude preparations of a nonreceptor constituent(s) having properties consistent with those of a membrane glycoprotein. A glycoprotein fraction from such crude soluble membrane preparations, freed from insulin receptor and subsequently partially purified using concanavalin-A-agarose, when combined with affinity-purified insulin receptor, causes both a reappearance of the complicated binding kinetics and an increase in the receptor's apparent Stokes radius from 38 Å to 72 Å. Similar results are observed for a glycoprotein fraction obtained from rat adipocyte membranes but are not observed for an identical fraction isolated from human erythrocyte membranes. We conclude that the insulin receptor in rat liver membranes can interact with another nonreceptor membrane glycoprotein that may represent either a nonrecognition moiety of the receptor oligomer or an effector molecule to the biological action of insulin. PMID:277909

  16. Prism-patterned Nafion membrane for enhanced water transport in polymer electrolyte membrane fuel cell

    NASA Astrophysics Data System (ADS)

    Kim, Sang Moon; Kang, Yun Sik; Ahn, Chiyeong; Jang, Segeun; Kim, Minhyoung; Sung, Yung-Eun; Yoo, Sung Jong; Choi, Mansoo

    2016-06-01

    Here, we report a simple and effective strategy to enhance the performance of the polymer electrolyte membrane fuel cell by imprinting prism-patterned arrays onto the Nafion membrane, which provides three combined effects directly related to the device performance. First, a locally thinned membrane via imprinted micro prism-structures lead to reduced membrane resistance, which is confirmed by electrochemical impedance spectroscopy. Second, increments of the geometrical surface area of the prism-patterned Nafion membrane compared to a flat membrane result in the increase in the electrochemical active surface area. Third, the vertically asymmetric geometry of prism structures in the cathode catalyst layer lead to enhanced water transport, which is confirmed by oxygen gain calculation. To explain the enhanced water transport, we propose a simple theoretical model on removal of water droplets existing in the asymmetric catalyst layer. These three combined effects achieved via incorporating prism patterned arrays into the Nafion membrane effectively enhance the performance of the polymer electrolyte membrane fuel cell.

  17. Membranes replace irradiated blast cells as growth requirement for leukemic blast progenitors in suspension culture

    SciTech Connect

    Nara, N.; McCulloch, E.A.

    1985-11-01

    The blast cells of acute myeloblastic leukemia (AML) may be considered as a renewal population, maintained by blast stem cells capable of both self-renewal and the generation of progeny with reduced or absent proliferative potential. This growth requires that two conditions be met: first, the cultures must contain growth factors in media conditioned either by phytohemagglutinin (PHA)-stimulated mononuclear leukocytes (PHA-LCM), or by cells of the continuous bladder carcinoma line HTB9 (HTB9-CM). Second, the cell density must be maintained at 10(6) blasts/ml; this may be achieved by adding irradiated cells to smaller numbers of intact blasts. The authors are concerned with the mechanism of the feeding function. They present evidence that (a) cell-cell contact is required. (b) Blasts are heterogeneous in respect to their capacity to support growth. (c) Fractions containing membranes from blast cells will substitute for intact cells in promoting the generation of new blast progenitors in culture. (d) This membrane function may be specific for AML blasts, since membranes from blasts of lymphoblastic leukemia or normal marrow cells were inactive.

  18. Adaptation of yeast cell membranes to ethanol

    SciTech Connect

    Jimenez, J.; Benitez, T.

    1987-05-01

    A highly ethanol-tolerant Saccharomyces wine strain is able, after growth in the presence of ethanol, to efficiently improve the ethanol tolerance of its membrane. A less-tolerant Saccharomyces laboratory strain, however, is unable to adapt its membrane to ethanol. Furthermore, after growth in the presence of ethanol, the membrane of the latter strain becomes increasingly sensitive, although this is a reversible process. Reversion to a higher tolerance occurs only after the addition of an energy source and does not take place in the presence of cycloheximide.

  19. Catalytic membranes for CO oxidation in fuel cells

    DOEpatents

    Sandi-Tapia, Giselle; Carrado Gregar, Kathleen; Kizilel, Riza

    2010-06-08

    A hydrogen permeable membrane, which includes a polymer stable at temperatures of about 200 C having clay impregnated with Pt or Au or Ru or Pd particles or mixtures thereof with average diameters of less than about 10 nanometers (nms) is disclosed. The membranes are useful in fuel cells or any device which requires hydrogen to be separated from carbon monoxide.

  20. Expression of basement membrane antigens in spindle cell melanoma.

    PubMed

    Prieto, V G; Woodruff, J M

    1998-07-01

    Spindle cell melanoma (SCM) is an uncommon form of melanoma that may be confused histologically with other tumors, including malignant peripheral nerve sheath tumors (MPNST). Tumors with neural differentiation and melanocytic nevi may both show basement membrane immunohistochemically and at the ultrastructural level. However, most ultrastructural studies of melanoma have failed to demonstrate well formed basement membrane around tumor cells. The presence of basement membrane has been used by some authors as evidence favoring MPNST, as opposed to SCM. To evaluate this distinction immunohistochemically, 22 primary and metastatic cutaneous melanomas having a spindle cell component (SCM) were studied using monoclonal antibodies against laminin and Type IV collagen. S100 protein and HMB45 antigen expression were also studied. All but one of the SCM were reactive for S100 protein in at least 25% of the cells. Thirteen of 20 tumors (65%) were focally reactive with HMB45. Laminin was expressed in 42% of the tumors (only membranous pattern in 3; cytoplasmic and membranous in 5). Seventeen tumors (77%) expressed type IV collagen (only membranous pattern in 7; cytoplasmic and membranous pattern in 10). Laminin and type IV collagen, known components of basement membrane, are often found in SCM. Therefore, their detection cannot be used to distinguish SCM from MPNST.

  1. Cell-Cell Communication Via Extracellular Membrane Vesicles and Its Role in the Immune Response

    PubMed Central

    Hwang, Inkyu

    2013-01-01

    The host immune response involves a variety of cell types, including specialized immune and non-immune cells. The delicate coordination among these cells via close communication is central for the proper operation of immune system. Cell-cell communication is mediated by a complex network that includes soluble factors such as cytokines, chemokines, and metabolites exported from cells, as well as membrane-bound receptors and their ligands. Cell-cell communication is also mediated by membrane vesicles (e.g., exosomes, ectosomes), which are either shed by distant cells or exchanged by cells that are making direct contact. Intercellular communication via extracellular membrane vesicles has drawn much attention recently, as they have been shown to carry various biomolecules that modulate the activities of recipient cells. In this review, I will discuss current views on cell-cell communication via extra-cellular membrane vesicles, especially shedded membrane vesicles, and their effects on the control of the immune system. PMID:23807045

  2. Monitoring of lung tumour cell growth in artificial membranes.

    PubMed

    Yang, Ying; Sulé-Suso, Josep; El Haj, Alicia J; Hoban, Paul R; Wang, Ruikang

    2004-10-15

    Morbidity of many tumour types is associated with invasion of tumour cells through the basement membrane and subsequent metastasis to vital organs. Tumour invasion is frequently detected late on as many patients present with advanced disease. The method of detecting invasion is through conventional histological staining techniques, which are time consuming and require processing of the sample. This can affect interpretation of the results. In this study, a new imaging technique, optical coherence tomography (OCT), was used to monitor lung tumour cell growth in two artificial membranes composed of either collagen type I or Matrigel. In parallel, standard histological section analysis was performed to validate the accuracy of the monitoring by OCT. Cross-sectional images from OCT revealed that lung tumour cells infiltrated only when low cell seeding density (5 x 10(5)) and low collagen concentration (1.5 mg/ml) were combined. The cells could be easily differentiated from the artificial membranes and appeared as either a brighter layer on the top of the membrane or brighter foci embedded within the darker membrane. These cell-membrane morphologies matched remarkably to the standard histological section images. Our results suggest that OCT has a great potential to become a useful tool for fast and robust imaging of cell growth in vivo and as a potential assessment of cell invasion.

  3. Sulfated Titania-Silica Reinforced Nafion Nanocomposite Membranes for Proton Exchange Membrane Fuel Cells.

    PubMed

    Abu Sayeed, M D; Kim, Hee Jin; Gopalan, A I; Kim, Young Ho; Lee, Kwang-Pill; Choi, Sang-June

    2015-09-01

    Sulfated titania-silica (SO4(2-)-/TiO2-SiO2) composites were prepared by a sol-gel method with sulfate reaction and characterized by X-ray diffraction (XRD) and energy-dispersive X-ray spectroscopy (EDS). The nanometric diameter and geometry of the sulfated titania-silica (STS) was investigated by transmission electron microscopy (TEM). A small amount of the STS composite in the range of 0.5-3 wt% was then added as reinforcing into the Nafion membrane by water-assisted solution casting method to prepare STS reinforced Nafion nanocomposite membranes (STS-Nafion nanocomposite membranes). The additional functional groups, sulfate groups, of the nanocomposite membrane having more surface oxygenated groups enhanced the fuel cell membrane properties. The STS-Nafion nanocomposite membranes exhibited improved water uptake compared to that of neat Nafion membranes, whereas methanol uptake values were decreased dramatically improved thermal property of the prepared nanocomposite membranes were measured by thermogravimetric analysis (TGA). Furthermore, increased ion exchange capacity values were obtained by thermoacidic pretreatment of the nanocomposite membranes.

  4. CO2 permeability of cell membranes is regulated by membrane cholesterol and protein gas channels.

    PubMed

    Itel, Fabian; Al-Samir, Samer; Öberg, Fredrik; Chami, Mohamed; Kumar, Manish; Supuran, Claudiu T; Deen, Peter M T; Meier, Wolfgang; Hedfalk, Kristina; Gros, Gerolf; Endeward, Volker

    2012-12-01

    Recent observations that some membrane proteins act as gas channels seem surprising in view of the classical concept that membranes generally are highly permeable to gases. Here, we study the gas permeability of membranes for the case of CO(2), using a previously established mass spectrometric technique. We first show that biological membranes lacking protein gas channels but containing normal amounts of cholesterol (30-50 mol% of total lipid), e.g., MDCK and tsA201 cells, in fact possess an unexpectedly low CO(2) permeability (P(CO2)) of ∼0.01 cm/s, which is 2 orders of magnitude lower than the P(CO2) of pure planar phospholipid bilayers (∼1 cm/s). Phospholipid vesicles enriched with similar amounts of cholesterol also exhibit P(CO2) ≈ 0.01 cm/s, identifying cholesterol as the major determinant of membrane P(CO2). This is confirmed by the demonstration that MDCK cells depleted of or enriched with membrane cholesterol show dramatic increases or decreases in P(CO2), respectively. We demonstrate, furthermore, that reconstitution of human AQP-1 into cholesterol-containing vesicles, as well as expression of human AQP-1 in MDCK cells, leads to drastic increases in P(CO2), indicating that gas channels are of high functional significance for gas transfer across membranes of low intrinsic gas permeability.

  5. Ultrafiltration by a compacted clay membrane-I. Oxygen and hydrogen isotopic fractionation

    USGS Publications Warehouse

    Coplen, T.B.; Hanshaw, B.B.

    1973-01-01

    Laboratory experiments were carried out to determine the magnitude of the isotopic fractionation of distilled water and of 0.01 N NaCl forced to flow at ambient temperature under a hydraulic pressure drop of 100 bars across a montmorillonite disc compacted to a porosity of 35 per cent by a pressure of 330 bars. The ultrafiltrates in both experiments were depleted in D by 2.5%. and in O18 by 0.8%. relative to the residual solution. No additional isotopic fractionation due to a salt filtering mechanism was observed at NaCl concentrations up to 0.01 N. Adsorption is most likely the principal mechanism which produces isotopic fractionation, but molecular diffusion may play a minor role. The results suggest that oxygen and hydrogen isotopic fractionation of ground water during passage through compacted clayey sediments should be a common occurrence, in accord with published interpretations of isotopic data from the Illinois and Alberta basins. ?? 1973.

  6. Cell-free system for synthesizing membrane proteins cell free method for synthesizing membrane proteins

    DOEpatents

    Laible, Philip D; Hanson, Deborah K

    2013-06-04

    The invention provides an in vitro method for producing proteins, membrane proteins, membrane-associated proteins, and soluble proteins that interact with membrane-associated proteins for assembly into an oligomeric complex or that require association with a membrane for proper folding. The method comprises, supplying intracytoplasmic membranes from organisms; modifying protein composition of intracytoplasmic membranes from organism by modifying DNA to delete genes encoding functions of the organism not associated with the formation of the intracytoplasmic membranes; generating appropriate DNA or RNA templates that encode the target protein; and mixing the intracytoplasmic membranes with the template and a transcription/translation-competent cellular extract to cause simultaneous production of the membrane proteins and encapsulation of the membrane proteins within the intracytoplasmic membranes.

  7. Low Crossover Polymer Electrolyte Membranes for Direct Methanol Fuel Cells

    NASA Technical Reports Server (NTRS)

    Prakash, G. K. Surya; Smart, Marshall; Atti, Anthony R.; Olah, George A.; Narayanan, S. R.; Valdez, T.; Surampudi, S.

    1996-01-01

    Direct Methanol Fuel Cells (DMFC's) using polymer electrolyte membranes are promising power sources for portable and vehicular applications. State of the art technology using Nafion(R) 117 membranes (Dupont) are limited by high methanol permeability and cost, resulting in reduced fuel cell efficiencies and impractical commercialization. Therefore, much research in the fuel cell field is focused on the preparation and testing of low crossover and cost efficient polymer electrolyte membranes. The University of Southern California in cooperation with the Jet Propulsion Laboratory is focused on development of such materials. Interpenetrating polymer networks are an effective method used to blend polymer systems without forming chemical links. They provide the ability to modify physical and chemical properties of polymers by optimizing blend compositions. We have developed a novel interpenetrating polymer network based on poly (vinyl - difluoride)/cross-linked polystyrenesulfonic acid polymer composites (PVDF PSSA). Sulfonation of polystyrene accounts for protonic conductivity while the non-polar, PVDF backbone provides structural integrity in addition to methanol rejection. Precursor materials were prepared and analyzed to characterize membrane crystallinity, stability and degree of interpenetration. USC JPL PVDF-PSSA membranes were also characterized to determine methanol permeability, protonic conductivity and sulfur distribution. Membranes were fabricated into membrane electrode assemblies (MEA) and tested for single cell performance. Tests include cell performance over a wide range of temperatures (20 C - 90 C) and cathode conditions (ambient Air/O2). Methanol crossover values are measured in situ using an in-line CO2 analyzer.

  8. Selectivity of biopolymer membranes using HepG2 cells.

    PubMed

    Lü, Dongyuan; Gao, Yuxin; Luo, Chunhua; Lü, Shouqian; Wang, Qian; Xu, Xianghong; Sun, Shujin; Wang, Chengzhi; Long, Mian

    2015-03-01

    Bioartificial liver (BAL) system has emerged as an alternative treatment to bridge acute liver failure to either liver transplantation or liver regeneration. One of the main reasons that the efficacy of the current BAL systems was not convincing in clinical trials is attributed to the lack of friendly interface between the membrane and the hepatocytes in liver bioreactor, the core unit of BAL system. Here, we systematically compared the biological responses of hepatosarcoma HepG2 cells seeded on eight, commercially available biocompatible membranes made of acetyl cellulose-nitrocellulose mixed cellulose (CA-NC), acetyl cellulose (CA), nylon (JN), polypropylene (PP), nitrocellulose (NC), polyvinylidene fluoride (PVDF), polycarbonate (PC) and polytetrafluoroethylene (PTFE). Physicochemical analysis and mechanical tests indicated that CA, JN and PP membranes yield high adhesivity and reasonable compressive and/or tensile features with friendly surface topography for cell seeding. Cells prefer to adhere on CA, JN, PP or PTFE membranes with high proliferation rate in spheriod-like shape. Actin, albumin and cytokeratin 18 expressions are favorable for cells on CA or PP membrane, whereas protein filtration is consistent among all the eight membranes. These results further the understandings of cell growth, morphology and spreading, as well as protein filtration on distinct membranes in designing a liver bioreactor.

  9. Drug Delivery via Cell Membrane Fusion Using Lipopeptide Modified Liposomes

    PubMed Central

    2016-01-01

    Efficient delivery of drugs to living cells is still a major challenge. Currently, most methods rely on the endocytotic pathway resulting in low delivery efficiency due to limited endosomal escape and/or degradation in lysosomes. Here, we report a new method for direct drug delivery into the cytosol of live cells in vitro and invivo utilizing targeted membrane fusion between liposomes and live cells. A pair of complementary coiled-coil lipopeptides was embedded in the lipid bilayer of liposomes and cell membranes respectively, resulting in targeted membrane fusion with concomitant release of liposome encapsulated cargo including fluorescent dyes and the cytotoxic drug doxorubicin. Using a wide spectrum of endocytosis inhibitors and endosome trackers, we demonstrate that the major site of cargo release is at the plasma membrane. This method thus allows for the quick and efficient delivery of drugs and is expected to have many invitro, ex vivo, and invivo applications. PMID:27725960

  10. Decreasing Outer Hair Cell Membrane Cholesterol Increases Cochlear Electromechanics

    NASA Astrophysics Data System (ADS)

    Brownell, William E.; Jacob, Stefan; Hakizimana, Pierre; Ulfendahl, Mats; Fridberger, Anders

    2011-11-01

    The effect of decreasing membrane cholesterol on the mechanical response of the cochlea to acoustic and/or electrical stimulation was monitored using laser interferometry. In contrast to pharmacological interventions that typically decrease cochlear electromechanics, reducing membrane cholesterol increased the response. The electromechanical response in untreated preparations was asymmetric with greater displacements in response to positive currents and cholesterol depletion increased the asymmetry. The results confirm that outer hair cell electromotility is enhanced by low membrane cholesterol. The asymmetry of the response indicates the outer hair cell resting membrane potential is hyperpolarized relative to the voltage of maximum gain for the outer hair cell voltage-displacement function. The magnitude of the response increase suggests a non-uniform distribution of cholesterol along the lateral wall of normal adult outer hair cells.

  11. Apparatus measures swelling of membranes in electrochemical cells

    NASA Technical Reports Server (NTRS)

    Hennigan, T. J.

    1965-01-01

    Apparatus consisting of a pressure plate unit, four springs of known spring constant and a micrometer measures the swelling and force exerted by the polymer membranes of alkaline electrochemical cells.

  12. The Antioxidative Fraction of White Mulberry Induces Apoptosis through Regulation of p53 and NFκB in EAC Cells

    PubMed Central

    Khan, Muhammad Ali; Kabir, Syed Rashel; Reza, Md Abu; Rahman, Md Mahbubur; Islam, Mohammad Saiful; Rahman, Md Aziz Abdur; Rashid, Mamunur; Sadik, Md Golam

    2016-01-01

    In this study, the antioxidative fraction of white mulberry (Morus alba) was found to have an apotogenic effect on Ehrlich’s ascites carcinoma cell-induced mice (EAC mice) that correlate with upregulated p53 and downregulated NFκB signaling. The antioxidant activities and polyphenolic contents of various mulberry fractions were evaluated by spectrophotometry and the ethyl acetate fraction (EAF) was selected for further analysis. Strikingly, the EAF caused 70.20% tumor growth inhibition with S-phase cell cycle arrest, normalized blood parameters including red/white blood cell counts and suppressed the tumor weight of EAC mice compared with untreated controls. Fluorescence microscopy analysis of EAF-treated EAC cells revealed DNA fragmentation, cell shrinkage, and plasma membrane blebbing. These characteristic morphological features of apoptosis influenced us to further investigate pro- and anti-apoptotic signals in EAF-treated EAC mice. Interestingly, apoptosis correlated with the upregulation of p53 and its target genes PARP-1 and Bax, and also with the down-regulation of NFκB and its target genes Bcl-2 and Bcl-xL. Our results suggest that the tumor- suppressive effect of the antioxidative fraction of white mulberry is likely due to apoptosis mediated by p53 and NFκB signaling. PMID:27936037

  13. Differential effects of lipid fractions from silver carp brain on human cervical carcinoma cells in vitro.

    PubMed

    Wang, Caixia; Xia, Wenshui; Jiang, Qixing; Xu, Yanshun; Yu, Peipei

    2014-09-01

    Previous research has revealed that n3 polyunsaturated fatty acids (PUFAs) exhibit anticancer activities. Lipids from a fish brain contain substantial n3 PUFAs. However, no research has been conducted on the action and mechanism of their potent anticancer activities. In this study, total lipids (TLs) from silver carp brain were isolated into polar lipids (PLs) and neutral lipids (NLs), and the anticancer potential of the lipid fractions (LFs) was investigated using the human cervical carcinoma HeLa cell line. LFs effectively inhibited the cell proliferation of HeLa cells in a time- and dose-dependent manner by cell cycle arrest at the S stage and by inducing apoptosis. Further analyses indicated that the loss of mitochondrial membrane potential could be one of mechanisms of apoptosis induced by LFs. Among the TLs, PLs have proven to be more effective in inducing cervical carcinoma cell death than NLs. This work will play a role in promoting lipids from silver carp brain as a potential preventive and therapeutic agent against human cervical carcinoma.

  14. Membrane Composition Tunes the Outer Hair Cell Motor

    NASA Astrophysics Data System (ADS)

    Rajagopalan, L.; Sfondouris, J.; Oghalai, J. S.; Pereira, F. A.; Brownell, W. E.

    2009-02-01

    Cholesterol and docosahexaenoic acid (DHA), an ω-3 fatty acid, affect membrane mechanical properties in different ways and modulate the function of membrane proteins. We have probed the functional consequence of altering cholesterol and DHA levels in the membranes of OHCs and prestin expressing HEK cells. Large, dynamic and reversible changes in prestin-associated charge movement and OHC motor activity result from altering the concentration of membrane cholesterol. Increasing membrane cholesterol shifts the q/V function ~ 50 mV in the hyperpolarizing direction, possibly a response related to increases in membrane stiffness. The voltage shift is linearly related to total membrane cholesterol. Increasing cholesterol also decreases the total charge moved in a linear fashion. Decreasing membrane cholesterol shifts the q/V function ~ 50 mV in the depolarizing direction with little or no effect on the amount of charge moved. In vivo increases in membrane cholesterol transiently increase but ultimately lead to decreases in DPOAE. Docosahexaenoic acid shifts the q/V function in the hyperpolarizing direction < 15 mV and increases total charge moved. Tuning of cochlear function by membrane cholesterol contributes to the exquisite temporal and frequency processing of mammalian hearing by optimizing the cochlear amplifier.

  15. Penetration of Cell Membranes and Synthetic Lipid Bilayers by Nanoprobes

    PubMed Central

    Angle, Matthew R.; Wang, Andrew; Thomas, Aman; Schaefer, Andreas T.; Melosh, Nicholas A.

    2014-01-01

    Nanoscale devices have been proposed as tools for measuring and controlling intracellular activity by providing electrical and/or chemical access to the cytosol. Unfortunately, nanostructures with diameters of 50–500 nm do not readily penetrate the cell membrane, and rationally optimizing nanoprobes for cell penetration requires real-time characterization methods that are capable of following the process of membrane penetration with nanometer resolution. Although extensive work has examined the rupture of supported synthetic lipid bilayers, little is known about the applicability of these model systems to living cell membranes with complex lipid compositions, cytoskeletal attachment, and membrane proteins. Here, we describe atomic force microscopy (AFM) membrane penetration experiments in two parallel systems: live HEK293 cells and stacks of synthetic lipid bilayers. By using the same probes in both systems, we were able to clearly identify membrane penetration in synthetic bilayers and compare these events with putative membrane penetration events in cells. We examined membrane penetration forces for three tip geometries and 18 chemical modifications of the probe surface, and in all cases the median forces required to penetrate cellular and synthetic lipid bilayers with nanoprobes were greater than 1 nN. The penetration force was sensitive to the probe's sharpness, but not its surface chemistry, and the force did not depend on cell surface or cytoskeletal properties, with cells and lipid stacks yielding similar forces. This systematic assessment of penetration under various mechanical and chemical conditions provides insights into nanoprobe-cell interactions and informs the design of future intracellular nanoprobes. PMID:25418094

  16. Effect of Chemicals on the Cell Membrane Transport of Nucleosides.

    DTIC Science & Technology

    1983-08-01

    lipid synthesis , a direct inhibition of the purine carrier by PFDA would not be expected. When efflux of AP from L5178Y cells was estimated with PFDA in...turnover of the carrier protein. PFDA may be an inhibitor -of carrier protein synthesis in the cell membrane. Another hypothesis suggests that the...inactive form. The activity level *may be controlled through inhibition of protein synthesis or the interaction 4between the carrier and the membrane

  17. Identification of a Zn(2+)-sensitive component of Ehrlich cell plasma membrane redox system by CHAPS-agarose-polyacrylamide electrophoresis and in situ staining of activity.

    PubMed

    Rodríguez-Caso, L; Rodríguez-Agudo, D; del Castillo-Olivares, A; Márquez, J; Núñez de Castro, I; Medina, M A

    1997-01-01

    A procedure based on CHAPS-agarose-polyacrylamide electrophoresis and in situ staining of activity was used to detect a Zn(2+)-sensitive component of Ehrlich cell plasma membrane redox system. The procedure is so powerful that it allows to use crude plasma membrane fractions and can be easily adapted for use in an electrophoretic approach to the purification of this protein.

  18. Live cell imaging of membrane/cytoskeleton interactions and membrane topology.

    PubMed

    Chierico, Luca; Joseph, Adrian S; Lewis, Andrew L; Battaglia, Giuseppe

    2014-09-10

    We elucidate the interaction between actin and specific membrane components, using real time live cell imaging, by delivering probes that enable access to components, that cannot be accessed genetically. We initially investigated the close interplay between Phosphatidylinositol 4,5-bisphosphate (PIP2) and the F-actin network. We show that, during the early stage of cell adhesion, PIP2 forms domains within the filopodia membrane. We studied these domains alongside cell spreading and observed that these very closely follow the actin tread-milling. We show that this mechanism is associated with an active transport of PIP2 rich organelles from the cell perinuclear area to the edge, along actin fibers. Finally, mapping other phospholipids and membrane components we observed that the PIP2 domains formation is correlated with sphingosine and cholesterol rafts.

  19. Modelling the structure of the red cell membrane.

    PubMed

    Burton, Nicholas M; Bruce, Lesley J

    2011-04-01

    The red cell membrane has long been the focus of extensive study. The macromolecules embedded within the membrane carry the blood group antigens and perform many functions including the vital task of gas exchange. Links between the intramembrane macromolecules and the underlying cytoskeleton stabilize the biconcave morphology of the red cell and allow deformation during microvascular transit. Much is now known about the proteins of the red cell membrane and how they are organised. In many cases we have an understanding of which proteins are expressed, the number of each protein per cell, their oligomeric state(s), and how they are collected in large multi-protein complexes. However, our typical view of these structures is as cartoon shapes in schematic figures. In this study we have combined knowledge of the red cell membrane with a wealth of protein structure data from crystallography, NMR, and homology modelling to generate the first, tentative models of the complexes which link the membrane to the cytoskeleton. Measurement of the size of these complexes and comparison with known cytoskeletal distance parameters suggests the idea of interaction between the membrane complexes, which may have profound implications for understanding red cell function and deformation.

  20. Controlled permeation of cell membrane by single bubble acoustic cavitation.

    PubMed

    Zhou, Y; Yang, K; Cui, J; Ye, J Y; Deng, C X

    2012-01-10

    Sonoporation is the membrane disruption generated by ultrasound and has been exploited as a non-viral strategy for drug and gene delivery. Acoustic cavitation of microbubbles has been recognized to play an important role in sonoporation. However, due to the lack of adequate techniques for precise control of cavitation activities and real-time assessment of the resulting sub-micron process of sonoporation, limited knowledge has been available regarding the detail processes and correlation of cavitation with membrane disruption at the single cell level. In the current study, we developed a combined approach including optical, acoustical, and electrophysiological techniques to enable synchronized manipulation, imaging, and measurement of cavitation of single bubbles and the resulting cell membrane disruption in real-time. Using a self-focused femtosecond laser and high frequency ultrasound (7.44MHz) pulses, a single microbubble was generated and positioned at a desired distance from the membrane of a Xenopus oocyte. Cavitation of the bubble was achieved by applying a low frequency (1.5MHz) ultrasound pulse (duration 13.3 or 40μs) to induce bubble collapse. Disruption of the cell membrane was assessed by the increase in the transmembrane current (TMC) of the cell under voltage clamp. Simultaneous high-speed bright field imaging of cavitation and measurements of the TMC were obtained to correlate the ultrasound-generated bubble activities with the cell membrane poration. The change in membrane permeability was directly associated with the formation of a sub-micrometer pore from a local membrane rupture generated by bubble collapse or bubble compression depending on ultrasound amplitude and duration. The impact of the bubble collapse on membrane permeation decreased rapidly with increasing distance (D) between the bubble (diameter d) and the cell membrane. The effective range of cavitation impact on membrane poration was determined to be D/d=0.75. The maximum mean

  1. Spray deposition of Nafion membranes: Electrode-supported fuel cells

    NASA Astrophysics Data System (ADS)

    Bayer, Thomas; Pham, Hung Cuong; Sasaki, Kazunari; Lyth, Stephen Matthew

    2016-09-01

    Fuel cells are a key technology for the successful transition towards a hydrogen society. In order to accelerate fuel cell commercialization, improvements in performance are required. Generally, polymer electrolyte membrane fuel cells (PEFCs) are membrane-supported; the electrocatalyst layer is sprayed onto both sides of the membrane, and sandwiched between carbon-based gas diffusion layers (GDLs). In this work we redesign the membrane electrode assembly (MEA) and fabricate an electrode-supported PEFC. First the electrocatalyst layer is sprayed onto the GDL, and then Nafion dispersion is sprayed over the top of this to form a thin membrane. This method has the advantage of simplifying the fabrication process, allowing the fabrication of extremely thin electrolyte layers (down to ∼10 μm in this case), and reducing the amount of ionomer required in the cell. Electrode-supported PEFCs operate at significantly increased power density compared to conventional membrane-supported PEFCs, with a maximum of 581 mW/cm2 at 80 °C (atmospheric pressure, air at the cathode). Impedance spectroscopy confirmed that the origin of the improved performance was an 80% reduction in the membrane resistance due the thinner Nafion layer. This novel fabrication method is a step towards cheaper, thinner, fully printable PEFCs with high power density and efficiency.

  2. Proton conducting membranes for high temperature fuel cells with solid state water free membranes

    NASA Technical Reports Server (NTRS)

    Narayanan, Sekharipuram R. (Inventor); Yen, Shiao-Pin S. (Inventor)

    2006-01-01

    A water free, proton conducting membrane for use in a fuel cell is fabricated as a highly conducting sheet of converted solid state organic amine salt, such as converted acid salt of triethylenediamine with two quaternized tertiary nitrogen atoms, combined with a nanoparticulate oxide and a stable binder combined with the converted solid state organic amine salt to form a polymeric electrolyte membrane. In one embodiment the membrane is derived from triethylenediamine sulfate, hydrogen phosphate or trifiate, an oxoanion with at least one ionizable hydrogen, organic tertiary amine bisulfate, polymeric quaternized amine bisulfate or phosphate, or polymeric organic compounds with quaternizable nitrogen combined with Nafion to form an intimate network with ionic interactions.

  3. Catalyst layers for proton exchange membrane fuel cells prepared by electrospray deposition on Nafion membrane

    NASA Astrophysics Data System (ADS)

    Chaparro, A. M.; Ferreira-Aparicio, P.; Folgado, M. A.; Martín, A. J.; Daza, L.

    The electrospray deposition method has been used for preparation of catalyst layers for proton exchange membrane fuel cells (PEMFC) on Nafion membrane. Deposition of Pt/C + ionomer suspensions on Nafion 212 gives rise to layers with a globular morphology, in contrast with the dendritic growth observed for the same layers when deposited on the gas diffusion layer, GDL (microporous carbon black layer on carbon cloth) or on metallic Al foils. Such a change is discussed in the light of the influence of the Nafion substrate on the electrospray deposition process. Nafion, which is a proton conductor and electronic insulator, gives rise to the discharge of particles through proton release and transport towards the counter electrode, compared with the direct electron transfer that takes place when depositing on an electronic conductor. There is also a change in the electric field distribution in the needle to counter-electrode gap due to the presence of Nafion, which may alter conditions for the electrospray effect. If discharging of particles is slow enough, for instances with a low membrane protonic conductivity, the Nafion substrate may be charged positively yielding a change in the electric field profile and, with it, in the properties of the film. Single cell characterization is carried out with Nafion 212 membranes catalyzed by electrospray on the cathode side. It is shown that the internal resistance of the cell decreases with on-membrane deposited cathodic catalyst layers, with respect to the same layers deposited on GDL, giving rise to a considerable improvement in cell performance. The lower internal resistance is due to higher proton conductivity at the catalyst layer-membrane interface resulting from on-membrane deposition. On the other hand, electroactive area and catalyst utilization appear little modified by on-membrane deposition, compared with on-GDL deposition.

  4. Manganese accumulation in membrane fractions of primary astrocytes is associated with decreased γ-aminobutyric acid (GABA) uptake, and is exacerbated by oleic acid and palmitate.

    PubMed

    Fordahl, Steve C; Erikson, Keith M

    2014-05-01

    Manganese (Mn) exposure interferes with GABA uptake; however, the effects of Mn on GABA transport proteins (GATs) have not been identified. We sought to characterize how Mn impairs GAT function in primary rat astrocytes. Astrocytes exposed to Mn (500 μM) had significantly reduced (3)H-GABA uptake despite no change in membrane or cytosolic GAT3 protein levels. Co-treatment with 100 μM oleic or palmitic acids (both known to be elevated in Mn neurotoxicity), exacerbated the Mn-induced decline in (3)H-GABA uptake. Mn accumulation in the membrane fraction of astrocytes was enhanced with fatty acid administration, and was negatively correlated with (3)H-GABA uptake. Furthermore, control cells exposed to Mn only during the experimental uptake had significantly reduced (3)H-GABA uptake, and the addition of GABA (50 μM) blunted cytosolic Mn accumulation. These data indicate that reduced GAT function in astrocytes is influenced by Mn and fatty acids accumulating at or interacting with the plasma membrane.

  5. Membrane Vesicles Released by Intestinal Epithelial Cells Infected with Rotavirus Inhibit T-Cell Function

    PubMed Central

    Barreto, Alfonso; Rodríguez, Luz-Stella; Rojas, Olga Lucía; Wolf, Marie; Greenberg, Harry B.; Franco, Manuel A.

    2010-01-01

    Abstract Rotavirus (RV) predominantly replicates in intestinal epithelial cells (IEC), and “danger signals” released by these cells may modulate viral immunity. We have recently shown that human model IEC (Caco-2 cells) infected with rhesus-RV release a non-inflammatory group of immunomodulators that includes heat shock proteins (HSPs) and TGF-β1. Here we show that both proteins are released in part in association with membrane vesicles (MV) obtained from filtrated Caco-2 supernatants concentrated by ultracentrifugation. These MV express markers of exosomes (CD63 and others), but not of the endoplasmic reticulum (ER) or nuclei. Larger quantities of proteins associated with MV were released by RV-infected cells than by non-infected cells. VP6 co-immunoprecipitated with CD63 present in these MV, and VP6 co-localized with CD63 in RV-infected cells, suggesting that this viral protein is associated with the MV, and that this association occurs intracellularly. CD63 present in MV preparations from stool samples from 36 children with gastroenteritis due or not due to RV were analyzed. VP6 co-immunoprecipitated with CD63 in 3/8 stool samples from RV-infected children, suggesting that these MV are released by RV-infected cells in vivo. Moreover, fractions that contained MV from RV-infected cells induced death and inhibited proliferation of CD4+ T cells to a greater extent than fractions from non-infected cells. These effects were in part due to TGF-β, because they were reversed by treatment of the T cells with the TGF-β-receptor inhibitor ALK5i. MV from RV-infected and non-infected cells were heterogeneous, with morphologies and typical flotation densities described for exosomes (between 1.10 and 1.18 g/mL), and denser vesicles (>1.24 g/mL). Both types of MV from RV-infected cells were more efficient at inhibiting T-cell function than were those from non-infected cells. We propose that RV infection of IEC releases MV that modulate viral immunity. PMID:21142445

  6. Membrane protein production in Escherichia coli cell-free lysates.

    PubMed

    Henrich, Erik; Hein, Christopher; Dötsch, Volker; Bernhard, Frank

    2015-07-08

    Cell-free protein production has become a core technology in the rapidly spreading field of synthetic biology. In particular the synthesis of membrane proteins, highly problematic proteins in conventional cellular production systems, is an ideal application for cell-free expression. A large variety of artificial as well as natural environments for the optimal co-translational folding and stabilization of membrane proteins can rationally be designed. The high success rate of cell-free membrane protein production allows to focus on individually selected targets and to modulate their functional and structural properties with appropriate supplements. The efficiency and robustness of lysates from Escherichia coli strains allow a wide diversity of applications and we summarize current strategies for the successful production of high quality membrane protein samples.

  7. Anhydrous Proton-Conducting Membranes for Fuel Cells

    NASA Technical Reports Server (NTRS)

    Narayanan, Sekharipuram; Yen, Shiao-Pin S.

    2005-01-01

    Polymeric electrolyte membranes that do not depend on water for conduction of protons are undergoing development for use in fuel cells. Prior polymeric electrolyte fuel-cell membranes (e.g., those that contain perfluorosulfonic acid) depend on water and must be limited to operation below a temperature of 125 C because they retain water poorly at higher temperatures. In contrast, the present developmental anhydrous membranes are expected to function well at temperatures up to 200 C. The developmental membranes exploit a hopping-and-reorganization proton- conduction process that can occur in the solid state in organic amine salts and is similar to a proton-conduction process in a liquid. This process was studied during the 1970s, but until now, there has been no report of exploiting organic amine salts for proton conduction in fuel cells.

  8. Effect of gas diffusion layer and membrane properties in an annular proton exchange membrane fuel cell

    NASA Astrophysics Data System (ADS)

    Khazaee, I.; Ghazikhani, M.; Esfahani, M. Nasr

    2012-01-01

    A complete three-dimensional and single phase computational dynamics model for annular proton exchange membrane (PEM) fuel cell is used to investigate the effect of changing gas diffusion layer and membrane properties on the performances, current density and gas concentration. The proposed model is a full cell model, which includes all the parts of the PEM fuel cell, flow channels, gas diffusion electrodes, catalyst layers and the membrane. Coupled transport and electrochemical kinetics equations are solved in a single domain; therefore no interfacial boundary condition is required at the internal boundaries between cell components. This computational fluid dynamics code is used as the direct problem solver, which is used to simulate the two-dimensional mass, momentum and species transport phenomena as well as the electron- and proton-transfer process taking place in a PEMFC that cannot be investigated experimentally. The results show that by increasing the thickness and decreasing the porosity of GDL the performance of the cell enhances that it is different with planner PEM fuel cell. Also the results show that by decreasing the thickness of the membrane the performance of the cell increases.

  9. Protein diffusion in plant cell plasma membranes: the cell-wall corral

    PubMed Central

    Martinière, Alexandre; Runions, John

    2013-01-01

    Studying protein diffusion informs us about how proteins interact with their environment. Work on protein diffusion over the last several decades has illustrated the complex nature of biological lipid bilayers. The plasma membrane contains an array of membrane-spanning proteins or proteins with peripheral membrane associations. Maintenance of plasma membrane microstructure can be via physical features that provide intrinsic ordering such as lipid microdomains, or from membrane-associated structures such as the cytoskeleton. Recent evidence indicates, that in the case of plant cells, the cell wall seems to be a major player in maintaining plasma membrane microstructure. This interconnection / interaction between cell-wall and plasma membrane proteins most likely plays an important role in signal transduction, cell growth, and cell physiological responses to the environment. PMID:24381579

  10. Cytotoxicity of bovine and porcine collagen membranes in mononuclear cells.

    PubMed

    Moura, Camilla Christian Gomes; Soares, Priscilla Barbosa Ferreira; Carneiro, Karine Fernandes; Souza, Maria Aparecida de; Magalhães, Denildo

    2012-01-01

    This study compared the cytotoxicity and the release of nitric oxide induced by collagen membranes in human mononuclear cells. Peripheral blood was collected from each patient and the separation of mononuclear cells was performed by Ficoll. Then, 2x10(5) cells were plated in 48-well culture plates under the membranes in triplicate. The polystyrene surface was used as negative control. Cell viability was assessed by measuring mitochondrial activity (MTT) at 4, 12 and 24 h, with dosage levels of nitrite by the Griess method for the same periods. Data had non-normal distribution and were analyzed by the Kruskal-Wallis test (p<0.05). Statistically significant differences (p<0.05) were observed between the membranes and the control in the experimental period, although there was a significant reduction in viability over time (p<0.01). At 4 and 12 h, the porcine membrane induced a higher release of nitrite compared with the control and bovine membrane, respectively (p<0.01), and this difference was maintained at 24 h (p<0.05). This in vitro study showed that the porcine collagen membrane induces an increased production of proinflammatory mediators by mononuclear cells in the first hours of contact, decreasing with time.

  11. Graphene-Induced Pore Formation on Cell Membranes

    PubMed Central

    Duan, Guangxin; Zhang, Yuanzhao; Luan, Binquan; Weber, Jeffrey K.; Zhou, Royce W.; Yang, Zaixing; Zhao, Lin; Xu, Jiaying; Luo, Judong; Zhou, Ruhong

    2017-01-01

    Examining interactions between nanomaterials and cell membranes can expose underlying mechanisms of nanomaterial cytotoxicity and guide the design of safer nanomedical technologies. Recently, graphene has been shown to exhibit potential toxicity to cells; however, the molecular processes driving its lethal properties have yet to be fully characterized. We here demonstrate that graphene nanosheets (both pristine and oxidized) can produce holes (pores) in the membranes of A549 and Raw264.7 cells, substantially reducing cell viability. Electron micrographs offer clear evidence of pores created on cell membranes. Our molecular dynamics simulations reveal that multiple graphene nanosheets can cooperate to extract large numbers of phospholipids from the membrane bilayer. Strong dispersion interactions between graphene and lipid-tail carbons result in greatly depleted lipid density within confined regions of the membrane, ultimately leading to the formation of water-permeable pores. This cooperative lipid extraction mechanism for membrane perforation represents another distinct process that contributes to the molecular basis of graphene cytotoxicity. PMID:28218295

  12. Membrane Targeting of P-type ATPases in Plant Cells

    SciTech Connect

    Jeffrey F. Harper, Ph.D.

    2004-06-30

    How membrane proteins are targeted to specific subcellular locations is a very complex and poorly understood area of research. Our long-term goal is to use P-type ATPases (ion pumps), in a model plant system Arabidopsis, as a paradigm to understand how members of a family of closely related membrane proteins can be targeted to different subcellular locations. The research is divided into two specific aims. The first aim is focused on determining the targeting destination of all 10 ACA-type calcium pumps (Arabidopsis Calcium ATPase) in Arabidopsis. ACAs represent a plant specific-subfamily of plasma membrane-type calcium pumps. In contrast to animals, the plant homologs have been found in multiple membrane systems, including the ER (ACA2), tonoplast (ACA4) and plasma membrane (ACA8). Their high degree of similarity provides a unique opportunity to use a comparative approach to delineate the membrane specific targeting information for each pump. One hypothesis to be tested is that an endomembrane located ACA can be re-directed to the plasma membrane by including targeting information from a plasma membrane isoform, ACA8. Our approach is to engineer domain swaps between pumps and monitor the targeting of chimeric proteins in plant cells using a Green Fluorescence Protein (GFP) as a tag. The second aim is to test the hypothesis that heterologous transporters can be engineered into plants and targeted to the plasma membrane by fusing them to a plasma membrane proton pump. As a test case we are evaluating the targeting properties of fusions made between a yeast sodium/proton exchanger (Sod2) and a proton pump (AHA2). This fusion may potentially lead to a new strategy for engineering salt resistant plants. Together these aims are designed to provide fundamental insights into the biogenesis and function of plant cell membrane systems.

  13. Polyarylenethioethersulfone Membranes for Fuel Cells (Postprint)

    DTIC Science & Technology

    2007-09-01

    precipitated copolymer was washed several times with deionized water in an attempt to completely remove the salts and then soxhlet - extracted in methanol...mation such as membrane resistance, charge-transfer resistance, and pore resistance was extracted from impedance plots using Nafion- and SPTES-50

  14. Fuel cell electrolyte membrane with basic polymer

    DOEpatents

    Larson, James M.; Pham, Phat T.; Frey, Matthew H.; Hamrock, Steven J.; Haugen, Gregory M.; Lamanna, William M.

    2012-12-04

    The present invention is an electrolyte membrane comprising an acid and a basic polymer, where the acid is a low-volatile acid that is fluorinated and is either oligomeric or non-polymeric, and where the basic polymer is protonated by the acid and is stable to hydrolysis.

  15. Fuel cell electrolyte membrane with basic polymer

    DOEpatents

    Larson, James M.; Pham, Phat T.; Frey, Matthew H.; Hamrock, Steven J.; Haugen, Gregory M.; Lamanna, William M.

    2010-11-23

    The present invention is an electrolyte membrane comprising an acid and a basic polymer, where the acid is a low-volatile acid that is fluorinated and is either oligomeric or non-polymeric, and where the basic polymer is protonated by the acid and is stable to hydrolysis.

  16. A Journey of Cytolethal Distending Toxins through Cell Membranes

    PubMed Central

    Boesze-Battaglia, Kathleen; Alexander, Desiree; Dlakić, Mensur; Shenker, Bruce J.

    2016-01-01

    The multifunctional role of lipids as structural components of membranes, signaling molecules, and metabolic substrates makes them an ideal partner for pathogens to hijack host cell processes for their own survival. The properties and composition of unique membrane micro-domains such as membrane rafts make these regions a natural target for pathogens as it affords them an opportunity to hijack cell signaling and intracellular trafficking pathways. Cytolethal distending toxins (Cdts), members of the AB2 family of toxins are comprised of three subunits, the active, CdtB unit, and the binding, CdtA-CdtC unit. Cdts are cyclomodulins leading to cell cycle arrest and apoptosis in a wide variety of cell types. Cdts from several species share a requirement for membrane rafts, and often cholesterol specifically for cell binding and CdtB mediated cytotoxicity. In this review we focus on how host–cell membrane bilayer organization contributes to the cell surface association, internalization, and action of bacteria derived cytolethal distending toxins (Cdts), with an emphasis on Aggregatibacter actinomycetemcomitans Cdt. PMID:27559534

  17. Gradiently crosslinked polymer electrolyte membranes in fuel cells

    NASA Astrophysics Data System (ADS)

    An, De; Wu, Bin; Zhang, Genlei; Zhang, Wen; Wang, Yuxin

    2016-01-01

    Polymer electrolyte membranes in fuel cells should be high in both ionic conductivity and mechanical strength. However, the two are often exclusive to each other. To solve this conundrum, a novel strategy is proposed in this paper, with extensively researched sulfonated poly (ether ether ketone) (SPEEK) membrane as a paradigm. A SPEEK membrane of high sulfonation degree is simply post-treated with NaBH4 and H2SO4 solution at ambient temperature for a certain time to afford the membrane with a gradient crosslinking structure. Measurements via 1H NMR, ATR-FTIR and SEM-EDS are conducted to verify such structural changes. The gradient crosslinks make practically no damage to proton conductance, but effectively restrain the membrane from over swelling and greatly enhance its tensile strength. A H2-O2 fuel cell with the gradiently crosslinked SPEEK membrane shows a maximal power density of 533 mW cm-2 at 80 °C, whereas the fuel cell with the pristine SPEEK membrane cannot be operated beyond 30 °C.

  18. Cell-free transfer of sterols by plant fractions

    SciTech Connect

    Morre, D.J.; Wilkinson, F.E.; Morre, D.M. ); Moreau, P. ); Sandelius, A.S. ); Penel, C.; Greppin, H. )

    1990-05-01

    Microsomes from etiolated hypocotyls of soybean or leaves of light-grown spinach radiolabeled in vivo with ({sup 3}H)acetate or in vitro with ({sup 3}H)squalene or ({sup 3}H)cholesterol as donor transferred radioactivity to unlabeled acceptor membranes immobilized on nitrocellulose. Most efficient transfer was with plasma membrane or tonoplast as the acceptor. The latter were highly purified by aqueous two-phase partition (plasma membrane) and preparative free-flow electrophoresis (tonoplast and plasma membrane). Plasma membrane- and tonoplast-free microsomes and purified mitochondria were less efficient acceptors. Sterol transfer was verified by thin-layer chromatography of extracted lipids. Transfer was time- and temperature-dependent, required ATP but was not promoted by cytosol. The nature of the donor (endoplasmic reticulum, Golgi apparatus or both) and of the transfer mechanism is under investigation.

  19. Isolation and partial characterization of antigens from basement membranes and streptococcal cell membrane (SCM) employing anti-SCM monoclonal antibody.

    PubMed

    Zelman, M E; Lange, C F

    1989-09-01

    Monoclonal antibodies (mAb) against streptococcal cell membrane (SCM) antigen were used to identify specific cross-reactive peptides prepared by trypsin digestion of purified glomerular basement membrane (GBM) and lung basement membrane (LBM). Anti-SCM mAb-coupled HPLC columns were used to affinity isolate soluble LBM, GBM, and SCM antigens which then were sized by HPLC. Alternatively, SCM, GBM, and LBM digests were subjected to an initial separation by HPLC into component polypeptides, followed by affinity purification and ELISA of these fractions using anti-SCM mAb. Comparison of the antigenic reactivities by ELISA of the sized polypeptides on a nanomolar basis permitted the estimation of their individual relative epitope densities. The results for SCM antigens showed increasing epitope density with increasing molecular size, which suggests that intact SCM consists of repeating epitopes. Low mol. wt GBM polypeptides in nanogram amounts inhibited mAb binding to SCM, indicating that these small GBM polypeptides may similarly contain more than a single cross-reactive epitope. The identification of these cross-reactive epitopes in LBM and GBM has important implications for the etiology of post-streptococcal sequelae.

  20. Direct visualization of membrane architecture of myelinating cells in transgenic mice expressing membrane-anchored EGFP.

    PubMed

    Deng, Yaqi; Kim, BongWoo; He, Xuelian; Kim, Sunja; Lu, Changqing; Wang, Haibo; Cho, Ssang-Goo; Hou, Yiping; Li, Jianrong; Zhao, Xianghui; Lu, Q Richard

    2014-04-01

    Myelinogenesis is a complex process that involves substantial and dynamic changes in plasma membrane architecture and myelin interaction with axons. Highly ramified processes of oligodendrocytes in the central nervous system (CNS) make axonal contact and then extrapolate to wrap around axons and form multilayer compact myelin sheathes. Currently, the mechanisms governing myelin sheath assembly and axon selection by myelinating cells are not fully understood. Here, we generated a transgenic mouse line expressing the membrane-anchored green fluorescent protein (mEGFP) in myelinating cells, which allow live imaging of details of myelinogenesis and cellular behaviors in the nervous systems. mEGFP expression is driven by the promoter of 2'-3'-cyclic nucleotide 3'-phosphodiesterase (CNP) that is expressed in the myelinating cell lineage. Robust mEGFP signals appear in the membrane processes of oligodendrocytes in the CNS and Schwann cells in the peripheral nervous system (PNS), wherein mEGFP expression defines the inner layers of myelin sheaths and Schmidt-Lanterman incisures in adult sciatic nerves. In addition, mEGFP expression can be used to track the extent of remyelination after demyelinating injury in a toxin-induced demyelination animal model. Taken together, the membrane-anchored mEGFP expression in the new transgenic line would facilitate direct visualization of dynamic myelin membrane formation and assembly during development and process remodeling during remyelination after various demyelinating injuries.

  1. Extraction methods of red blood cell membrane proteins for Multidimensional Protein Identification Technology (MudPIT) analysis.

    PubMed

    De Palma, Antonella; Roveri, Antonella; Zaccarin, Mattia; Benazzi, Louise; Daminelli, Simone; Pantano, Giorgia; Buttarello, Mauro; Ursini, Fulvio; Gion, Massimo; Mauri, Pier Luigi

    2010-08-13

    Since red blood cells (RBCs) lack nuclei and organelles, cell membrane is their main load-bearing component and, according to a dynamic interaction with the cytoskeleton compartment, plays a pivotal role in their functioning. Even if erythrocyte membranes are available in large quantities, the low abundance and the hydrophobic nature of cell membrane proteins complicate their purification and detection by conventional 2D gel-based proteomic approaches. So, in order to increase the efficiency of RBC membrane proteome identification, here we took advantage of a simple and reproducible membrane sub-fractionation method coupled to Multidimensional Protein Identification Technology (MudPIT). In addition, the adoption of a stringent RBC filtration strategy from the whole blood, permitted to remove exhaustively contaminants, such as platelets and white blood cells, and to identify a total of 275 proteins in the three RBC membrane fractions collected and analysed. Finally, by means of software for the elaboration of the great quantity of data obtained and programs for statistical analysis and protein classification, it was possible to determine the validity of the entire system workflow and to assign the proper sub-cellular localization and function for the greatest number of the identified proteins.

  2. Antioxidant activity of membrane-fractionated coffee extracts in dependence of the storage conditions

    NASA Astrophysics Data System (ADS)

    Mitev, D.; Peshev, D.; Peev, G.; Peeva, L.

    2016-10-01

    Present paper aims at one of the important aspects of the application of products with antioxidant activity: namely the preservation and change of their properties during the storage in different conditions, as well as their reliable characterisation. The tests of antioxidant properties were conducted with membrane-separated coffee extracts, isolated using a “Microdyn Nadir NP030P” type of commercial nanofiltration membrane (30% retention of NaCl; MWCO∼400). Prepared coffee permeates and retentates were stored 0÷10 days in cool/warm conditions, with/without air access and at different illumination conditions. The kinetics of content changes was evaluated according to Folin-Ciocalteu method of total phenolic/reducing content determination.

  3. Performance of cell-penetrating peptide-linked polymers physically mixed with poorly membrane-permeable molecules on cell membranes.

    PubMed

    Sakuma, Shinji; Suita, Masaya; Yamamoto, Takafumi; Masaoka, Yoshie; Kataoka, Makoto; Yamashita, Shinji; Nakajima, Noriko; Shinkai, Norihiro; Yamauchi, Hitoshi; Hiwatari, Ken-Ichiro; Hashizume, Akio; Tachikawa, Hiroyuki; Kimura, Ryoji; Ishimaru, Yuki; Kasai, Atsushi; Maeda, Sadaaki

    2012-05-01

    We are investigating a new class of penetration enhancers that enable poorly membrane-permeable molecules physically mixed with them to effectively penetrate cell membranes without their concomitant cellular uptake. Since we previously revealed that poly(N-vinylacetamide-co-acrylic acid) modified with d-octaarginine, which is a typical cell-penetrating peptide, significantly enhanced the nasal absorption of insulin, we examined the performance of the polymers on cell membranes. When Caco-2 cells were incubated with 5(6)-carboxyfluorescein (CF) for 30 min, approximately 0.1% of applied CF was internalized into the cells. This poor membrane permeability was dramatically enhanced by d-octaarginine-linked polymers; a 25-fold increase in the cellular uptake of CF was observed when the polymer concentration was adjusted to 0.2mg/mL. None of the individual components, for example, d-octaarginine, had any influence on CF uptake, demonstrating that only d-octaarginine anchored chemically to the polymeric platform enhanced the membrane permeation of CF. The polymer-induced CF uptake was consistently high even when the incubation time was extended to 120 min. Confocal laser scanning microphotographs of cells incubated with d-octaarginine-linked polymers bearing rhodamine red demonstrated that the cell outline was stained with red fluorescence. The polymer-induced CF uptake was significantly suppressed by 5-(N-ethyl-N-isopropyl)amiloride, which is an inhibitor of macropinocytosis. Results indicated that d-octaarginine-linked polymers remained on the cell membrane and poorly membrane-permeable CF was continuously internalized into cells mainly via macropinocytosis repeated for the individual peptidyl branches in the polymer backbone.

  4. Distance Measurement on an Endogenous Membrane Transporter in E. coli Cells and Native Membranes Using EPR Spectroscopy.

    PubMed

    Joseph, Benesh; Sikora, Arthur; Bordignon, Enrica; Jeschke, Gunnar; Cafiso, David S; Prisner, Thomas F

    2015-05-18

    Membrane proteins may be influenced by the environment, and they may be unstable in detergents or fail to crystallize. As a result, approaches to characterize structures in a native environment are highly desirable. Here, we report a novel general strategy for precise distance measurements on outer membrane proteins in whole Escherichia coli cells and isolated outer membranes. The cobalamin transporter BtuB was overexpressed and spin-labeled in whole cells and outer membranes and interspin distances were measured to a spin-labeled cobalamin using pulse EPR spectroscopy. A comparative analysis of the data reveals a similar interspin distance between whole cells, outer membranes, and synthetic vesicles. This approach provides an elegant way to study conformational changes or protein-protein/ligand interactions at surface-exposed sites of membrane protein complexes in whole cells and native membranes, and provides a method to validate outer membrane protein structures in their native environment.

  5. Demonstration of a Sample Preparation Method for Biological Detection Based on a Novel Membrane Fractionation Technology

    DTIC Science & Technology

    2008-12-31

    oftarget nucleic acid in the sample) Deoxyribonucleic acid kilo Dalton Leonard Wood Institute Midwest Research Institute Quantitative Polymerase...membranes. Desalination 227: 111-119. 7) Kong S, Titchener- Hooker N, Levy MS, 2006. Plasmid DNA processing for gene therapy and vaccination: Studies on...Computerized Helical Scanning Technique. JAm Soc Nephrol, 13: S53-S61. 15. Bricefio, M.I. , Joseph , D.D. 2003. Self-lubricated transport of aqueous

  6. A Comparative Spin-Label Study of Isolated Plasma Membranes and Plasma Membranes of Whole Cells and Protoplasts from Cold-Hardened and Nonhardened Winter Rye

    PubMed Central

    Windle, John J.

    1988-01-01

    Lipid-lipid and lipid-protein interactions in the plasma membranes of whole cells and protoplasts and an isolated plasma membrane fraction from winter rye (Secale cereale L. cv Puma) have been studied by spin labeling. Spectra were recorded between −40°C and 40°C using the freely diffusing spin-label, 16-doxyl stearic acid, as a midbilayer membrane probe. The probe was reduced by the whole cells and protoplasts and reoxidized by external potassium ferricyanide. The reoxidized probe was assumed to be localized in the plasma membrane. The spectra consisted of the superposition of a narrow and a broad component indicating that both fluid and immobilized lipids were present in the plasma membrane. The two components were separated by digital subtraction of the immobilized component. Temperature profiles of the membranes were developed using the percentage of immobilized lipid present at each temperature and the separation between the outermost hyperfine lines for the fluid lipid component. Lipid immobilization was attributed to lipid-protein interactions, lipid-cell wall interactions, and temperature-induced lipid phase transitions to the gel-state. Temperature profiles were compared for both cold-hardened and nonhardened protoplasts, plasma membranes, and plasma membrane lipids, respectively. Although cold-hardening extended the range of lipid fluidity by 5°C, it had no effect on lipid-protein interactions or activation energies of lipid mobility. Differences were found, however, between the temperature profiles for the different samples, suggesting that alterations in the plasma membrane occurred as a consequence of the isolation methods used. PMID:16666471

  7. Tight binding of proteins to membranes from older human cells.

    PubMed

    Truscott, Roger J W; Comte-Walters, Susana; Ablonczy, Zsolt; Schwacke, John H; Berry, Yoke; Korlimbinis, Anastasia; Friedrich, Michael G; Schey, Kevin L

    2011-12-01

    The lens is an ideal model system for the study of macromolecular aging and its consequences for cellular function, since there is no turnover of lens fibre cells. To examine biochemical processes that take place in the lens and that may also occur in other long-lived cells, membranes were isolated from defined regions of human lenses that are synthesised at different times during life, and assayed for the presence of tightly bound cytosolic proteins using quantitative iTRAQ proteomics technology. A majority of lens beta crystallins and all gamma crystallins became increasingly membrane bound with age, however, the chaperone proteins alpha A and alpha B crystallin, as well as the thermally-stable protein, βB2 crystallin, did not. Other proteins such as brain-associated signal protein 1 and paralemmin 1 became less tightly bound in the older regions of the lens. It is evident that protein-membrane interactions change significantly with age. Selected proteins that were formerly cytosolic become increasingly tightly bound to cell membranes with age and are not removed even by treatment with 7 M urea. It is likely that such processes reflect polypeptide denaturation over time and the untoward binding of proteins to membranes may alter membrane properties and contribute to impairment of communication between older cells.

  8. 3D visualization of membrane failures in fuel cells

    NASA Astrophysics Data System (ADS)

    Singh, Yadvinder; Orfino, Francesco P.; Dutta, Monica; Kjeang, Erik

    2017-03-01

    Durability issues in fuel cells, due to chemical and mechanical degradation, are potential impediments in their commercialization. Hydrogen leak development across degraded fuel cell membranes is deemed a lifetime-limiting failure mode and potential safety issue that requires thorough characterization for devising effective mitigation strategies. The scope and depth of failure analysis has, however, been limited by the 2D nature of conventional imaging. In the present work, X-ray computed tomography is introduced as a novel, non-destructive technique for 3D failure analysis. Its capability to acquire true 3D images of membrane damage is demonstrated for the very first time. This approach has enabled unique and in-depth analysis resulting in novel findings regarding the membrane degradation mechanism; these are: significant, exclusive membrane fracture development independent of catalyst layers, localized thinning at crack sites, and demonstration of the critical impact of cracks on fuel cell durability. Evidence of crack initiation within the membrane is demonstrated, and a possible new failure mode different from typical mechanical crack development is identified. X-ray computed tomography is hereby established as a breakthrough approach for comprehensive 3D characterization and reliable failure analysis of fuel cell membranes, and could readily be extended to electrolyzers and flow batteries having similar structure.

  9. Synaptic and Golgi membrane recycling in cochlear hair cells.

    PubMed

    Siegel, J H; Brownell, W E

    1986-06-01

    Membrane recycling in the mechanoreceptive sensory cells of the mammalian cochlea was studied by observing membrane-bound horseradish peroxidase (HRP) reaction product following brief in vivo exposure to the enzyme. In the inner hair cell (IHC), peroxidase was taken up into coated vesicles and became incorporated into synaptic vesicles surrounding presynaptic bodies, but much HRP was also transported to the apical zone where reaction product appeared in all components of the Golgi complex. Neither the subsurface cisternae nor a tubular network associated with clusters of mitochondria were labelled. Outer hair cells (OHCs) showed considerably less membrane-bound reaction product than IHCs, indicating less rapid plasmalemmal recycling. Most membrane-bound reaction product was contained in coated vesicles and small vacuoles in the synaptic zone, but was occasionally seen in multivesicular bodies in the most apical zone. No labelled organelles were detected in the large central region of the OHC. A diffuse staining of the cytoplasm, particularly pronounced in OHCs, often interfered with the evaluation of membrane-bound reaction product in OHCs. This staining pattern could be qualitatively reproduced in both IHCs and OHCs by incubating fixed segments of the organ of Corti in oxidized diaminobenzidine. The presence of labelled synaptic vesicles associated with presynaptic bodies of IHCs and OHCs suggests that they are formed from membrane retrieved from the plasmalemma. We found no evidence that the subsurface cisternae of IHCs or the laminated cisternae of OHCs are derived from the cell surface as they never contained reaction product.

  10. Cryopreservation of cells: FT-IR monitoring of lipid membrane at freeze-thaw cycles.

    PubMed

    Giugliarelli, A; Sassi, P; Urbanelli, L; Paolantoni, M; Caponi, S; Ricci, M; Emiliani, C; Fioretto, D; Morresi, A

    2016-01-01

    In the present study, FTIR spectroscopy was used to monitor the freeze-thaw cycle of two cellular lines (HuDe and Jurkat) suspended in three different media: phosphate buffer solution (PBS); dimethylsulfoxide (DMSO)/PBS solution at 0.1 DMSO molar fraction; and CryoSure (0.1 DMSO molar fraction PBS solution+dextran 5% w/v) solution. The Trypan Blue test was also applied before freezing and after thawing each cell sample to estimate the recovery of membrane integrity after thermal treatment, and correlate this datum with spectroscopic results. By following the temperature evolution of two different spectral components (the libration and bending combination mode νc(H2O) at 2000-2500 cm(-1), and the methylene symmetric stretching vibration νsym(CH2) at about 2850 cm(-1)) in the -120÷28°C range, we evidenced the main transition of lipid membrane in connection with cell dehydration, as induced by ice formation in the extracellular medium. In particular, in DMSO/PBS and CryoSure samples we observed a transition to a more rigid state of the lipid membrane together with an increased amount of non-freezable water in the extracellular medium; these results are connected to the role of DMSO as a cryoprotective agent irrespective of the nature of cell type.

  11. Cytotoxic effects of Kingella kingae outer membrane vesicles on human cells

    PubMed Central

    Maldonado, R; Wei, R; Kachlany, SC; Kazi, M; Balashova, NV

    2011-01-01

    Kingella kingae is an emerging pathogen causing osteoarticular infections in pediatric patients. Electron microscopy of K. kingae clinical isolates revealed the heterogeneously-sized membranous structures blebbing from the outer membrane that were classified as outer membrane vesicles (OMVs). OMVs purified from the secreted fraction of a septic arthritis K. kingae isolate were characterized. Among several major proteins, K. kingae OMVs contained virulence factors RtxA toxin and PilC2 pilus adhesin. RtxA was also found secreted as a soluble protein in the extracellular environment indicating that the bacterium may utilize different mechanisms for the toxin delivery. OMVs were shown to be hemolytic and possess some leukotoxic activity while high leukotoxicity was detected in the non-hemolytic OMV-free component of the secreted fraction. OMVs were internalized by human osteoblasts and synovial cells. Upon interaction with OMVs, the cells produced increased levels of human granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleuskin 6 (IL-6) suggesting that these cytokines might be involved in the signaling response of infected joint and bone tissues during natural K. kingae infection. This study is the first report of OMV production by K. kingae and demonstrates that OMVs are a complex virulence factor of the organism causing cytolytic and inflammatory effects on host cells. PMID:21443941

  12. Cytotoxic effects of Kingella kingae outer membrane vesicles on human cells.

    PubMed

    Maldonado, R; Wei, R; Kachlany, S C; Kazi, M; Balashova, N V

    2011-01-01

    Kingella kingae is an emerging pathogen causing osteoarticular infections in pediatric patients. Electron microscopy of K. kingae clinical isolates revealed the heterogeneously-sized membranous structures blebbing from the outer membrane that were classified as outer membrane vesicles (OMVs). OMVs purified from the secreted fraction of a septic arthritis K. kingae isolate were characterized. Among several major proteins, K. kingae OMVs contained virulence factors RtxA toxin and PilC2 pilus adhesin. RtxA was also found secreted as a soluble protein in the extracellular environment indicating that the bacterium may utilize different mechanisms for the toxin delivery. OMVs were shown to be hemolytic and possess some leukotoxic activity while high leukotoxicity was detected in the non-hemolytic OMV-free component of the secreted fraction. OMVs were internalized by human osteoblasts and synovial cells. Upon interaction with OMVs, the cells produced increased levels of human granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 6 (IL-6) suggesting that these cytokines might be involved in the signaling response of infected joint and bone tissues during natural K. kingae infection. This study is the first report of OMV production by K. kingae and demonstrates that OMVs are a complex virulence factor of the organism causing cytolytic and inflammatory effects on host cells.

  13. Anaerobic digestion of the organic fraction of municipal solid waste in a two-stage membrane process.

    PubMed

    Trzcinski, A P; Stuckey, D C

    2009-01-01

    A batch of the Organic Fraction of Municipal Solid Waste (OFMSW) was treated in a two-step process with effluent recirculation comprising a novel hydrolytic reactor (HR) followed by a Submerged Anaerobic Membrane Bioreactor (SAMBR) operating at a stable permeate flux of 5.6 L/m(2) hr (LMH). A soluble COD removal higher than 95% was obtained from the SAMBR. The soluble COD as well as the Total Suspended Solids (TSS) did not build up due to efficient hydrolysis inside the SAMBR, and no VFA accumulation occurred due to the complete retention of methanogens by the membrane as well as the formation of syntrophic associations. Because of the microfiltration membrane in the second reactor a stabilized leachate was obtained from the very first days of the treatment and the highly stable process enabled shorter treatment periods compared to traditional leach bed processes. This experiment showed that the recycle of the stabilised leachate does not lead to a build up of SCOD. Size exclusion chromatography analysis confirmed that high molecular weight compounds were completely degraded and did not appear in the SAMBR permeate, and that low molecular weight fulvic-like and medium MW material were present in the permeate of the SAMBR but their concentration remained stable with time.

  14. Effects of Extracellular Calcium on Cell Membrane Resealing during Sonoporation

    NASA Astrophysics Data System (ADS)

    Zhou, Yun; Cui, Jianmin; Deng, Cheri X.

    2006-05-01

    Sonoporation has been exploited as a novel strategy for intracellular drug and gene delivery. In sonoporation, ultrasound application generates transient pores or openings in the cell membrane that allow entry of extracellular agents normally not permeable to the cell membrane. In order to improve the sonoporation outcome, we seek to obtain improved understanding of the sonoporation mechanism and investigate the factors affecting sonoporation process. We established a voltage clamp technique for real time measurement of sonoporation at single cell level using Xenopus oocytes as a model system. As both cell survival and intracellular delivery efficiency of drug or genes depend on the sonoporation dynamic process, and Calcium plays important roles in cellular processes, we focus on studying of the effect of extracellular Calcium concentration on the formation, extension, and resealing of membrane pores in sonoporation. We obtained experimental results demonstrating that the cell membrane reseals in the order of seconds in the presence of physiological level of extracellular [Ca]. We measured the resealing as function of extracellular [Ca] (0-1.8mM) and observed that the resealing rate decreases as extracellular [Ca] decreases from normal physiological level. No resealing was demonstrated when 1mM EGTA was added in the extracellular medium to chelate the [Ca] extracellularly. Our experimental findings suggest that extracellular Calcium plays an important role in controlling membrane resealing in sonoporation and thus the sonoporation outcome such as cell survival and delivery efficiency.

  15. Anion selective membrane. [ion exchange resins and ion exchange membrane electrolytes for electrolytic cells

    NASA Technical Reports Server (NTRS)

    Alexander, S. S.; Geoffroy, R. R.; Hodgdon, R. B.

    1975-01-01

    Experimental anion permselective membranes were prepared and tested for their suitability as cell separators in a chemical redox power storage system being developed at NASA-Lewis Research Center. The goals of long-term (1000 hr) oxidative and thermal stability at 80 C in FeCl3 and CrCl3 electrolytes were met by most of the weak base and strong base amino exchange groups considered in the program. Good stability is exhibited by several of the membrane substrate resins. These are 'styrene' divinylbenzene copolymer and PVC film. At least four membrane systems produce strong flexible films with electrochemical properties (resistivity, cation transfer) superior to those of the 103QZL, the most promising commercial membrane. The physical and chemical properties of the resins are listed.

  16. Cell Membrane-Cloaked Nanoparticles for Targeted Therapeutics

    NASA Astrophysics Data System (ADS)

    Luk, Brian Tsengchi

    The advent of nanoparticle-based delivery systems has made a significant impact on clinical patient outcomes. In recent decades, myriad nanoparticle-based therapeutic agents have been developed for the treatment and management of ailments such as cancer, diabetes, pain, bacterial infections, and asthma, among many others. Nanotherapeutics offer many distinct advantages over conventional free drug formulations. For example, nanoparticles are able to accumulate at tumor sites by extravasation through leaky vasculature at tumor sites via the enhanced permeability and retention (EPR) effect; nanoparticles can also be tailored to have desirable characteristics, such as prolonged circulation in the blood stream, improved drug encapsulation, and sustained or triggered drug release. Currently, a growing number of nanoformulations with favorable pharmacological profiles and promising efficacy are being used in clinical trials for the treatment of various cancers. Building on the success of these encouraging clinical results, new engineering strategies have emerged that combine synthetic nanoparticles with natural biomaterials to create nature-inspired biomimetic delivery systems. The work presented in this dissertation focuses on the biointerfacing between synthetic and natural materials, namely in the manifestation of cell membrane-coated nanoparticles. By exploiting the natural functionalities of source cell membranes, cell membrane-cloaked nanoparticles have huge potential in the delivery of therapeutic agents for a variety of applications. The first portion of this thesis will focus on understanding the fundamentals underlying cell membrane coating on synthetic nanoparticles. First introduced in 2011, cell membrane-cloaked nanoparticles showed immediate promise in drug delivery applications, but further understanding was necessary to be able to harness the full potential of the membrane coating platform. The first section provides further insight into the interfacial

  17. A water and heat management model for proton-exchange-membrane fuel cells

    SciTech Connect

    Nguyen, T.V.; White, R.E. . Dept. of Chemical Engineering)

    1993-08-01

    Proper water and heat management are essential for obtaining high-power-density performance at high energy efficiency for proton-exchange-membrane fuel cells. A water and heat management model was developed and used to investigate the effectiveness of various humidification designs. The model accounts for water transport across the membrane by electro-osmosis and diffusion, heat transfer from the solid phase to the gas phase and latent heat associated with water evaporation and condensation in the flow channels. Results from the model showed that at high current (> 1A/cm[sup 2]) ohmic loss in the membrane accounts for a large fraction of the voltage loss in the cell and back diffusion of water from the cathode side of the membrane is insufficient to keep the membrane hydrated (i.e., conductive). Consequently, to minimize this ohmic loss the anode stream must be humidified, and when air is used instead of pure oxygen the cathode stream must also be humidified.

  18. Interaction of injectable neurotropic drugs with the red cell membrane.

    PubMed

    Reinhart, Walter H; Lubszky, Szabina; Thöny, Sandra; Schulzki, Thomas

    2014-10-01

    The normal red blood cell (RBC) shape is a biconcave discocyte. An intercalation of a drug in the outer half of the membrane lipid bilayer leads to echinocytosis, an intercalation in the inner half to stomatocytosis. We have used the shape transforming capacity of RBCs as a model to analyse the membrane interaction potential of various neurotropic drugs. Chlorpromazine, clomipramine, citalopram, clonazepam, and diazepam induced a reversible stomatocytosis, phenytoin induced echinocytosis, while the anticonvulsants levetiracetam, valproic acid and phenobarbital had no effect. This diversity of RBC shape transformations suggests that the pharmacological action is not linked to the membrane interaction. We conclude that this simple RBC shape transformation assay could be a useful tool to screen for potential drug interactions with cell membranes.

  19. Direct Cytoskeleton Forces Cause Membrane Softening in Red Blood Cells

    PubMed Central

    Rodríguez-García, Ruddi; López-Montero, Iván; Mell, Michael; Egea, Gustavo; Gov, Nir S.; Monroy, Francisco

    2015-01-01

    Erythrocytes are flexible cells specialized in the systemic transport of oxygen in vertebrates. This physiological function is connected to their outstanding ability to deform in passing through narrow capillaries. In recent years, there has been an influx of experimental evidence of enhanced cell-shape fluctuations related to metabolically driven activity of the erythroid membrane skeleton. However, no direct observation of the active cytoskeleton forces has yet been reported to our knowledge. Here, we show experimental evidence of the presence of temporally correlated forces superposed over the thermal fluctuations of the erythrocyte membrane. These forces are ATP-dependent and drive enhanced flickering motions in human erythrocytes. Theoretical analyses provide support for a direct force exerted on the membrane by the cytoskeleton nodes as pulses of well-defined average duration. In addition, such metabolically regulated active forces cause global membrane softening, a mechanical attribute related to the functional erythroid deformability. PMID:26083919

  20. A new material concept for the red cell membrane.

    PubMed

    Evans, E A

    1973-09-01

    The proposition is made that the red cell membrane is a two-dimensional, incompressible material and a general stress-strain law is developed for finite deformations. In the linear form, the character of such a material is analogous to a two-dimensional Mooney material (e.g., rubber), indicating that the molecular structure in the plane of the membrane would consist of long chains, randomly kinked and cross-linked in the natural state. The loose network could be provided by the protein component and the lipid phase could exist interstitially as a liquid bilayer, giving the membrane its two-dimensional incompressibility. The material provides the capability of large deformations exhibited by the discocyte and yet the rigidity associated with the osmotic spherocyte state. It is demonstrated that a membrane of this type can form a sphere at constant area. An illustrative example of the application to single cell discocyte-to-osmotic spherocyte transformations is presented.

  1. Attachment of killed Mycoplasma gallisepticum cells and membranes to erythrocytes

    SciTech Connect

    Banai, M.; Kahane, I.; Feldner, J.; Razin, S.

    1981-11-01

    To correlate viability with attachment capacity, Mycoplasma gallisepticum cells harvested at different growth phases and treated by various agents were tested for their capacity to attach to human erythrocytes. The results show that viability per se is not essential for M. gallisepticum attachment to erythrocytes, as cells killed by ultraviolet irradiation and membranes isolated by lysing M. gallisepticum cells by various means retained attachment capacity. However, treatment of the mycoplasmas by protein-denaturing agents, such as heart, glutaraldehyde, or prolonged exposure to low pH, drastically affected or even abolished attachment, supporting the protein nature of the mycoplasma membrane components responsible for specific binding to the sialoglycoprotein receptors on the erythrocytes.

  2. Discovery of novel hematopoietic cell adhesion molecules from human bone marrow stromal cell membrane protein extracts by a new cell-blotting technique.

    PubMed

    Seshi, B

    1994-05-01

    In an attempt to define the role of cell adhesion molecules (CAMs) within the bone marrow (BM) microenvironment in normal hematopoiesis and in leukemia development, a novel cell-blotting technique that involved cell adhesion to protein bands after separation by lithium dodecyl sulfate-polyacrylamide gel electrophoresis (LDS-PAGE) and blotting onto polyvinylidene difluoride (PVDF) membrane has been developed. Human BM stromal cell membrane fractions have been prepared from Dexter-type cultures after cell lysis by sonification and differential centrifugations of the sonification contents. The 20,000 g pellets representing membrane fractions have been solubilized by 2% Triton X-100, 0.575% LDS, and 8 mol/L urea in sequential order. The protein extracts are fractionated by LDS-PAGE and screened for CAMs by the new cell-blotting technique. This led to identification of nine protein bands in lanes containing LDS extracts showing adhesion of KG1a (CD34+ progenitor myeloid) cells. Evidence that the BM proteins exhibiting KG1a cell adhesion are novel CAMs is based on the observations that these proteins, in comparison with known CAMs, specifically VCAM-1, CD54, and CD44, show (1) contrasting detergent-solubility properties, (2) different temperature requirement for mediating cell adhesion function, and (3) markedly distinct electrophoretic mobilities. The various cell types tested, notably KG1a, NALM-6, WIL-2, Ramos, HS-Sultan, K562, JY B lymphoblastoid cells, and T lymphoblasts, showed distinctive patterns of binding to different subsets of BM CAMs. These results demonstrate a new approach to studies of molecular mechanisms that may determine specificity of hematopoietic cellular localization within BM microenvironment and may play an important role in controlling hematopoiesis.

  3. Coating nanofiber scaffolds with beta cell membrane to promote cell proliferation and function

    NASA Astrophysics Data System (ADS)

    Chen, Wansong; Zhang, Qiangzhe; Luk, Brian T.; Fang, Ronnie H.; Liu, Younian; Gao, Weiwei; Zhang, Liangfang

    2016-05-01

    The cell membrane cloaking technique has emerged as an intriguing strategy in nanomaterial functionalization. Coating synthetic nanostructures with natural cell membranes bestows the nanostructures with unique cell surface antigens and functions. Previous studies have focused primarily on development of cell membrane-coated spherical nanoparticles and the uses thereof. Herein, we attempt to extend the cell membrane cloaking technique to nanofibers, a class of functional nanomaterials that are drastically different from nanoparticles in terms of dimensional and mechanophysical characteristics. Using pancreatic beta cells as a model cell line, we demonstrate successful preparation of cell membrane-coated nanofibers and validate that the modified nanofibers possess an antigenic exterior closely resembling that of the source beta cells. When such nanofiber scaffolds are used to culture beta cells, both cell proliferation rate and function are significantly enhanced. Specifically, glucose-dependent insulin secretion from the cells is increased by near five-fold compared with the same beta cells cultured in regular, unmodified nanofiber scaffolds. Overall, coating cell membranes onto nanofibers could add another dimension of flexibility and controllability in harnessing cell membrane functions and offer new opportunities for innovative applications.

  4. Phytosphingosine kills Candida albicans by disrupting its cell membrane.

    PubMed

    Veerman, Enno C I; Valentijn-Benz, Marianne; van't Hof, Wim; Nazmi, Kamran; van Marle, Jan; Amerongen, Arie V Nieuw

    2010-01-01

    The mechanism of action of phytosphingosine (PHS), a member of the sphingosine family which has candidacidal activity when added externally, was investigated. Previously, it has been reported that the fungicidal activity of PHS is based on the induction of caspase-independent apoptosis. In contrast, we found that addition of PHS causes a direct permeabilization of the plasma membrane of yeast, highlighted by the influx of the membrane probe propidium iodide, and the efflux of small molecules (i.e., adenine nucleotides) as well as large cellular constituents such as proteins. Freeze-fracture electron microscopy revealed that PHS treatment causes severe damage of the plasma membrane of the cell, which seems to have lost its integrity completely. We also found that PHS reverts the azide-induced insensitivity to histatin 5 (Hst5) of Candida albicans. In a previous study, we had found that the decreased sensitivity to Hst5 of energy-depleted cells is due to rigidification of the plasma membrane, which could be reverted by the membrane fluidizer benzyl alcohol. In line with the increased membrane permeabilization and ultrastructural damage, this reversal of the azide-induced insensitivity by PHS also points to a direct interaction between PHS and the cytoplasmic membrane of C. albicans.

  5. Epidermal cells adhere preferentially to type IV (basement membrane) collagen

    PubMed Central

    1979-01-01

    Epidermal cells from adult guinea pig skin attach and differentiate preferentially on substrates of type IV (basement membrane) collagen, compared to those of types I--III collagen. In contrast, guinea pig dermal fibroblasts attach equally well to all four collagen substrates. Fibronectin mediates the attachment of fibroblasts but not of epidermal cells to collagen. PMID:422650

  6. Numerical modeling transport phenomena in proton exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Suh, DongMyung

    To study the coupled phenomena occurring in proton exchange membrane fuel cells, a two-phase, one-dimensional, non-isothermal model is developed in the chapter 1. The model includes water phase change, proton transport in the membrane and electro-osmotic effect. The thinnest, but most complex layer in the membrane electrode assembly, catalyst layer, is considered an interfacial boundary between the gas diffusion layer and the membrane. Mass and heat transfer and electro-chemical reaction through the catalyst layer are formulated into equations, which are applied to boundary conditions for the gas diffusion layer and the membrane. Detail accounts of the boundary equations and the numerical solving procedure used in this work are given. The polarization curve is calculated at different oxygen pressures and compared with the experimental results. When the operating condition is changed along the polarization curve, the change of physicochemical variables in the membrane electrode assembly is studied. In particular, the over-potential diagram presents the usage of the electrochemical energy at each layer of the membrane electrode assembly. Humidity in supplying gases is one of the most important factors to consider for improving the performance of PEMFE. Both high and low humidity conditions can result in a deteriorating cell performance. The effect of humidity on the cell performance is studied in the chapter 2. First, a numerical model based on computational fluid dynamics is developed. Second, the cell performances are simulated, when the relative humidity is changed from 0% to 100% in the anode and the cathode channel. The simulation results show how humidity in the reactant gases affects the water content distribution in the membrane, the over-potential at the catalyst layers and eventually the cell performance. In particular, the rapid enhancement in the cell performance caused by self-hydrating membrane is captured by the simulation. Fully humidifying either H2

  7. Scalable nanostructured membranes for solid-oxide fuel cells.

    PubMed

    Tsuchiya, Masaru; Lai, Bo-Kuai; Ramanathan, Shriram

    2011-05-01

    The use of oxide fuel cells and other solid-state ionic devices in energy applications is limited by their requirement for elevated operating temperatures, typically above 800°C (ref. 1). Thin-film membranes allow low-temperature operation by reducing the ohmic resistance of the electrolytes. However, although proof-of-concept thin-film devices have been demonstrated, scaling up remains a significant challenge because large-area membranes less than ~ 100 nm thick are susceptible to mechanical failure. Here, we report that nanoscale yttria-stabilized zirconia membranes with lateral dimensions on the scale of millimetres or centimetres can be made thermomechanically stable by depositing metallic grids on them to function as mechanical supports. We combine such a membrane with a nanostructured dense oxide cathode to make a thin-film solid-oxide fuel cell that can achieve a power density of 155 mW cm⁻² at 510 °C. We also report a total power output of more than 20 mW from a single fuel-cell chip. Our large-area membranes could also be relevant to electrochemical energy applications such as gas separation, hydrogen production and permeation membranes.

  8. Sequential CD34 cell fractionation by magnetophoresis in a magnetic dipole flow sorter.

    PubMed

    Schneider, Thomas; Karl, Stephan; Moore, Lee R; Chalmers, Jeffrey J; Williams, P Stephen; Zborowski, Maciej

    2010-01-01

    Cell separation and fractionation based on fluorescent and magnetic labeling procedures are common tools in contemporary research. These techniques rely on binding of fluorophores or magnetic particles conjugated to antibodies to target cells. Cell surface marker expression levels within cell populations vary with progression through the cell cycle. In an earlier work we showed the reproducible magnetic fractionation (single pass) of the Jurkat cell line based on the population distribution of CD45 surface marker expression. Here we present a study on magnetic fractionation of a stem and progenitor cell (SPC) population using the established acute myelogenous leukemia cell line KG-1a as a cell model. The cells express a CD34 cell surface marker associated with the hematopoietic progenitor cell activity and the progenitor cell lineage commitment. The CD34 expression level is approximately an order of magnitude lower than that of the CD45 marker, which required further improvements of the magnetic fractionation apparatus. The cells were immunomagnetically labeled using a sandwich of anti-CD34 antibody-phycoerythrin (PE) conjugate and anti-PE magnetic nanobead and fractionated into eight components using a continuous flow dipole magnetophoresis apparatus. The CD34 marker expression distribution between sorted fractions was measured by quantitative PE flow cytometry (using QuantiBRITE PE calibration beads), and it was shown to be correlated with the cell magnetophoretic mobility distribution. A flow outlet addressing scheme based on the concept of the transport lamina thickness was used to control cell distribution between the eight outlet ports. The fractional cell distributions showed good agreement with numerical simulations of the fractionation based on the cell magnetophoretic mobility distribution in the unsorted sample.

  9. Sequential CD34 cell fractionation by magnetophoresis in a magnetic dipole flow sorter

    PubMed Central

    Schneider, Thomas; Karl, Stephan; Moore, Lee R.; Chalmers, Jeffrey J.; Williams, P. Stephen; Zborowski, Maciej

    2010-01-01

    Cell separation and fractionation based on fluorescent and magnetic labeling procedures are common tools in contemporary research. These techniques rely on binding of fluorophores or magnetic particles conjugated to antibodies to target cells. Cell surface marker expression levels within cell populations vary with progression through the cell cycle. In an earlier work we showed the reproducible magnetic fractionation (single pass) of the Jurkat cell line based on the population distribution of CD45 surface marker expression. Here we present a study on magnetic fractionation of a stem and progenitor cell (SPC) population using the established acute myelogenous leukemia cell line KG-1a as a cell model. The cells express a CD34 cell surface marker associated with the hematopoietic progenitor cell activity and the progenitor cell lineage commitment (related to the CD34 marker expression level). The CD34 expression level is approximately an order of magnitude lower than that of the CD45 marker, which required further improvements of the magnetic fractionation apparatus. The cells were immuno-magnetically labeled using a sandwich of anti CD34 antibody-phycoerythrin (PE) conjugate and anti PE magnetic nanobead and fractionated into eight components using a continuous flow dipole magnetophoresis apparatus. The CD34 marker expression distribution between sorted fractions was measured by quantitative PE flow cytometry (using QuantiBRITE™ PE calibration beads), and it was shown to be correlated with the cell magnetophoretic mobility distribution. A flow outlet addressing scheme based on the concept of the transport lamina thickness was used to control cell distribution between the eight outlet ports. The fractional cell distributions showed good agreement with numerical simulations of the fractionation based on the cell magnetophoretic mobility distribution in the unsorted sample. PMID:20024182

  10. Understanding the transport processes in polymer electrolyte membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Cheah, May Jean

    Polymer electrolyte membrane (PEM) fuel cells are energy conversion devices suitable for automotive, stationary and portable applications. An engineering challenge that is hindering the widespread use of PEM fuel cells is the water management issue, where either a lack of water (resulting in membrane dehydration) or an excess accumulation of liquid water (resulting in fuel cell flooding) critically reduces the PEM fuel cell performance. The water management issue is addressed by this dissertation through the study of three transport processes occurring in PEM fuel cells. Water transport within the membrane is a combination of water diffusion down the water activity gradient and the dragging of water molecules by protons when there is a proton current, in a phenomenon termed electro-osmotic drag, EOD. The impact of water diffusion and EOD on the water flux across the membrane is reduced due to water transport resistance at the vapor/membrane interface. The redistribution of water inside the membrane by EOD causes an overall increase in the membrane resistance that regulates the current and thus EOD, thereby preventing membrane dehydration. Liquid water transport in the PEM fuel cell flow channel was examined at different gas flow regimes. At low gas Reynolds numbers, drops transitioned into slugs that are subsequently pushed out of the flow channel by the gas flow. The slug volume is dependent on the geometric shape, the surface wettability and the orientation (with respect to gravity) of the flow channel. The differential pressure required for slug motion primarily depends on the interfacial forces acting along the contact lines at the front and the back of the slug. At high gas Reynolds number, water is removed as a film or as drops depending on the flow channel surface wettability. The shape of growing drops at low and high Reynolds number can be described by a simple interfacial energy minimization model. Under flooding conditions, the fuel cell local current

  11. Properties of electrophoretic fractions of human embryonic kidney cells separated on space shuttle flight STS-8

    NASA Technical Reports Server (NTRS)

    Morrison, D. R.; Lewis, M. L.; Barlow, G. H.; Todd, P. W.; Kunze, M. E.; Sarnoff, B. E.; Li, Z. K.

    1985-01-01

    Suspensions of cultured primary human embryonic kidney cells were subjected to continuous flow electrophoresis on Space Shuttle flight STS-8. The objectives of the experiments were to obtain electrophoretically separated fractions of the original cell populations and to test these fractions for the amount and kind of urokinase (a kidney plasminogen activator that is used medically for digesting blood clots), the morphologies of cells in the individual fractions, and their cellular electrophoretic mobilities after separation and subsequent proliferation. Individual fractions were successfully cultured after return from orbit, and they were found to differ substantially from one another and from the starting sample with respect to all of these properties.

  12. Plasma membrane reorganization induced by tumor promoters in an epithelial cell line

    SciTech Connect

    PACKARD, BEVERLY S.; SAXTON, MICHAEL J.; BISSELL, MINA J.; KLEIN, MELVIN P.

    1984-01-01

    The effects of phorbol ester tumor promoters on the lateral diffusion in plasma membrane lipid environments were examined by the technique of fluorescence recovery after photobleaching. To this end, the probe collarein, a fluorescent lipid analog that has the property of exclusive localization in the plasma membrane, was synthesized. Measured decreases in three parameters [percentage of fluorescence bleached (30%), percentage of recovery (52%), and half-time for recovery (52%)] connoted the appearance of an immobile fraction upon exposure to tumor promoters. These data are consistent with lipid reorganization in response to a reorganization of the intra- and perimembranous macromolecular scaffolding upon the interaction of cells with tumor promoters. The idea of induced reorganization is supported by experiments in which cell shape change, brought about by either exposure to cytochalasin B or growth on matrices of collagen, fibronectin, or laminin, resulted in values in the fluorescence recovery after photobleaching technique similar to those with active phorbol esters.

  13. Accumulation of Multipotent Progenitor Cells on Polymethylpentene Membranes During Extracorporeal Membrane Oxygenation.

    PubMed

    Lehle, Karla; Friedl, Lucas; Wilm, Julius; Philipp, Alois; Müller, Thomas; Lubnow, Matthias; Schmid, Christof

    2016-06-01

    Multipotent progenitor cells were mobilized during pediatric extracorporeal membrane oxygenation (ECMO). We hypothesize that these cells also adhered onto polymethylpentene (PMP) fibers within the membrane oxygenator (MO) during adult ECMO support. Mononuclear cells were removed from the surface of explanted PMP-MOs (n = 16). Endothelial-like outgrowth and mesenchymal-like cells were characterized by flow cytometric analysis using different surface markers. Spindle-shaped attaching cells were identified early, but without proliferative activity. After long-term cultivation palisading type or cobblestone-type outgrowth cells with high proliferative activity appeared and were characterized as (i) leukocytoid CD45+/CD31+ (CD133+/VEGFR-II+/CD90+/CD14+/CD146dim/CD105dim); (ii) endothelial-like CD45-/CD31+ (VEGF-RII+/CD146+/CD105+/CD133-/CD14-/CD90-); and (iii) mesenchymal-like cells CD45-/CD31- (CD105+/CD90+/CD133dim/VEGFR-II-/CD146-/CD14-). The distribution of the cell populations depended on the MO and cultivation time. Endothelial-like cells formed capillary-like structures and did uptake Dil-acetylated low-density lipoprotein. Endothelial- and mesenchymal-like cells adhered on the surface of PMP-MOs. Further research is needed to identify the clinical relevance of these cells.

  14. Corona discharge in electroporation of cell membranes

    NASA Astrophysics Data System (ADS)

    Cramariuc, R.; Tudorache, A.; Popa, M. E.; Branduse, E.; Nisiparu, L.; Mitelut, A.; Turtoi, M. O.; Fotescu, L.

    2008-12-01

    The objective of the present work is to demonstrate that electrical corona discharge is very efficient in cellular membrane electroporation due to current pulses with sharp front (2-5 ns) and to the fact that corona discharge is associated with UV radiation and micro particles emission. A comparison between DC and AC at 800 Hz and a special waveform to corona application is presented. The comparison is analyzed by means of applying all these in the maceration process (electroplasmolysis) of red wine production and in the processes of different types of the microbes.

  15. Development of membrane electrode assembly for high temperature proton exchange membrane fuel cell by catalyst coating membrane method

    NASA Astrophysics Data System (ADS)

    Liang, Huagen; Su, Huaneng; Pollet, Bruno G.; Pasupathi, Sivakumar

    2015-08-01

    Membrane electrode assembly (MEA), which contains cathode and anode catalytic layer, gas diffusion layers (GDL) and electrolyte membrane, is the key unit of a PEMFC. An attempt to develop MEA for ABPBI membrane based high temperature (HT) PEMFC is conducted in this work by catalyst coating membrane (CCM) method. The structure and performance of the MEA are examined by scanning electron microscopy (SEM), electrochemical impedance spectroscopy (EIS) and I-V curve. Effects of the CCM preparation method, Pt loading and binder type are investigated for the optimization of the single cell performance. Under 160 °C and atmospheric pressure, the peak power density of the MEA, with Pt loading of 0.5 mg cm-2 and 0.3 mg cm-2 for the cathode and the anode, can reach 277 mW cm-2, while a current density of 620 A cm-2 is delivered at the working voltage of 0.4 V. The MEA prepared by CCM method shows good stability operating in a short term durability test: the cell voltage maintained at ∼0.45 V without obvious drop when operated at a constant current density of 300 mA cm-2 and 160 °C under ambient pressure for 140 h.

  16. Adenosine deaminase in cell transformation. Biophysical manifestation of membrane dynamics.

    PubMed

    Porat, N; Gill, D; Parola, A H

    1988-10-15

    Cell transformation is associated with a dramatic collapse of a graphic fingerprint characteristic of normal cells, as measured by phase fluorimetry. This is demonstrated on adenosine deaminase (ADA, EC 3.5.4.4), an established malignancy marker. ADA activity is known to decrease markedly in chick embryo fibroblasts (CEF) transformed by Rous sarcoma virus. The high affinity between the catalytic small subunit ADA (SS-ADA) and its membranal complexing protein (ADCP) (which abounds on the plasma membrane of CEF) allowed the hybridization of fluorescent labeled SS-ADA with native ADCP on CEF. Multifrequency differential phase fluorimetry responded remarkably to the state of this hybrid membrane protein. The transformation process is shown to have led to increased membrane fluidity and rotational mobility of ADCP as well as to its reduced availability to SS-ADA binding. The hypothesis of protein vertical sinking into the lipid core of the membrane is now given support by our spectroscopic data. Additional models are considered. A regulatory role is thus suggested for the complexing protein, which may also account for (a) reduced ADA activity in transformed cells and (b) detachment, exclusive to normal cells, upon addition of SS-ADA in excess.

  17. Boron Induces Hyperpolarization of Sunflower Root Cell Membranes and Increases Membrane Permeability to K+1

    PubMed Central

    Schon, Mary K.; Novacky, Anton; Blevins, Dale G.

    1990-01-01

    Although many studies have alluded to a role for boron (B) in membrane function, there is little evidence for a direct effect of B on the plasmalemma of higher plant cells. These studies were conducted to demonstrate, by electrophysiological techniques, a direct effect of B on the membrane potential (Em) of sunflower (Helianthus annuus [L.], cv Mammoth Grey Stripe) root tip cells and to determine if the response to B occurs rapidly enough to account for the previously observed effects of B on ion uptake. By inserting a glass microelectrode into an individual cell in the root tip, the Em of the cell was determined in basal salt medium (BSM), pH 6.0. The perfusion solution surrounding the root tissue was then changed to BSM + 50 micromolar H3BO3, pH 6.0. The exposure to B induced a significant plasmalemma hyperpolarization in sunflower root cells within 20 minutes. After just 3 minutes of exposure to B, the change in Em was already significantly different from the negligible change in Em observed over time in root cells never exposed to B. Membrane hyperpolarization could be caused by a stimulation of the proton pump or by a change in the conductance of one or more permeable ions. Since B has been shown to affect K+ uptake by plants, the electrophysiological techniques described above were used to determine if B has an effect on membrane permeability to K+, and could thereby lead to an increased diffusion potential. When sunflower root tips were pretreated in 50 micromolar B for 2 hours, cell membranes exhibited a significantly greater depolarization with each 10-fold increase in external [K+] than minus-B cells. Subsequent studies demonstrated that the depolarization due to increased external [K+] was also significantly greater when tissue was exposed to B at the same time as the 10-fold increase in [K+], indicating that the effect of B on K+ permeability was immediate. Analysis of sunflower root tips demonstrated that treatment in 50 micromolar B caused a

  18. Detection of Goodpasture antigen in fractions prepared from collagenase digests of human glomerular basement membrane.

    PubMed Central

    Fish, A J; Lockwood, M C; Wong, M; Price, R G

    1984-01-01

    Preparations of human glomerular basement membrane (GBM) were digested with collagenase, and a Goodpasture (GP) antigen rich pool from gel filtration column runs was identified by antibody inhibition radioimmunoassay. The components of the GP antigen pool were separated on polyacrylamide gels, and transferred to nitrocellulose sheets by the 'western' blotting technique. The blots were separately reacted with thirteen GP sera as primary antibody, followed by peroxidase labelled goat anti-human IgG and revealed 45-50K (two bands) and 25-28K (one-three bands) components. No corresponding reactivity was observed using convalescent GP sera or other control sera (normal human serum, rapidly progressive glomerulonephritis with or without pulmonary haemorrhage, and lupus erythematosus) as primary antibody. Images Fig. 3 PMID:6319059

  19. Edelfosine and miltefosine effects on lipid raft properties: membrane biophysics in cell death by antitumor lipids.

    PubMed

    Castro, Bruno M; Fedorov, Aleksander; Hornillos, Valentin; Delgado, Javier; Acuña, A Ulises; Mollinedo, Faustino; Prieto, Manuel

    2013-07-03

    Edelfosine (1-O-octadecyl-2-O-methyl-sn-glycero-phosphocholine) and miltefosine (hexadecylphosphocholine) are synthetic alkylphospholipids (ALPs) that are reported to selectively accumulate in tumor cell membranes, inducing Fas clustering and activation on lipid rafts, triggering apoptosis. However, the exact mechanism by which these lipids elicit these events is still not fully understood. Recent studies propose that their mode of action might be related with alterations of lipid rafts biophysical properties caused by these lipid drugs. To achieve a clear understanding of this mechanism, we studied the effects of pharmacologically relevant amounts of edelfosine and miltefosine in the properties of model and cellular membranes. The influence of these molecules on membrane order, lateral organization, and lipid rafts molar fraction and size were studied by steady-state and time-resolved fluorescence methods, Förster resonance energy transfer (FRET), confocal and fluorescence lifetime imaging microscopy (FLIM). We found that the global membrane and lipid rafts biophysical properties of both model and cellular membranes were not significantly affected by both the ALPs. Nonetheless, in model membranes, a mild increase in membrane fluidity induced by both alkyl lipids was detected, although this effect was more noticeable for edelfosine than miltefosine. This absence of drastic alterations shows for the first time that ALPs mode of action is unlikely to be directly linked to alterations of lipid rafts biophysical properties caused by these drugs. The biological implications of this result are discussed in the context of ALPs effects on lipid metabolism, mitochondria homeostasis modulation, and their relationship with tumor cell death.

  20. Transport parameters in the human red cell membrane: solute-membrane interactions of amides and ureas.

    PubMed

    Toon, M R; Solomon, A K

    1991-04-02

    We have studied the permeability of a series of hydrophilic amides and ureas through the red cell membrane by determining the three phenomenological coefficients which describe solute-membrane interaction: the hydraulic permeability (Lp), the phenomenological permeability coefficient (omega i) and the reflection coefficient (sigma i). In 55 experiments on nine solutes, we have determined that the reflection coefficient (after a small correction for solute permeation by membrane dissolution) is significantly less than 1.0 (P less than 0.003, t-test), which provides very strong evidence that solute and water fluxes are coupled as they cross the red cell membrane. It is proposed that the aqueous channel is a tripartite assembly, comprising H-bond exchange regions at both faces of the membrane, joined by a narrower sieve-specific region which crosses the lipid. The solutes bind to the H-bond exchange regions to exchange their solvation shell with the H-bonds of the channel; the existence of these regions is confirmed by the finding that the permeation of all the amides and ureas requires binding to well-characterized sites with Km values of 0.1-0.5 M. The sieve-specific regions provide the steric restraints which govern the passage of the solutes according to their size; their existence is shown by the findings that: (1) the reflection coefficient (actually the function [1-corrected sigma i]) is linearly dependent upon the solute molecular diameter; and (2) the permeability coefficient is linearly dependent upon solute molar volume. These several observations, taken together, provide strong arguments which lead to the conclusion that the amides and urea cross the red cell membrane in an aqueous pore.

  1. Biosynthesis of membrane dependent proteins in insect cell lysates: identification of limiting parameters for folding and processing.

    PubMed

    Merk, Helmut; Rues, Ralf-Bernhardt; Gless, Christine; Beyer, Kerstin; Dong, Fang; Dötsch, Volker; Gerrits, Michael; Bernhard, Frank

    2015-09-01

    G protein-coupled receptors, like many other membrane proteins, are notoriously difficult to synthesize in conventional cellular systems. Although expression in insect cells is considered the preferred technique for structural characterizations in particular, inefficient membrane translocation, instability, toxic effects and low yields still pose clear limitations for their production in living cells. Recent studies started to explore alternative strategies for the in vitro production of problematic membrane proteins in cell-free lysates in combination with supplied membranes. We provide a detailed study on the production efficiencies and quality of G protein-coupled receptors, Fab fragments and other proteins synthesized in insect cell lysates containing endogenous microsomes. Effects of different reaction kinetics, redox conditions and sample preparations on the specific activities of synthesized proteins have been analyzed. The extent of glycosylation, membrane translocation and percentages of ligand binding active fractions of synthesized protein samples have been determined. We provide strong evidence that membrane insertion of integral membrane proteins can represent a prime limiting factor for their preparative scale in vitro production. Improved expression protocols resulting into higher production rates yielded more active protein in case of Fab fragments, but not in case of the human endothelin B receptor.

  2. Membrane Mechanics of Endocytosis in Cells with Turgor

    PubMed Central

    Dmitrieff, Serge; Nédélec, François

    2015-01-01

    Endocytosis is an essential process by which cells internalize a piece of plasma membrane and material from the outside. In cells with turgor, pressure opposes membrane deformations, and increases the amount of force that has to be generated by the endocytic machinery. To determine this force, and calculate the shape of the membrane, we used physical theory to model an elastic surface under pressure. Accurate fits of experimental profiles are obtained assuming that the coated membrane is highly rigid and preferentially curved at the endocytic site. The forces required from the actin machinery peaks at the onset of deformation, indicating that once invagination has been initiated, endocytosis is unlikely to stall before completion. Coat proteins do not lower the initiation force but may affect the process by the curvature they induce. In the presence of isotropic curvature inducers, pulling the tip of the invagination can trigger the formation of a neck at the base of the invagination. Hence direct neck constriction by actin may not be required, while its pulling role is essential. Finally, the theory shows that anisotropic curvature effectors stabilize membrane invaginations, and the loss of crescent-shaped BAR domain proteins such as Rvs167 could therefore trigger membrane scission. PMID:26517669

  3. Membrane Mechanics of Endocytosis in Cells with Turgor.

    PubMed

    Dmitrieff, Serge; Nédélec, François

    2015-10-01

    Endocytosis is an essential process by which cells internalize a piece of plasma membrane and material from the outside. In cells with turgor, pressure opposes membrane deformations, and increases the amount of force that has to be generated by the endocytic machinery. To determine this force, and calculate the shape of the membrane, we used physical theory to model an elastic surface under pressure. Accurate fits of experimental profiles are obtained assuming that the coated membrane is highly rigid and preferentially curved at the endocytic site. The forces required from the actin machinery peaks at the onset of deformation, indicating that once invagination has been initiated, endocytosis is unlikely to stall before completion. Coat proteins do not lower the initiation force but may affect the process by the curvature they induce. In the presence of isotropic curvature inducers, pulling the tip of the invagination can trigger the formation of a neck at the base of the invagination. Hence direct neck constriction by actin may not be required, while its pulling role is essential. Finally, the theory shows that anisotropic curvature effectors stabilize membrane invaginations, and the loss of crescent-shaped BAR domain proteins such as Rvs167 could therefore trigger membrane scission.

  4. Red Blood Cell Membrane-Cloaked Nanoparticles For Drug Delivery

    NASA Astrophysics Data System (ADS)

    Carpenter, Cody Westcott

    Herein we describe the development of the Red Blood Cell coated nanoparticle, RBC-NP. Purified natural erythrocyte membrane is used to coat drug-loaded poly(lacticco-glycolic acid) (PLGA). Synthetic PLGA co-polymer is biocompatible and biodegradable and has already received US FDA approval for drug-delivery and diagnostics. This work looks specifically at the retention of immunosuppressive proteins on RBC-NPs, right-sidedness of natural RBC membranes interfacing with synthetic polymer nanoparticles, sustained and retarded drug release of RBC-NPs as well as further surface modification of RBC-NPs for increased targeting of model cancer cell lines.

  5. Microstructured Electrolyte Membranes to Improve Fuel Cell Performance

    NASA Astrophysics Data System (ADS)

    Wei, Xue

    Fuel cells, with the advantages of high efficiency, low greenhouse gas emission, and long lifetime are a promising technology for both portable power and stationary power sources. The development of efficient electrolyte membranes with high ionic conductivity, good mechanical durability and dense structure at low cost remains a challenge to the commercialization of fuel cells. This thesis focuses on exploring novel composite polymer membranes and ceramic electrolytes with the microstructure engineered to improve performance in direct methanol fuel cells (DMFCs) and solid oxide fuel cells (SOFCs), respectively. Polymer/particle composite membranes hold promise to meet the demands of DMFCs at lower cost. The structure of composite membranes was controlled by aligning proton conducting particles across the membrane thickness under an applied electric field. The field-induced structural changes caused the membranes to display an enhanced water uptake, proton conductivity, and methanol permeability in comparison to membranes prepared without an applied field. Although both methanol permeability and proton conductivity are enhanced by the applied field, the permeability increase is relatively lower than the proton conductivity improvement, which results in enhanced proton/methanol selectivity and improved DMFC performance. Apatite ceramics are a new class of fast ion conductors being studied as alternative SOFC electrolytes in the intermediate temperature range. An electrochemical/hydrothermal deposition method was developed to grow fully dense apatite membranes containing well-developed crystals with c-axis alignment to promote ion conductivity. Hydroxyapatite seed crystals were first deposited onto a metal substrate electrochemically. Subsequent ion substitution during the hydrothermal growth process promoted the formation of dense, fully crystalline films with microstructure optimal for ion transport. The deposition parameters were systematically investigated, such as

  6. New High-Temperature Membranes Developed for Proton Exchange Membrane Fuel Cells

    NASA Technical Reports Server (NTRS)

    Kinder, James D.

    2004-01-01

    Fuel cells are receiving a considerable amount of attention for potential use in a variety of areas, including the automotive industry, commercial power generation, and personal electronics. Research at the NASA Glenn Research Center has focused on the development of fuel cells for use in aerospace power systems for aircraft, unmanned air vehicles, and space transportation systems. These applications require fuel cells with higher power densities and better durability than what is required for nonaerospace uses. In addition, membrane cost is a concern for any fuel cell application. The most widely used membrane materials for proton exchange membrane (PEM) fuel cells are based on sulfonated perfluorinated polyethers, typically Nafion 117, Flemion, or Aciplex. However, these polymers are costly and do not function well at temperatures above 80 C. At higher temperatures, conventional membrane materials dry out and lose their ability to conduct protons, essential for the operation of the fuel cell. Increasing the operating temperature of PEM fuel cells from 80 to 120 C would significantly increase their power densities and enhance their durability by reducing the susceptibility of the electrode catalysts to carbon monoxide poisoning. Glenn's Polymers Branch has focused on developing new, low-cost membranes that can operate at these higher temperatures. A new series of organically modified siloxane (ORMOSIL) polymers were synthesized for use as membrane materials in a high-temperature PEM fuel cell. These polymers have an organic portion that can allow protons to transport through the polymer film and a cross-linked silica network that gives the polymers dimensional stability. These flexible xerogel polymer films are thermally stable, with decomposition onset as high as 380 C. Two types of proton-conducting ORMOSIL films have been produced: (1) NASA-A, which can coordinate many highly acid inorganic salts that facilitate proton conduction and (2) NASA-B, which has been

  7. Highly Water Resistant Anion Exchange Membrane for Fuel Cells.

    PubMed

    Yang, Zhengjin; Hou, Jianqiu; Wang, Xinyu; Wu, Liang; Xu, Tongwen

    2015-07-01

    For anion exchange membranes (AEMs), achieving efficient hydroxide conductivity without excessive hydrophilicity presents a challenge. Hence, new strategies for constructing mechanically strengthened and hydroxide conductive (especially at controlled humidity) membranes are critical for developing better AEMs. Macromolecular modification involving ylide chemistry (Wittig reaction) for the fabrication of novel AEMs with an interpenetrating polymer network structure is reported. The macromolecular modification is cost effective, facile, and based on a one-pot synthesis. AEM water uptake is reduced to 3.6 wt% and a high hydroxide conductivity (69.7 mS cm(-1) , 90 °C) is achieved simultaneously. More importantly, the membrane exhibits similar tensile strength (>35 MPa) and comparable flexibility in both dry and wet states. These AEMs could find further applications within anion exchange membrane fuel cells with low humidity or photoelectric assemblies.

  8. Rotational diffusion of band 3 in erythrocyte membranes. 1. Comparison of ghosts and intact cells.

    PubMed

    Matayoshi, E D; Jovin, T M

    1991-04-09

    The rotational diffusion of eosin-labeled 3 in human erythrocyte cells and hemoglobin-free ghosts at 37 degrees C has been studied in detail by polarized delayed luminescence. The time-resolved anisotropy with both cells and freshly prepared ghosts is similar, decaying with well-resolved rotational correlation times of 0.03, 0.2, and greater than or equal to 1 ms. Mild proteolytic removal of the water-soluble 41-kDa cytoplasmic domain of band 3 in ghosts results in a drastic increase in the fractional contributions of the two fastest depolarizing components. Our results, taken together with other data in the literature, imply that several classes of band 3 that differ greatly in mobility exist in ghosts and intact cells. The mobility of one class is hindered due to complexation with other membrane or cytoplasmic proteins mediated via the 41-kDa cytoplasmic domain. However, another class of band 3 molecules exists as homo-or heterooligomeric complexes larger than a dimer that are stabilized by hydrophobic interactions involving the intramembranal domain. Finally, the presence of the (previously undetected) 0.03-ms anisotropy component strongly suggests that a significant fraction of band 3 in both ghosts and intact cells is highly mobile and diffuses at the rate expected for a freely rotating dimer in the erythrocyte membrane.

  9. Lipid profiles of detergent resistant fractions of the plasma membrane in oat and rye in association with cold acclimation and freezing tolerance

    PubMed Central

    Takahashi, Daisuke; Imai, Hiroyuki; Kawamura, Yukio; Uemura, Matsuo

    2016-01-01

    Cold acclimation (CA) results in alteration of the plasma membrane (PM) lipid composition in plants, which plays a crucial role in the acquisition of freezing tolerance via membrane stabilization. Recent studies have indicated that PM structure is consistent with the fluid mosaic model but is laterally non-homogenous and contains microdomains enriched in sterols, sphingolipids and specific proteins. In plant cells, the function of these microdomains in relation to CA and freezing tolerance is not yet fully understood. The present study aimed to investigate the lipid compositions of detergent resistant fractions of the PM (DRM) which are considered to represent microdomains. They were prepared from leaves of low-freezing tolerant oat and high-freezing tolerant rye. The DRMs contained higher proportions of sterols, sphingolipids and saturated phospholipids than the PM. In particular, one of the sterol lipid classes, acylated sterylglycoside, was the predominant sterol in oat DRM while rye DRM contained free sterol as the major sterol. Oat and rye showed different patterns (or changes) of sterols and 2-hydroxy fatty acids of sphingolipids of DRM lipids during CA. Taken together, these results suggest that CA-induced changes of lipid classes and molecular species in DRMs are associated with changes in the thermodynamic properties and physiological functions of microdomains during CA and hence, influence plant freezing tolerance. PMID:26904981

  10. Lactic acid fermentation in cell-recycle membrane bioreactor.

    PubMed

    Choudhury, B; Swaminathan, T

    2006-02-01

    Traditional lactic acid fermentation suffers from low productivity and low product purity. Cell-recycle fermentation has become one of the methods to obtain high cell density, which results in higher productivity. Lactic acid fermentation was investigated in a cell-recycle membrane bioreactor at higher substrate concentrations of 100 and 120 g/dm3. A maximum cell density of 145 g/dm3 and a maximum productivity of 34 g/(dm3.h) were achieved in cell-recycle fermentation. In spite of complete consumption of substrate, there was a continuous increase in cell density in cell-recycle fermentation. Control of cell density in cell-recycle fermentation was attempted by cell bleeding and reduction in yeast extract concentration.

  11. Membrane patterned by pulsed laser micromachining for proton exchange membrane fuel cell with sputtered ultra-low catalyst loadings

    NASA Astrophysics Data System (ADS)

    Cuynet, S.; Caillard, A.; Kaya-Boussougou, S.; Lecas, T.; Semmar, N.; Bigarré, J.; Buvat, P.; Brault, P.

    2015-12-01

    Proton exchange membranes were nano- and micro-patterned on their cathode side by pressing them against stainless steel molds previously irradiated by a Ti:Sapphire femtosecond laser. The membranes were associated to ultra-low loaded thin catalytic layers (25 μgPt cm-2) prepared by plasma magnetron sputtering. The Pt catalyst was sputtered either on the membrane or on the porous electrode. The fuel cell performance in dry conditions were found to be highly dependent on the morphology of the membrane surface. When nanometric ripples covered by a Pt catalyst were introduced on the surface of the membrane, the fuel cell outperformed the conventional one with a flat membrane. By combining nano- and micro-patterns (nanometric ripples and 11-24 μm deep craters), the performance of the cells was clearly enhanced. The maximum power density achieved by the fuel cell was multiplied by a factor of 3.6 (at 50 °C and 3 bar): 438 mW cm-2 vs 122 mW cm-2. This improvement is due to high catalyst utilization with a high membrane conductivity. When Pt is sputtered on the porous electrode (and not on the membrane), the contribution of the patterned membrane to the fuel cell efficiency was less significant, except in the presence of nanometric ripples. This result suggests that the patterning of the membrane must be consistent with the way the catalyst is synthesized, on the membrane or on the porous electrode.

  12. Radiation effects on membranes - 1. Cellular permeability and cell survival

    SciTech Connect

    Khare, S.; Jayakumar, A.; Trivedi, A.; Kesavan, P.C.; Prasad, R.

    1982-05-01

    The effect of various doses of ..gamma.. radiation (5-60 krad) on the membrane permeability and cell survival of Candida albicans, a pathogenic yeast, was investigated. A reduction in the cell survival and in the accumulation of amino acids (proline, glycine, lysine, and glutamic acid) was observed following irradiation. The rate of oxygen uptake, which is often associated with transport, was also reduced. There was no damage to available sulfhydryl groups following the exposure of cells to various doses of ..gamma.. radiation. The membrane lipid composition of C. albicans cells can be altered by growing them in alkanes of varying chain lengths. The effects of such altered lipid composition on radiosensitivity was examined. It was observed that C. albicans cells with altered lipid content acquire resistance to ..gamma.. radiation.

  13. Evidence for Bidirectional Endocannabinoid Transport across Cell Membranes*

    PubMed Central

    Chicca, Andrea; Marazzi, Janine; Nicolussi, Simon; Gertsch, Jürg

    2012-01-01

    Despite extensive research on the trafficking of anandamide (AEA) across cell membranes, little is known about the membrane transport of other endocannabinoids, such as 2-arachidonoylglycerol (2-AG). Previous studies have provided data both in favor and against a cell membrane carrier-mediated transport of endocannabinoids, using different methodological approaches. Because AEA and 2-AG undergo rapid and almost complete intracellular hydrolysis, we employed a combination of radioligand assays and absolute quantification of cellular and extracellular endocannabinoid levels. In human U937 leukemia cells, 100 nm AEA and 1 μm 2-AG were taken up through a fast and saturable process, reaching a plateau after 5 min. Employing differential pharmacological blockage of endocannabinoid uptake, breakdown, and interaction with intracellular binding proteins, we show that eicosanoid endocannabinoids harboring an arachidonoyl chain compete for a common membrane target that regulates their transport, whereas other N-acylethanolamines did not interfere with AEA and 2-AG uptake. By combining fatty acid amide hydrolase or monoacyl glycerol lipase inhibitors with hydrolase-inactive concentrations of the AEA transport inhibitors UCM707 (1 μm) and OMDM-2 (5 μm), a functional synergism on cellular AEA and 2-AG uptake was observed. Intriguingly, structurally unrelated AEA uptake inhibitors also blocked the cellular release of AEA and 2-AG. We show, for the first time, that UCM707 and OMDM-2 inhibit the bidirectional movement of AEA and 2-AG across cell membranes. Our findings suggest that a putative endocannabinoid cell membrane transporter controls the cellular AEA and 2-AG trafficking and metabolism. PMID:22879589

  14. Cell cycle dependent changes in the plasma membrane organization of mammalian cells.

    PubMed

    Denz, Manuela; Chiantia, Salvatore; Herrmann, Andreas; Mueller, Peter; Korte, Thomas; Schwarzer, Roland

    2017-03-01

    Lipid membranes are major structural elements of all eukaryotic and prokaryotic organisms. Although many aspects of their biology have been studied extensively, their dynamics and lateral heterogeneity are still not fully understood. Recently, we observed a cell-to-cell variability in the plasma membrane organization of CHO-K1 cells (Schwarzer et al., 2014). We surmised that cell cycle dependent changes of the individual cells from our unsynchronized cell population account for this phenomenon. In the present study, this hypothesis was tested. To this aim, CHO-K1 cells were arrested in different cell cycle phases by chemical treatments, and the order of their plasma membranes was determined by various fluorescent lipid analogues using fluorescence lifetime imaging microscopy. Our experiments exhibit significant differences in the membrane order of cells arrested in the G2/M or S phase compared to control cells. Our single-cell analysis also enabled the specific selection of mitotic cells, which displayed a significant increase of the membrane order compared to the control. In addition, the lipid raft marker GPImYFP was used to study the lateral organization of cell cycle arrested cells as well as mitotic cells and freely cycling samples. Again, significant differences were found between control and arrested cells and even more pronounced between control and mitotic cells. Our data demonstrate a direct correlation between cell cycle progression and plasma membrane organization, underlining that cell-to-cell heterogeneities of membrane properties have to be taken into account in cellular studies especially at the single-cell level.

  15. Fractionation of Plant Bioactives from Black Carrots (Daucus carota subspecies sativus varietas atrorubens Alef.) by Adsorptive Membrane Chromatography and Analysis of Their Potential Anti-Diabetic Activity.

    PubMed

    Esatbeyoglu, Tuba; Rodríguez-Werner, Miriam; Schlösser, Anke; Liehr, Martin; Ipharraguerre, Ignacio; Winterhalter, Peter; Rimbach, Gerald

    2016-07-27

    Black and purple carrots have attracted interest as colored extracts for coloring food due to their high content of anthocyanins. This study aimed to investigate the polyphenol composition of black carrots. Particularly, the identification and quantification of phenolic compounds of the variety Deep Purple carrot (DPC), which presents a very dark color, was performed by HPLC-PDA and HPLC-ESI-MS(n) analyses. The separation of polyphenols from a DPC XAD-7 extract into an anthocyanin fraction (AF) and co-pigment fraction (CF; primarily phenolic acids) was carried out by membrane chromatography. Furthermore, possible anti-diabetic effects of the DPC XAD-7 extract and its AF and CF were determined. DPC samples (XAD-7, CF, and AF) inhibited α-amylase and α-glucosidase in a dose-dependent manner. Moreover, DPC XAD-7 and chlorogenic acid, but not DPC CF and DPC AF, caused a moderate inhibition of intestinal glucose uptake in Caco-2 cells. However, DPC samples did not affect glucagon-like peptide-1 (GLP-1) secretion and dipeptidyl peptidase IV (DPP-4) activity. Overall, DPC exhibits an inhibitory effect on α-amylase and α-glucosidase activity and on cellular glucose uptake indicating potential anti-diabetic properties.

  16. Electronic circuit model for proton exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Yu, Dachuan; Yuvarajan, S.

    The proton exchange membrane (PEM) fuel cell is being investigated as an alternate power source for various applications like transportation and emergency power supplies. The paper presents a novel circuit model for a PEM fuel cell that can be used to design and analyze fuel cell power systems. The PSPICE-based model uses bipolar junction transistors (BJTs) and LC elements available in the PSPICE library with some modification. The model includes the phenomena like activation polarization, ohmic polarization, and mass transport effect present in a PEM fuel cell. The static and dynamic characteristics obtained through simulation are compared with experimental results obtained on a commercial fuel cell module.

  17. PIG7 promotes leukemia cell chemosensitivity via lysosomal membrane permeabilization

    PubMed Central

    Niu, Ting; Wu, Yu; Li, Jianjun; Wang, Fangfang; Zheng, Yuhuan; Liu, Ting

    2016-01-01

    PIG7 localizes to lysosomal membrane in leukemia cells. Our previous work has shown that transduction of pig7 into a series of leukemia cell lines did not result in either apoptosis or differentiation of most tested cell lines. Interestingly, it did significantly sensitize these cell lines to chemotherapeutic drugs. Here, we further investigated the mechanism underlying pig7-induced improved sensitivity of acute leukemia cells to chemotherapy. Our results demonstrated that the sensitization effect driven by exogenous pig7 was more effective in drug-resistant leukemia cell lines which had lower endogenous pig7 expression. Overexpression of pig7 did not directly activate the caspase apoptotic pathway, but decreased the lysosomal stability. The expression of pig7 resulted in lysosomal membrane permeabilization (LMP) and lysosomal protease (e.g. cathepsin B, D, L) release. Moreover, we also observed increased reactive oxygen species (ROS) and decreased mitochondrial membrane potential (ΔΨm) induced by pig7. Some autophagy markers such as LC3I/II, ATG5 and Beclin-1, and necroptosis maker MLKL were also stimulated. However, intrinsic antagonism such as serine/cysteine protease inhibitors Spi2A and Cystatin C prevented downstream effectors from triggering leukemia cells, which were only on the “verge of apoptosis”. When combined with chemotherapy, LMP increased and more proteases were released. Once this process was beyond the limit of intrinsic antagonism, it induced programmed cell death cooperatively via caspase-independent and caspase-dependent pathways. PMID:26716897

  18. PIG7 promotes leukemia cell chemosensitivity via lysosomal membrane permeabilization.

    PubMed

    Liu, Jiazhuo; Peng, Leiwen; Niu, Ting; Wu, Yu; Li, Jianjun; Wang, Fangfang; Zheng, Yuhuan; Liu, Ting

    2016-01-26

    PIG7 localizes to lysosomal membrane in leukemia cells. Our previous work has shown that transduction of pig7 into a series of leukemia cell lines did not result in either apoptosis or differentiation of most tested cell lines. Interestingly, it did significantly sensitize these cell lines to chemotherapeutic drugs. Here, we further investigated the mechanism underlying pig7-induced improved sensitivity of acute leukemia cells to chemotherapy. Our results demonstrated that the sensitization effect driven by exogenous pig7 was more effective in drug-resistant leukemia cell lines which had lower endogenous pig7 expression. Overexpression of pig7 did not directly activate the caspase apoptotic pathway, but decreased the lysosomal stability. The expression of pig7 resulted in lysosomal membrane permeabilization (LMP) and lysosomal protease (e.g. cathepsin B, D, L) release. Moreover, we also observed increased reactive oxygen species (ROS) and decreased mitochondrial membrane potential (ΔΨm) induced by pig7. Some autophagy markers such as LC3I/II, ATG5 and Beclin-1, and necroptosis maker MLKL were also stimulated. However, intrinsic antagonism such as serine/cysteine protease inhibitors Spi2A and Cystatin C prevented downstream effectors from triggering leukemia cells, which were only on the "verge of apoptosis". When combined with chemotherapy, LMP increased and more proteases were released. Once this process was beyond the limit of intrinsic antagonism, it induced programmed cell death cooperatively via caspase-independent and caspase-dependent pathways.

  19. The role of cell membranes in the regulation of lignification in pine cells

    NASA Technical Reports Server (NTRS)

    Hendrix, D. L.

    1978-01-01

    The identity of pine cell membranes bearing PAL enzyme activity, the isolation of a plasma membrane preparation from pine cells for testing as a regulatory barrier in lignification, and the measurement of the geopotential effect in pine stems are presented. A model to describe and predict the interaction of gravity and lignification of higher plants was developed.

  20. Brucella fractions behave as nonspecific mitogens and polyclonal B-cell activators for human lymphocytes.

    PubMed Central

    Vendrell, J P; Rabesandratana, H; Huguet, M F; Cannat, A; Serre, A

    1985-01-01

    Two lipid-A-free fractions which were extracted from Brucella melitensis and were designated PI and SF stimulated human unsensitized mononuclear cells to proliferate and to secrete immunoglobulins. Both of these effects were observed in cultures of peripheral blood, tonsils, and cord blood lymphocytes. Neither B cells nor T cells alone proliferated in the presence of these fractions, whereas the proliferative response of T cells plus B cells was largely independent of accessory cells. Polyclonal activation was estimated by counting the cells which secreted immunoglobulins of different isotypes into culture supernatants. This phenomenon was strongly T dependent. PMID:3876286

  1. Time courses of mammalian cell electropermeabilization observed by millisecond imaging of membrane property changes during the pulse.

    PubMed Central

    Gabriel, B; Teissié, J

    1999-01-01

    Time courses of electropermeabilization were analyzed during the electric field application using a rapid fluorescent imaging system. Exchanges of calcium ions through electropermeabilized membrane of Chinese hamster ovary cells were found to be asymmetrical. Entry of calcium ions during a millisecond pulse occurred on the anode-facing cell hemisphere. Entry through the region facing the cathode was observed only after the pulse. Leakage of intracellular calcium ions from electropermeabilized cell in low-calcium content medium was observed only from the anode-facing side. The exchanges during the pulse were mostly due to diffusion-driven processes, i.e., governed by the concentration gradient. Interaction of propidium iodide, a dye sensitive to the structural alteration of membrane, with cell membrane was asymmetrical during electropermeabilization. Localized enhancement of the dye fluorescence was observed during and after the pulsation on the cell surface. Specific staining of a limited anode-facing part of the membrane was observed as soon as the pulse was applied. The membrane fluorescence level increased during and immediately after the pulse whereas the geometry of the staining was unchanged. The membrane regions stained by propidium iodide were the same as those where calcium exchanges occurred. The fraction of the membrane on which structural alterations occurred was defined by the field strength. The density of defects was governed by the pulse duration. Electropermeabilization is a localized but asymmetrical process. The membrane defects are created unequally on the two cell sides during the pulse, implying a vectorial effect of the electric field on the membrane. PMID:10096909

  2. Membrane with internal passages to permit fluid flow and an electrochemical cell containing the same

    NASA Technical Reports Server (NTRS)

    Cisar, Alan J. (Inventor); Gonzalez-Martin, Anuncia (Inventor); Hitchens, G. Duncan (Inventor); Murphy, Oliver J. (Inventor)

    1997-01-01

    The invention provides an improved proton exchange membrane for use in electrochemical cells having internal passages parallel to the membrane surface, an apparatus and process for making the membrane, membrane and electrode assemblies fabricated using the membrane, and the application of the membrane and electrode assemblies to a variety of devices, both electrochemical and otherwise. The passages in the membrane extend from one edge of the membrane to another and allow fluid flow through the membrane and give access directly to the membrane for purposes of hydration.

  3. Macrophages engulf endothelial cell membrane particles preceding pupillary membrane capillary regression.

    PubMed

    Poché, Ross A; Hsu, Chih-Wei; McElwee, Melissa L; Burns, Alan R; Dickinson, Mary E

    2015-07-01

    Programmed capillary regression and remodeling are essential developmental processes. However, the cellular and molecular mechanisms that regulate vessel regression are only the beginning to be understood. Here, using in vivo, dynamic, confocal imaging of mouse transgenic reporters as well as static confocal and electron microscopy, we studied the embryonic development and postnatal regression of the transient mouse pupillary membrane (PM) vasculature. This approach allowed us to directly observe the precise temporal sequence of cellular events preceding and during the elimination of the PM from the mouse eye. Imaging of Tcf/Lef-H2B::GFP Wnt-reporter mice uncovered that, unlike the hyaloid vasculature of the posterior eye, a PM endothelial cell (EC) Wnt/β-catenin response is unlikely to be part of the regression mechanism. Live imaging of EC and macrophage dynamics revealed highly active Csf1r-GFP+ macrophages making direct contact with the Flk1-myr::mCherry+ vessel surface and with membrane protrusions or filopodia extending from the ECs. Flk1-myr::mCherry+ EC membrane particles were observed on and around ECs as well as within macrophages. Electron microscopy studies confirmed that they were in phagosomes within macrophages, indicating that the macrophages engulfed the membrane particles. Interestingly, EC plasma membrane uptake by PM macrophages did not correlate with apoptosis and was found shortly after vessel formation at mid-gestation stages in the embryo; long before vessel regression begins during postnatal development. Additionally, genetic ablation of macrophages showed that EC membrane particles were still shed in the absence of macrophages suggesting that macrophages do not induce the formation or release of EC microparticles. These studies have uncovered a novel event during programmed capillary regression in which resident macrophages scavenge endothelial cell microparticles released from the PM vessels. This finding suggests that there may be an

  4. Changes in Band 3 oligomeric state precede cell membrane phospholipid loss during blood bank storage of red blood cells

    PubMed Central

    Karon, Brad S.; Hoyer, James D.; Stubbs, James R.; Thomas, David D.

    2013-01-01

    BACKGROUND Lipid loss in the form of vesicles contributes to the red blood cell (RBC) storage lesion, and this loss of lipid is correlated with changes in membrane protein function. Sensitive spectroscopic techniques were used to measure changes in Band 3 oligomeric state during storage of RBCs, compared to metabolic changes and phospholipid loss. The aim of the study was to determine whether changes in the macromolecular organization of membrane proteins occur before, coincident with, or after lipid loss during RBC storage. STUDY DESIGN AND METHODS Five RBC units were collected from normal volunteers and stored under standard blood bank conditions, and both metabolic changes and lipid loss were measured by multiple assays. Band 3 oligomeric state was assessed by time-resolved phosphorescence anisotropy and fluorescence resonance energy transfer of eosin-5-maleimide–labeled RBC ghosts. RESULTS Extracellular pH decreased and extracellular potassium increased rapidly during cold storage of blood. Band 3 on the RBC membrane exhibited a shift from small to large oligomers early in the storage period and before detectable loss of phospholipid from the RBC membrane. The immobilized fraction of Band 3, that which is tethered to the cytoskeletal network via spectrin and ankyrin, did not change during cold storage. CONCLUSION Our results demonstrate that changes in the macromolecular organization of membrane proteins on the RBC occur early in storage, and these changes may induce phospholipid loss, irreversible morphologic changes, and loss of function during RBC storage. PMID:19389033

  5. Durable, Low-cost, Improved Fuel Cell Membranes

    SciTech Connect

    Chris Roger; David Mountz; Wensheng He; Tao Zhang

    2011-03-17

    The development of low cost, durable membranes and membranes electrode assemblies (MEAs) that operate under reduced relative humidity (RH) conditions remain a critical challenge for the successful introduction of fuel cells into mass markets. It was the goal of the team lead by Arkema, Inc. to address these shortages. Thus, this project addresses the following technical barriers from the fuel cells section of the Hydrogen Fuel Cells and Infrastructure Technologies Program Multi-Year Research, Development and Demonstration Plan: (A) Durability (B) Cost Arkema’s approach consisted of using blends of polyvinylidenefluoride (PVDF) and proprietary sulfonated polyelectrolytes. In the traditional approach to polyelectrolytes for proton exchange membranes (PEM), all the required properties are “packaged” in one macromolecule. The properties of interest include proton conductivity, mechanical properties, durability, and water/gas transport. This is the case, for example, for perfluorosulfonic acid-containing (PFSA) membranes. However, the cost of these materials is high, largely due to the complexity and the number of steps involved in their synthesis. In addition, they suffer other shortcomings such as mediocre mechanical properties and insufficient durability for some applications. The strength and originality of Arkema’s approach lies in the decoupling of ion conductivity from the other requirements. Kynar® PVDF provides an exceptional combination of properties that make it ideally suited for a membrane matrix (Kynar® is a registered trademark of Arkema Inc.). It exhibits outstanding chemical resistance in highly oxidative and acidic environments. In work with a prior grant, a membrane known as M41 was developed by Arkema. M41 had many of the properties needed for a high performance PEM, but had a significant deficiency in conductivity at low RH. In the first phase of this work, the processing parameters of M41 were explored as a means to increase its proton

  6. Identification of proteins from a cell wall fraction of the diatom Thalassiosira pseudonana: insights into silica structure formation.

    PubMed

    Frigeri, Luciano G; Radabaugh, Timothy R; Haynes, Paul A; Hildebrand, Mark

    2006-01-01

    Diatoms are unicellular eucaryotic algae with cell walls containing silica, intricately and ornately structured on the nanometer scale. Overall silica structure is formed by expansion and molding of the membrane-bound silica deposition vesicle. Although molecular details of silica polymerization are being clarified, we have limited insight into molecular components of the silica deposition vesicle, particularly of membrane-associated proteins that may be involved in structure formation. To identify such proteins, we refined existing procedures to isolate an enriched cell wall fraction from the diatom Thalassiosira pseudonana, the first diatom with a sequenced genome. We applied tandem mass spectrometric analysis to this fraction, identifying 31 proteins for further evaluation. mRNA levels for genes encoding these proteins were monitored during synchronized progression through the cell cycle and compared with two previously identified silaffin genes (involved in silica polymerization) having distinct mRNA patterns that served as markers for cell wall formation. Of the 31 proteins identified, 10 had mRNA patterns that correlated with the silaffins, 13 had patterns that did not, and seven had patterns that correlated but also showed additional features. The possible involvements of these proteins in cell wall synthesis are discussed. In particular, glutamate acetyltransferase was identified, prompting an analysis of mRNA patterns for other genes in the polyamine biosynthesis pathway and identification of those induced during cell wall synthesis. Application of a specific enzymatic inhibitor for ornithine decarboxylase resulted in dramatic alteration of silica structure, confirming the involvement of polyamines and demonstrating that manipulation of proteins involved in cell wall synthesis can alter structure. To our knowledge, this is the first proteomic analysis of a diatom, and furthermore we identified new candidate genes involved in structure formation and

  7. Thermoase-Derived Flaxseed Protein Hydrolysates and Membrane Ultrafiltration Peptide Fractions Have Systolic Blood Pressure-Lowering Effects in Spontaneously Hypertensive Rats

    PubMed Central

    Nwachukwu, Ifeanyi D.; Girgih, Abraham T.; Malomo, Sunday A.; Onuh, John O.; Aluko, Rotimi E.

    2014-01-01

    Thermoase-digested flaxseed protein hydrolysate (FPH) samples and ultrafiltration membrane-separated peptide fractions were initially evaluated for in vitro inhibition of angiotensin I-converting enzyme (ACE) and renin activities. The two most active FPH samples and their corresponding peptide fractions were subsequently tested for in vivo antihypertensive activity in spontaneously hypertensive rats (SHR). The FPH produced with 3% thermoase digestion showed the highest ACE- and renin-inhibitory activities. Whereas membrane ultrafiltration resulted in significant (p < 0.05) increases in ACE inhibition by the <1 and 1–3 kDa peptides, only a marginal improvement in renin-inhibitory activity was observed for virtually all the samples after membrane ultrafiltration. The FPH samples and membrane fractions were also effective in lowering systolic blood pressure (SBP) in SHR with the largest effect occurring after oral administration (200 mg/kg body weight) of the 1–3 kDa peptide fraction of the 2.5% FPH and the 3–5 kDa fraction of the 3% FPH. Such potent SBP-lowering capacity indicates the potential of flaxseed protein-derived bioactive peptides as ingredients for the formulation of antihypertensive functional foods and nutraceuticals. PMID:25302619

  8. Sol-gel based silica electrodes for inorganic membrane direct methanol fuel cells

    NASA Astrophysics Data System (ADS)

    Kim, Hyea; Kohl, Paul A.

    Inorganic glass electrodes are of interest for use with inorganic proton exchange membranes for direct methanol fuel cells. Platinum-ruthenium glass electrodes (PtRu/C-SiO 2) have been prepared by incorporating the PtRu/C nanoparticles into a silica-based matrix. The SiO 2 matrix was synthesized through the sol-gel reaction of 3-(trihydroxysilyl)-1-propanesulfonic acid (3TPS) and 3-glycidoxypropyltrimethoxysilane (GPTMS). The distribution of the PtRu/C particles can be controlled by changing the properties of the gel matrix. The effect of gelation time, mole fraction of reactants within the sol, curing temperature, and glass ionomer content were investigated. The adhesion of the catalyst layer on the membrane, catalytic activity for methanol oxidation, and inhibition of methanol permeation through the membrane have been characterized and optimized. The electroless deposition of PtRu onto the PtRu/C nanoparticles was performed to increase the sheet conductivity of the electrode. It was found that the electrolessly deposited metal improved the catalytic activity for methanol oxidation and decreased the methanol cross-over. The methanol fuel cell performance using the inorganic membrane electrode assembly was 236 μA cm -2 at 0.4 V and was stable for more than 10 days.

  9. Purification of human adipose-derived stem cells from fat tissues using PLGA/silk screen hybrid membranes.

    PubMed

    Chen, Da-Chung; Chen, Li-Yu; Ling, Qing-Dong; Wu, Meng-Hsueh; Wang, Ching-Tang; Suresh Kumar, S; Chang, Yung; Munusamy, Murugan A; Alarfajj, Abdullah A; Wang, Han-Chow; Hsu, Shih-Tien; Higuchi, Akon

    2014-05-01

    The purification of human adipose-derived stem cells (hADSCs) from human adipose tissue cells (stromal vascular fraction) was investigated using membrane filtration through poly(lactide-co-glycolic acid)/silk screen hybrid membranes. Membrane filtration methods are attractive in regenerative medicine because they reduce the time required to purify hADSCs (i.e., less than 30 min) compared with conventional culture methods, which require 5-12 days. hADSCs expressing the mesenchymal stem cell markers CD44, CD73, and CD90 were concentrated in the permeation solution from the hybrid membranes. Expression of the surface markers CD44, CD73, and CD99 on the cells in the permeation solution from the hybrid membranes, which were obtained using 18 mL of feed solution containing 50 × 10⁴ cells, was statistically significantly higher than that of the primary adipose tissue cells, indicating that the hADSCs can be purified in the permeation solution by the membrane filtration method. Cells expressing the stem cell-associated marker CD34 could be successfully isolated in the permeation solution, whereas CD34⁺ cells could not be purified by the conventional culture method. The hADSCs in the permeation solution demonstrated a superior capacity for osteogenic differentiation based on their alkali phosphatase activity, their osterix gene expression, and the results of mineralization analysis by Alizarin Red S and von Kossa staining compared with the cells from the suspension of human adipose tissue. These results suggest that the hADSCs capable of osteogenic differentiation preferentially permeate through the hybrid membranes.

  10. Single cell electric impedance topography: mapping membrane capacitance.

    PubMed

    Dharia, Sameera; Ayliffe, Harold E; Rabbitt, Richard D

    2009-12-07

    Single-cell electric impedance topography (sceTopo), a technique introduced here, maps the spatial distribution of capacitance (i.e. displacement current) associated with the membranes of isolated, living cells. Cells were positioned in the center of a circular recording chamber surrounded by eight electrodes. Electrodes were evenly distributed on the periphery of the recording chamber. Electric impedance measured between adjacent electrode pairs (10 kHz-5 MHz) was used to construct topographical maps of the spatial distribution of membrane capacitance. Xenopus Oocytes were used as a model cell to develop sceTopo because these cells consist of two visually distinguishable hemispheres, each with distinct membrane composition and structure. Results showed significant differences in the imaginary component of the impedance between the two oocyte hemispheres. In addition, the same circumferential array was used to map the size of the extracellular electrical shunt path around the cell, providing a means to estimate the location and shape of the cell in the recording chamber.

  11. Single cell electric impedance topography: Mapping membrane capacitance

    PubMed Central

    Dharia, Sameera; Ayliffe, Harold E.

    2010-01-01

    Single-cell electric impedance topography (sceTopo), a technique introduced here, maps the spatial distribution of capacitance (i.e. displacement current) associated with the membranes of isolated, living cells. Cells were positioned in the center of a circular recording chamber surrounded by eight electrodes. Electrodes were evenly distributed on the periphery of the recording chamber. Electric impedance measured between adjacent electrode pairs (10 kHz–5 MHz) was used to construct topographical maps of the spatial distribution of membrane capacitance. Xenopus Oocytes were used as a model cell to develop sceTopo because these cells consist of two visually distinguishable hemispheres, each with distinct membrane composition and structure. Results showed significant differences in the imaginary component of the impedance between the two oocyte hemispheres. In addition, the same circumferential array was used to map the size of the extracellular electrical shunt path around the cell, providing a means to estimate the location and shape of the cell in the recording chamber. PMID:19904403

  12. Membrane Stabilization and Detoxification of Acetaminophen-Mediated Oxidative Onslaughts in the Kidneys of Wistar Rats by Standardized Fraction of Zea mays L. (Poaceae), Stigma maydis

    PubMed Central

    Sabiu, S.; O'Neill, F. H.

    2016-01-01

    This study evaluated membrane stabilization and detoxification potential of ethyl acetate fraction of Zea mays L., Stigma maydis in acetaminophen-induced oxidative onslaughts in the kidneys of Wistar rats. Nephrotoxic rats were orally pre- and posttreated with the fraction and vitamin C for 14 days. Kidney function, antioxidative and histological analyses were thereafter evaluated. The acetaminophen-mediated significant elevations in the serum concentrations of creatinine, urea, uric acid, sodium, potassium, and tissue levels of oxidized glutathione, protein-oxidized products, lipid peroxidized products, and fragmented DNA were dose-dependently assuaged in the fraction-treated animals. The fraction also markedly improved creatinine clearance rate, glutathione, and calcium concentrations as well as activities of superoxide dismutase, catalase, glutathione reductase, and glutathione peroxidase in the nephrotoxic rats. These improvements may be attributed to the antioxidative and membrane stabilization activities of the fraction. The observed effects compared favorably with that of vitamin C and are informative of the fraction's ability to prevent progression of renal pathological conditions and preserve kidney functions as evidently supported by the histological analysis. Although the effects were prominently exhibited in the fraction-pretreated groups, the overall data from the present findings suggest that the fraction could prevent or extenuate acetaminophen-mediated oxidative renal damage via fortification of antioxidant defense mechanisms. PMID:27579048

  13. Proton exchange membrane fuel cell diagnosis by spectral characterization of the electrochemical noise

    NASA Astrophysics Data System (ADS)

    Maizia, R.; Dib, A.; Thomas, A.; Martemianov, S.

    2017-02-01

    Electrochemical noise analysis (ENA) has been performed for the diagnosis of proton-exchange membrane fuel cell (PEMFC) under various operating conditions. Its interest is related with the possibility of a non-invasive on-line diagnosis of a commercial fuel cell. A methodology of spectral analysis has been developed and an evaluation of the stationarity of the signal has been proposed. It has been revealed that the spectral signature of fuel cell, is a linear slope with a fractional power dependence 1/fα where α = 2 for different relative humidities and current densities. Experimental results reveal that the electrochemical noise is sensitive to the water management, especially under dry conditions. At RHH2 = 20% and RHair = 20%, spectral analysis shows a three linear slopes signature on the spectrum at low frequency range (f < 100 Hz). This results indicates that power spectral density, calculated thanks to FFT, can be used for the detection of an incorrect fuel cell water balance.

  14. Binding of white spot syndrome virus to Artemia sp. cell membranes.

    PubMed

    Feng, Shuying; Li, Guangda; Feng, Wenpo; Huang, Jie

    2013-10-01

    Using differential velocity centrifugation, cell membranes of Artemia sp. were prepared, and their binding to white spot syndrome virus (WSSV) was analyzed in vitro. The results indicated that WSSV can specifically bind to Artemia cell membranes, and that WSSV receptor very likely existed in this membrane, which suggested that Artemia sp. may be a reservoir of WSSV. This study investigated the specific WSSV binding site by performing competitive inhibition experiments using shrimp gill cell membranes to bind WSSV to Artemia cell membranes. The results showed that shrimp gill cell membranes had a distinct inhibition effect on the specific binding of Artemia cell membranes to WSSV. Thus, potentially similar WSSV receptors or binding sites existed on Artemia sp. cell membranes and shrimp gill cell membranes. Taken together, these findings may provide experimental basis for the development of an effective approach to controlling WSSV, and theoretical basis for the study of WSSV receptors.

  15. Facile and green assembly of nanocomposite membranes for fuel cells.

    PubMed

    Quartarone, Eliana; Villa, Davide Carlo; Angioni, Simone; Mustarelli, Piercarlo

    2015-02-04

    We report on a facile spray deposition method, which allows obtaining nanocomposite membranes for high-temperature polymer fuel cells characterized by high homogeneity and excellent proton conductivity. The proposed method is also green, as it requires much smaller amounts of solvents with respect to standard casting.

  16. Sulfonated Nanoplates in Proton Conducting Membranes for Fuel Cells

    SciTech Connect

    Chen, W.F.; Ni’mah, H.; Yu-Cheng Shen, Y.-C.; Kuo, P.-L.

    2011-09-29

    Surface-functionalized nanoplates are synthesized by anchoring sulfonic acid containing siloxanes on zirconium phosphate, and in turn blended with Nafion to fabricate proton conducting membranes. The effects of these sulfonated nanoplates on proton conduction, hydro-characteristics and fuel cell performance are reported.

  17. Hereditary red cell membrane disorders and laboratory diagnostic testing.

    PubMed

    King, M-J; Zanella, A

    2013-06-01

    This overview describes two groups of nonimmune hereditary hemolytic anemias caused by defects in membrane proteins located in distinct layers of the red cell membrane. Hereditary spherocytosis (HS), hereditary elliptocytosis (HE), and hereditary pyropoikilocytosis (HPP) represent disorders of the red cell cytoskeleton. Hereditary stomatocytoses represents disorders of cation permeability in the red cell membrane. The current laboratory screening tests for HS are the osmotic fragility test, acid glycerol lysis time test (AGLT), cryohemolysis test, and eosin-5'-maleimide (EMA)-binding test. For atypical HS, SDS-polyacrylamide gel electrophoresis of erythrocyte membrane proteins is carried out to confirm the diagnosis. The diagnosis of HE/HPP is based on abnormal red cell morphology and the detection of protein 4.1R deficiency or spectrin variants using gel electrophoresis. None of screening tests can detect all HS cases. Some testing centers (a survey of 25 laboratories) use a combination of tests (e.g., AGLT and EMA). No specific screening test for hereditary stomatocytoses is available. The preliminary diagnosis is based on presenting a compensated hemolytic anemia, macrocytosis, and a temperature or time dependent pseudohyperkalemia in some patients. Both the EMA-binding test and the osmotic fragility test may help in differential diagnosis of HS and hereditary stomatocytosis.

  18. Ankyrin-B directs membrane tethering of periaxin and is required for maintenance of lens fiber cell hexagonal shape and mechanics.

    PubMed

    Maddala, Rupalatha; Walters, Mark; Brophy, Peter J; Bennett, Vann; Rao, Ponugoti V

    2016-01-15

    Periaxin (Prx), a PDZ domain protein expressed preferentially in myelinating Schwann cells and lens fibers, plays a key role in membrane scaffolding and cytoarchitecture. Little is known, however, about how Prx is anchored to the plasma membrane. Here we report that ankyrin-B (AnkB), a well-characterized adaptor protein involved in linking the spectrin-actin cytoskeleton to integral membrane proteins, is required for membrane association of Prx in lens fibers and colocalizes with Prx in hexagonal fiber cells. Under AnkB haploinsufficiency, Prx accumulates in the soluble fraction with a concomitant loss from the membrane-enriched fraction of mouse lenses. Moreover, AnkB haploinsufficiency induced age-dependent disruptions in fiber cell hexagonal geometry and radial alignment and decreased compressive stiffness in mouse lenses parallel to the changes observed in Prx null mouse lens. Both AnkB- and Prx-deficient mice exhibit disruptions in membrane organization of the spectrin-actin network and the dystrophin-glycoprotein complex in lens fiber cells. Taken together, these observations reveal that AnkB is required for Prx membrane anchoring and for maintenance of lens fiber cell hexagonal geometry, membrane skeleton organization, and biomechanics.

  19. Ankyrin-B directs membrane tethering of periaxin and is required for maintenance of lens fiber cell hexagonal shape and mechanics

    PubMed Central

    Maddala, Rupalatha; Walters, Mark; Brophy, Peter J.; Bennett, Vann

    2015-01-01

    Periaxin (Prx), a PDZ domain protein expressed preferentially in myelinating Schwann cells and lens fibers, plays a key role in membrane scaffolding and cytoarchitecture. Little is known, however, about how Prx is anchored to the plasma membrane. Here we report that ankyrin-B (AnkB), a well-characterized adaptor protein involved in linking the spectrin-actin cytoskeleton to integral membrane proteins, is required for membrane association of Prx in lens fibers and colocalizes with Prx in hexagonal fiber cells. Under AnkB haploinsufficiency, Prx accumulates in the soluble fraction with a concomitant loss from the membrane-enriched fraction of mouse lenses. Moreover, AnkB haploinsufficiency induced age-dependent disruptions in fiber cell hexagonal geometry and radial alignment and decreased compressive stiffness in mouse lenses parallel to the changes observed in Prx null mouse lens. Both AnkB- and Prx-deficient mice exhibit disruptions in membrane organization of the spectrin-actin network and the dystrophin-glycoprotein complex in lens fiber cells. Taken together, these observations reveal that AnkB is required for Prx membrane anchoring and for maintenance of lens fiber cell hexagonal geometry, membrane skeleton organization, and biomechanics. PMID:26538089

  20. Monocyte cell membrane-derived nanoghosts for targeted cancer therapy

    NASA Astrophysics Data System (ADS)

    Krishnamurthy, S.; Gnanasammandhan, M. K.; Xie, C.; Huang, K.; Cui, M. Y.; Chan, J. M.

    2016-03-01

    Core-shell type `nanoghosts' were synthesized with a drug-loaded biodegradable PLGA core and a monocyte cell membrane-derived shell. The nanoghosts were monodisperse with an average size <200 nm, and showed good serum stability for 120 h. Doxorubicin-loaded nanoghosts showed greater cellular uptake and cytotoxicity compared to non-coated nanoparticle controls in metastatic MCF-7 breast cancer cell lines.Core-shell type `nanoghosts' were synthesized with a drug-loaded biodegradable PLGA core and a monocyte cell membrane-derived shell. The nanoghosts were monodisperse with an average size <200 nm, and showed good serum stability for 120 h. Doxorubicin-loaded nanoghosts showed greater cellular uptake and cytotoxicity compared to non-coated nanoparticle controls in metastatic MCF-7 breast cancer cell lines. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr07588b

  1. Mitochondria and cell death: outer membrane permeabilization and beyond.

    PubMed

    Tait, Stephen W G; Green, Douglas R

    2010-09-01

    Mitochondrial outer membrane permeabilization (MOMP) is often required for activation of the caspase proteases that cause apoptotic cell death. Various intermembrane space (IMS) proteins, such as cytochrome c, promote caspase activation following their mitochondrial release. As a consequence, mitochondrial outer membrane integrity is highly controlled, primarily through interactions between pro- and anti-apoptotic members of the B cell lymphoma 2 (BCL-2) protein family. Following MOMP by pro-apoptotic BCL-2-associated X protein (BAX) or BCL-2 antagonist or killer (BAK), additional regulatory mechanisms govern the mitochondrial release of IMS proteins and caspase activity. MOMP typically leads to cell death irrespective of caspase activity by causing a progressive decline in mitochondrial function, although cells can survive this under certain circumstances, which may have pathophysiological consequences.

  2. Investigation of the bystander effect in MRC5 cells after acute and fractionated irradiation in vitro.

    PubMed

    Soleymanifard, Shokouhozaman; Toossi, Mohammad Taghi Bahreyni; Samani, Roghayeh Kamran; Mohebbi, Shokoufeh

    2014-04-01

    Radiation-induced bystander effect (RIBE) has been defined as radiation responses observed in nonirradiated cells. It has been the focus of investigators worldwide due to the deleterious effects it induces in nonirradiated cells. The present study was performed to investigate whether acute or fractionated irradiation will evoke a differential bystander response in MRC5 cells. A normal human cell line (MRC5), and a human lung tumor cell line (QU-DB) were exposed to 0, 1, 2, and 4Gy of single acute or fractionated irradiation of equal fractions with a gap of 6 h. The MRC5 cells were supplemented with the media of irradiated cells and their micronucleus frequency was determined. The micronucleus frequency after single and fractionated irradiation did not vary significantly in the MRC5 cells conditioned with autologous or QU-DB cell-irradiated media, except for 4Gy where the frequency of micronucleated cells was lower in those MRC5 cells cultured in the media of QU-DB-exposed with a single dose of 4Gy. Our study demonstrates that the radiation-induced bystander effect was almost similar after single acute and fractionated exposure in MRC5 cells.

  3. How to Evaluate the Electric Noise in a Cell Membrane?

    NASA Astrophysics Data System (ADS)

    Bier, M.

    2006-05-01

    There has been considerable public anxiety about possible health effects of electromagnetic radiation emitted by high voltage power lines. Power frequencies (60 Hz in the US, 50 Hz in many other countries) are sufficiently slow for the associated electric fields to distribute themselves across the highly resistive cell membranes. To assess the ambient power frequency fields, researchers have compared the voltage that these fields induce across cell membranes to the strength of the electric noise that the membranes generate themselves through Brownian motion. However, there has been disagreement among researchers on how to evaluate this equilibrium membrane electric noise. I will review the different approaches and present an {ITALIC ab initio} modeling of membrane electric fields. I will show that different manifestations of Brownian noise lead to an electric noise intensity that is many times larger than what conventional estimates have yielded. Next, the legitimacy of gauging a nonequilibrium external signal against internal equilibrium noise is questioned and a more meaningful criterion is proposed. Finally, an estimate will be derived of the nonequilibrium noise intensity due to the driven ion traffic through randomly opening and closing ion channels.

  4. A prototype biosensor: artificial cell membrane on porous silicon

    NASA Astrophysics Data System (ADS)

    Retamal, Maria Jose; Cisternas, Marcelo; Busch, Mark; Gutierrez, Sebastian; Huber, Patrick; Perez-Acle, Tomas; Kappl, Michael; Volkmann, Ulrich

    2014-03-01

    Biosensors have been studied in recent years because they are powerful instruments to detect physical or chemical parameters as, e.g., intracellular interactions. What we propose is a prototype biosensor based on an artificial cell membrane (DPPC) on porous silicon. Porous silicon is used as a sponge-like substrate to absorb water by capillarity and keep the membrane hydrated, which is essential for the membrane not to denature when performing temperature cycles. Thus, one can observe the phase changes of the cell membrane with temperature using optical and surface scanning methods. In this research we used the technique of Very High Resolution Ellipsometry (VHRE) to observe changes in the ellipsometric angles during temperature ramps, which are attributed to different lipid phase transitions. Imaging ellipsometry (IE) was used to observe surface changes at the microscopic level and Atomic Force Microscopy (AFM) to observe changes in the topography of the membrane at the nanoscale. This work was supported by Fondecyt 1100882, DAAD-Conicyt PCCI 044, Conicyt Scholarship and Project Anillo ACT 1107.

  5. Fractionation and Structural Characterization of Arabinogalactan-Proteins from the Cell Wall of Rose Cells.

    PubMed Central

    Serpe, M. D.; Nothnagel, E. A.

    1995-01-01

    Arabinogalactan-proteins (AGPs) have been purified from Paul's Scarlet rose (Rosa sp.) cell walls. As estimated by gel permeation chromatography, the apparent molecular masses of the two major cell-wall AGP fractions were 130 and 242 kD. Since the 130-kD AGP had a ratio of arabinose/glucuronic acid that was 12 times higher than that of the 242-kD AGP, the fractions were named cell-wall AGP1 (CW-AGP1) and glucuronogalactan-protein (GGP), respectively. CW-AGP1 and GGP contained predominantly t-arabinofuranosyl residues; 3-linked, 6-linked, and 3,6-branched galactopyranosyl residues; and 4-linked and t-glucuronopyranosyl residues. The 1H-nuclear magnetic resonance spectra of CW-AGP1 and GGP showed that the arabinofuranosyl and galactopyranosyl residues were predominantly in [alpha]- and [beta]-anomeric configuration, respectively, and that GGP contained a few O-acetyl residues. The protein moieties of CW-AGP1 and GGP were both rich in hydroxyproline and alanine but differed in the percentage of various amino acids, including hydroxyproline, alanine, serine, and glycine. Cell-wall AGPs bound to ([beta]-D-glucosyl)3 Yariv phenylglycoside, but the stoichiometry of binding was about 6 times greater in GGP than in other Rosa AGPs. GGP seems to be peculiar to the cell wall, since no similar molecule was found in the culture medium. PMID:12228648

  6. Analysis of MicroRNA-mRNA Interactions in Stem-cell Enriched Fraction of Oral Squamous Cell Carcinoma.

    PubMed

    Richard, Vinitha; Raju, Rajesh; Paul, Aswathy Mary; Girijadevi, Reshmi; Santhoshkumar, Thankayyan Retnabai; Pillai, Madhavan Radhakrishna

    2017-03-09

    This study is an integrated analysis of the transcriptome profile, MicroRNA (miRNA) and their experimentally validatedmRNA targets differentially expressed in the tumorigenic stem-like fraction of oral squamous cell carcinoma (OSCC). We had previously reported the co-existence of multiple drug resistant, tumorigenic fractions termed as side population (SP1, SP2 and MP2) and a non-tumorigenic fraction, main population (MP1) in oral cancer. These fractions displayed self-renewal, regeneration potential and expressed known stemness related cell surface markers despite functional differences. Flow cytometrically sorted pure fractions of SP1 and MP1 cells were subjected to differential expression analysis of both mRNAs and microRNAs. A significant upregulation of genes associated with inflammation, cell survival, cell proliferation, drug transporters and antiapoptotic pathways in addition to enhanced transcriptome reprogramming mediated by DNA- histone binding proteins and pattern recognition receptor-mediated signaling was found to play a crucial role in the transformation of non-tumorigenic MP1 fraction to tumorigenic SP1 fraction. We also identified several differentially expressed microRNAs that specifically target genes distinctive of tumorigenic SP1 fraction. MicroRNA mediated downregulation of stemness associated markers CD44, CD147 and upregulation of CD151 may also account for the emergence and persistence of multiple tumorigenic stem cell fractions with varying degrees of malignancy. The phenotypic switch of cancer cells to stem-like OSCC cells mediated by transcriptomal regulation is effectual in addressing biological tumor heterogeneity and subsequent therapeutic resistance leading to minimal residual disease (MRD) condition in oral cancer. Detailed study of the interplay of microRNAs, mRNA and the cellular phases involved in the gradual transition of non-tumorigenic cancer cells to tumorigenic stem-like cells in solid tumors would enable detection and

  7. Effect of BCD Plasma on a Bacteria Cell Membrane

    NASA Astrophysics Data System (ADS)

    Nasrin, Navabsafa; Hamid, Ghomi; Maryam, Nikkhah; Soheila, Mohades; Hossein, Dabiri; Saeed, Ghasemi

    2013-07-01

    Abstract Cell membrane rupture is considered to be one of the probable mechanisms for bacterial inactivation using barrier corona discharge (BCD) plasma. In this paper, the effect of the BCD plasma on the Escherichia coli (E. coli) bacteria cell wall was investigated through two analytical methods; Adenosine-5'-triphosphate (ATP) assay and Atomic Force Microscopy (AFM). The ATP assay results indicate an increase in the ATP content of samples which were exposed to the BCD plasma. This implies the bacteria cell rupture. Moreover, AFM images confirm a serious damage of the bacteria cell wall under the influence of the bactericidal agents of the plasma.

  8. The importance of extracellular speciation and corrosion of copper nanoparticles on lung cell membrane integrity.

    PubMed

    Hedberg, Jonas; Karlsson, Hanna L; Hedberg, Yolanda; Blomberg, Eva; Odnevall Wallinder, Inger

    2016-05-01

    Copper nanoparticles (Cu NPs) are increasingly used in various biologically relevant applications and products, e.g., due to their antimicrobial and catalytic properties. This inevitably demands for an improved understanding on their interactions and potential toxic effects on humans. The aim of this study was to investigate the corrosion of copper nanoparticles in various biological media and to elucidate the speciation of released copper in solution. Furthermore, reactive oxygen species (ROS) generation and lung cell (A549 type II) membrane damage induced by Cu NPs in the various media were studied. The used biological media of different complexity are of relevance for nanotoxicological studies: Dulbecco's modified eagle medium (DMEM), DMEM(+) (includes fetal bovine serum), phosphate buffered saline (PBS), and PBS+histidine. The results show that both copper release and corrosion are enhanced in DMEM(+), DMEM, and PBS+histidine compared with PBS alone. Speciation results show that essentially no free copper ions are present in the released fraction of Cu NPs in neither DMEM(+), DMEM nor histidine, while labile Cu complexes form in PBS. The Cu NPs were substantially more membrane reactive in PBS compared to the other media and the NPs caused larger effects compared to the same mass of Cu ions. Similarly, the Cu NPs caused much more ROS generation compared to the released fraction only. Taken together, the results suggest that membrane damage and ROS formation are stronger induced by Cu NPs and by free or labile Cu ions/complexes compared with Cu bound to biomolecules.

  9. Alternative Sources of Adult Stem Cells: Human Amniotic Membrane

    NASA Astrophysics Data System (ADS)

    Wolbank, Susanne; van Griensven, Martijn; Grillari-Voglauer, Regina; Peterbauer-Scherb, Anja

    Human amniotic membrane is a highly promising cell source for tissue engineering. The cells thereof, human amniotic epithelial cells (hAEC) and human amniotic mesenchymal stromal cells (hAMSC), may be immunoprivileged, they represent an early developmental status, and their application is ethically uncontroversial. Cell banking strategies may use freshly isolated cells or involve in vitro expansion to increase cell numbers. Therefore, we have thoroughly characterized the effect of in vitro cultivation on both phenotype and differentiation potential of hAEC. Moreover, we present different strategies to improve expansion including replacement of animal-derived supplements by human platelet products or the introduction of the catalytic subunit of human telomerase to extend the in vitro lifespan of amniotic cells. Characterization of the resulting cultures includes phenotype, growth characteristics, and differentiation potential, as well as immunogenic and immunomodulatory properties.

  10. Enrichment of putative stem cells from adipose tissue using dielectrophoretic field-flow fractionation

    PubMed Central

    Vykoukal, Jody; Vykoukal, Daynene M.; Freyberg, Susanne; Alt, Eckhard U.; Gascoyne, Peter R. C.

    2009-01-01

    We have applied the microfluidic cell separation method of dielectrophoretic field-flow fractionation (DEP-FFF) to the enrichment of a putative stem cell population from an enzyme-digested adipose tissue derived cell suspension. A DEP-FFF separator device was constructed using a novel microfluidic-microelectronic hybrid flex-circuit fabrication approach that is scaleable and anticipates future low-cost volume manufacturing. We report the separation of a nucleated cell fraction from cell debris and the bulk of the erythrocyte population, with the relatively rare (<2% starting concentration) NG2-positive cell population (pericytes and/or putative progenitor cells) being enriched up to 14-fold. This work demonstrates a potential clinical application for DEP-FFF and further establishes the utility of the method for achieving label-free fractionation of cell subpopulations. PMID:18651083

  11. The fractionation of isolated liver cells from normal and carcinogen treated rats.

    PubMed Central

    Horsfall, A. C.; Ketterer, B.

    1976-01-01

    Suspensions of isolated cells were obtained from livers of normal rats and rats treated with the hepatocarcinogen N,N-dimethyl-4-aminoazobenzene. Differential centrifugation of dispersed cells yielded a large parenchymal cell fraction and a small non-parencymal cell fraction. By means of rate sedimentation through different concnetrations of Ficoll, parenchymal cells were separated into cells with fast, intermediate and slow rates of sedimentation. Periods of sedimentation were brief and centrifugal forces low in order to retain the best possible state of preservation of cells. DNA, RNA and protein contents, acid phosphatase activity, cell size and nucleocytoplasmic ratios of parenchymal cells sedimenting at fast, intermediate and slow rates were measured. Cell fractions from normal livers had properties suggesting that faster sedimenting cells were derived from the centre and middle of the lobule whereas slowly sedimenting cells were periportal; however, much of the periportal cell population remained in a residue of undissociated tissue. Compared with normal cells, carcinogen treated cells appeared to fractionate according to different physical and chemical criteria and could not be related to their origin within the liver lobule. They were smaller, slower sedimenting, lower in protein and RNA content and acid phosphatase activity. The tissue residue contained abnromal histological structures. PMID:814912

  12. Reduction of DOM fractions and their trihalomethane formation potential in surface river water by in-line coagulation with ceramic membrane filtration.

    PubMed

    Rakruam, Pharkphum; Wattanachira, Suraphong

    2014-03-01

    This research was aimed at investigating the reduction of DOM fractions and their trihalomethane formation potential (THMFP) by in-line coagulation with 0.1 μm ceramic membrane filtration. The combination of ceramic membrane filtration with a coagulation process is an alternative technology which can be applied to enhance conventional coagulation processes in the field of water treatment and drinking water production. The Ping River water (high turbidity water) was selected as the raw surface water because it is currently the main raw water source for water supply production in the urban and rural areas of Chiang Mai Province. From the investigation, the results showed that the highest percent reductions of DOC, UV-254, and THMFP (47.6%, 71.0%, and 67.4%, respectively) were achieved from in-line coagulation with ceramic membrane filtration at polyaluminum chloride dosage 40 mg/L. Resin adsorption techniques were employed to characterize the DOM in raw surface water and filtered water. The results showed that the use of a ceramic membrane with in-line coagulation was able to most efficiently reduce the hydrophobic fraction (HPOA) (68.5%), which was then followed by the hydrophilic fraction (HPIA) (49.3%). The greater mass DOC reduction of these two fractions provided the highest THMFP reductions (55.1% and 37.2%, respectively). Furthermore, the in-line coagulation with ceramic membrane filtration was able to reduce the hydrophobic (HPOB) fraction which is characterized by high reactivity toward THM formation. The percent reduction of mass DOC and THMFP of HPOB by in-line coagulation with ceramic membrane filtration was 45.9% and 48.0%, respectively.

  13. Chemical Imaging of the Cell Membrane by NanoSIMS

    SciTech Connect

    Weber, P K; Kraft, M L; Frisz, J F; Carpenter, K J; Hutcheon, I D

    2010-02-23

    The existence of lipid microdomains and their role in cell membrane organization are currently topics of great interest and controversy. The cell membrane is composed of a lipid bilayer with embedded proteins that can flow along the two-dimensional surface defined by the membrane. Microdomains, known as lipid rafts, are believed to play a central role in organizing this fluid system, enabling the cell membrane to carry out essential cellular processes, including protein recruitment and signal transduction. Lipid rafts are also implicated in cell invasion by pathogens, as in the case of the HIV. Therefore, understanding the role of lipid rafts in cell membrane organization not only has broad scientific implications, but also has practical implications for medical therapies. One of the major limitations on lipid organization research has been the inability to directly analyze lipid composition without introducing artifacts and at the relevant length-scales of tens to hundreds of nanometers. Fluorescence microscopy is widely used due to its sensitivity and specificity to the labeled species, but only the labeled components can be observed, fluorophores can alter the behavior of the lipids they label, and the length scales relevant to imaging cell membrane domains are between that probed by fluorescence resonance energy transfer (FRET) imaging (<10 nm) and the diffraction limit of light. Topographical features can be imaged on this length scale by atomic force microscopy (AFM), but the chemical composition of the observed structures cannot be determined. Immuno-labeling can be used to study the distribution of membrane proteins at high resolution, but not lipid composition. We are using imaging mass spectrometry by secondary ion mass spectrometry (SIMS) in concert with other high resolution imaging methods to overcome these limitations. The experimental approach of this project is to combine molecule-specific stable isotope labeling with high-resolution SIMS using a

  14. Membrane protein synthesis in cell-free systems: from bio-mimetic systems to bio-membranes.

    PubMed

    Sachse, Rita; Dondapati, Srujan K; Fenz, Susanne F; Schmidt, Thomas; Kubick, Stefan

    2014-08-25

    When taking up the gauntlet of studying membrane protein functionality, scientists are provided with a plethora of advantages, which can be exploited for the synthesis of these difficult-to-express proteins by utilizing cell-free protein synthesis systems. Due to their hydrophobicity, membrane proteins have exceptional demands regarding their environment to ensure correct functionality. Thus, the challenge is to find the appropriate hydrophobic support that facilitates proper membrane protein folding. So far, various modes of membrane protein synthesis have been presented. Here, we summarize current state-of-the-art methodologies of membrane protein synthesis in biomimetic-supported systems. The correct folding and functionality of membrane proteins depend in many cases on their integration into a lipid bilayer and subsequent posttranslational modification. We highlight cell-free systems utilizing the advantages of biological membranes.

  15. Block copolymers for alkaline fuel cell membrane materials

    NASA Astrophysics Data System (ADS)

    Li, Yifan

    Alkaline fuel cells (AFCs) using anion exchange membranes (AEMs) as electrolyte have recently received considerable attention. AFCs offer some advantages over proton exchange membrane fuel cells, including the potential of non-noble metal (e.g. nickel, silver) catalyst on the cathode, which can dramatically lower the fuel cell cost. The main drawback of traditional AFCs is the use of liquid electrolyte (e.g. aqueous potassium hydroxide), which can result in the formation of carbonate precipitates by reaction with carbon dioxide. AEMs with tethered cations can overcome the precipitates formed in traditional AFCs. Our current research focuses on developing different polymer systems (blend, block, grafted, and crosslinked polymers) in order to understand alkaline fuel cell membrane in many aspects and design optimized anion exchange membranes with better alkaline stability, mechanical integrity and ionic conductivity. A number of distinct materials have been produced and characterized. A polymer blend system comprised of poly(vinylbenzyl chloride)-b-polystyrene (PVBC-b-PS) diblock copolymer, prepared by nitroxide mediated polymerization (NMP), with poly(2,6-dimethyl-1,4-phenylene oxide) (PPO) or brominated PPO was studied for conversion into a blend membrane for AEM. The formation of a miscible blend matrix improved mechanical properties while maintaining high ionic conductivity through formation of phase separated ionic domains. Using anionic polymerization, a polyethylene based block copolymer was designed where the polyethylene-based block copolymer formed bicontinuous morphological structures to enhance the hydroxide conductivity (up to 94 mS/cm at 80 °C) while excellent mechanical properties (strain up to 205%) of the polyethylene block copolymer membrane was observed. A polymer system was designed and characterized with monomethoxy polyethylene glycol (mPEG) as a hydrophilic polymer grafted through substitution of pendent benzyl chloride groups of a PVBC

  16. ``Lock and key mechanism'' for ligand binding with adrenergic receptors and the arising mechanical effects on the cell membrane

    NASA Astrophysics Data System (ADS)

    Lunghi, Laura; Deseri, Luca

    2013-03-01

    Chemicals hitting the surface of cell aggregates are known to give arise to cyclic Adenosine Mono Phosphate (cAMP), a second messenger that transduces inside the cell the effects of species that cannot get through the cell membrane. Ligands bind to a specific receptor following the so called ``lock and key mechanism'' (beta)-adrenergic receptors are proteins embedded in the lipid bilayer characterized by seven transmembrane helices. Thinning and thickening in cell membranes may be initiated by conformational changes of some of three of the seven domains above. The cell response is linked to the coupling of chemical, conformational and mechanical effects. Part of the cAMP remains intracellular, whereas the remaining fractions migrates outside the cell due to membrane transporters. A new Helmholtz free energy, accounting for receptor and transporter densities, receptor conformation field and membrane elasticity is investigated. It is shown how the density of active receptors is directly related to the conformation field and it enters the resulting balance equation for the membrane stress. Balance laws for fluxes of transporters and receptors, coupled with the former because of the outgoing cAMP flux caused by the transporters, as well as for the diffusive powers must be supplied. The Center for Nonlinear Analysis through the NSF Grant No. DMS-0635983 is gratefully acknowledged.

  17. Reticulated lipid probe fluorescence reveals MDCK cell apical membrane topography.

    PubMed

    Colarusso, Pina; Spring, Kenneth R

    2002-02-01

    High spatial resolution confocal microscopy of young MDCK cells stained with the lipophilic probe 1,1'-dihexadecyl-3,3,3',3'- tetramethylindocarbocyanine perchlorate (DiIC(16)) revealed a reticulated fluorescence pattern on the apical membrane. DiIC(16) was delivered as crystals to live cells to minimize possible solvent perturbations of the membrane lipids. The ratio of the integrated fluorescence intensities in the bright versus dim regions was 1.6 +/- 0.1 (n = 13). Deconvolved images of the cells were consistent with exclusive plasma membrane staining. Multi-spectral and fluorescence anisotropy microscopy did not reveal differences between bright and dim regions. Bright regions coincided with microvilli and microridges observed by differential interference contrast microscopy and were stable for several minutes. Fluorescence recovery after photobleaching yielded similar diffusion coefficients (pooled D = 1.5 +/- 0.6 x 10(-9) cm(2)/s, n = 40) for both bright and dim regions. Line fluorescence recovery after photobleaching showed that the reticulated pattern was maintained as the fluorescence recovered in the bleached areas. Cytochalasin D did not affect the staining pattern, but the pattern was eliminated by cholesterol depletion with methyl-beta-cyclodextrin. We conclude that the reticulated fluorescence pattern was caused by increased optical path lengths through the microvilli and microridges compared with the flat areas on the apical membrane.

  18. Reticulated lipid probe fluorescence reveals MDCK cell apical membrane topography.

    PubMed Central

    Colarusso, Pina; Spring, Kenneth R

    2002-01-01

    High spatial resolution confocal microscopy of young MDCK cells stained with the lipophilic probe 1,1'-dihexadecyl-3,3,3',3'- tetramethylindocarbocyanine perchlorate (DiIC(16)) revealed a reticulated fluorescence pattern on the apical membrane. DiIC(16) was delivered as crystals to live cells to minimize possible solvent perturbations of the membrane lipids. The ratio of the integrated fluorescence intensities in the bright versus dim regions was 1.6 +/- 0.1 (n = 13). Deconvolved images of the cells were consistent with exclusive plasma membrane staining. Multi-spectral and fluorescence anisotropy microscopy did not reveal differences between bright and dim regions. Bright regions coincided with microvilli and microridges observed by differential interference contrast microscopy and were stable for several minutes. Fluorescence recovery after photobleaching yielded similar diffusion coefficients (pooled D = 1.5 +/- 0.6 x 10(-9) cm(2)/s, n = 40) for both bright and dim regions. Line fluorescence recovery after photobleaching showed that the reticulated pattern was maintained as the fluorescence recovered in the bleached areas. Cytochalasin D did not affect the staining pattern, but the pattern was eliminated by cholesterol depletion with methyl-beta-cyclodextrin. We conclude that the reticulated fluorescence pattern was caused by increased optical path lengths through the microvilli and microridges compared with the flat areas on the apical membrane. PMID:11806917

  19. Capacitance-Voltage Measurement of Transporting Function at Cell Membrane

    NASA Astrophysics Data System (ADS)

    Sakata, Toshiya; Miyahara, Yuji

    In this paper, we report the detection of transporting function at cell membrane using capacitance-voltage (CV) measurement. The detection principle of our devices is based on the field-effect of electrostatic interaction between charged species at cell membrane in solution and surface electrons in silicon crystal through the gate insulator of Si3N4/SiO2 thin double-layer. We designed an oocyte-based field-effect capacitor, on which a Xenopus laevis oocyte was fixed. The transporter of human organic anion transporting peptide C (hOATP-C) was expressed at oocyte membrane by induction of cRNA. The electrical phenomena such as ion or molecular charge flux at the interface between cell membrane and gate surface could be detected as the change of flat band voltage in CV characteristics. The flat band voltage shift decreased with incubation time after introduction of substrate into the oocyte-based field-effect capacitor. The electrical signal is due to the change of charge flux from the oocyte at the gate surface inspired by transporter-substrate binding. The platform based on the oocyte-based field-effect capacitor is suitable for a simple and non-invasive detection system in order to analyze function of transporters related to drug efficacy.

  20. A Novel Unitized Regenerative Proton Exchange Membrane Fuel Cell

    NASA Technical Reports Server (NTRS)

    Murphy, O. J.; Cisar, A. J.; Gonzalez-Martin, A.; Salinas, C. E.; Simpson, S. F.

    1996-01-01

    A difficulty encountered in designing a unitized regenerative proton exchange membrane (PEM) fuel cell lies in the incompatibility of electrode structures and electrocatalyst materials optimized for either of the two functions (fuel cell or electrolyzer) with the needs of the other function. This difficulty is compounded in previous regenerative fuel cell designs by the fact that water, which is needed for proton conduction in the PEM during both modes of operation, is the reactant supplied to the anode in the electrolyzer mode of operation and the product formed at the cathode in the fuel cell mode. Drawbacks associated with existing regenerative fuel cells have been addressed. In a first innovation, electrodes function either as oxidation electrodes (hydrogen ionization or oxygen evolution) or as reduction electrodes (oxygen reduction or hydrogen evolution) in the fuel cell and electrolyzer modes, respectively. Control of liquid water within the regenerative fuel cell has been brought about by a second innovation. A novel PEM has been developed with internal channels that permit the direct access of water along the length of the membrane. Lateral diffusion of water along the polymer chains of the PEM provides the water needed at electrode/PEM interfaces. Fabrication of the novel single cell unitized regenerative fuel cell and results obtained on testing it are presented.

  1. Anti-angiogenic quassinoid-rich fraction from Eurycoma longifolia modulates endothelial cell function.

    PubMed

    Al-Salahi, Omar Saeed Ali; Kit-Lam, Chan; Majid, Amin Malik Shah Abdul; Al-Suede, Fouad Saleih R; Mohammed Saghir, Sultan Ayesh; Abdullah, Wan Zaidah; Ahamed, Mohamed B Khadeer; Yusoff, Narazah Mohd

    2013-11-01

    Targeting angiogenesis could be an excellent strategy to combat angiogenesis-dependent pathophysiological conditions such as cancer, rheumatoid arthritis, obesity, systemic lupus erythematosus, psoriasis, proliferative retinopathy and atherosclerosis. Recently a number of clinical investigations are being undertaken to assess the potential therapeutic application of various anti-angiogenic agents. Many of these angiogenesis inhibitors are directed against the functions of endothelial cells, which are considered as the building blocks of blood vessels. Similarly, roots of a traditional medicinal plant, Eurycoma longifolia, can be used as an alternative treatment to prevent and treat the angiogenesis-related diseases. In the present study, antiangiogenic potential of partially purified quassinoid-rich fraction (TAF273) of E. longifolia root extract was evaluated using ex vivo and in vivo angiogenesis models and the anti-angiogenic efficacy of TAF273 was investigated in human umbilical vein endothelial cells (HUVEC). TAF273 caused significant suppression in sprouting of microvessels in rat aorta with IC50 11.5μg/ml. TAF273 (50μg/ml) showed remarkable inhibition (63.13%) of neovascularization in chorioallantoic membrane of chick embryo. Tumor histology also revealed marked reduction in extent of vascularization. In vitro, TAF273 significantly inhibited the major angiogenesis steps such as proliferation, migration and differentiation of HUVECs. Phytochemical analysis revealed high content of quassinoids in TAF273. Specially, HPLC characterization showed that TAF273 is enriched with eurycomanone, 13α(21)-epoxyeurycomanone and eurycomanol. These results demonstrated that the antiangiogenic activity of TAF273 may be due to its inhibitory effect on endothelial cell proliferation, differentiation and migration which could be attributed to the high content of quassinoids in E. longifolia.

  2. Incorporation profiles of conjugated linoleic acid isomers in cell membranes and their positional distribution in phospholipids

    PubMed Central

    Subbaiah, Papasani V.; Gould, Ian G.; Lal, Samanta; Aizezi, Buzulagu

    2010-01-01

    Although the conjugated linoleic acids (CLA) have several isomer-specific biological effects including anti-carcinogenic and anti-adipogenic effects, their mechanisms of action remain unclear. To determine their potential effects on membrane structure and function, we studied the incorporation profiles of four CLA isomers (trans-10 cis-12 (A), trans-9 trans-11 (B), cis-9 trans-11 (C), and cis-9 cis-11 (D)) in CHO and HepG2 cells. All four isomers were incorporated into cellular lipids as efficiently as linoleic acid (LA), with the majority of the incorporated CLA present in membrane rafts. Of the four isomers, only CLA-A increased the cholesterol content of the raft fraction. Over 50% of the incorporated CLAs were recovered in phosphatidylcholine of CHO cells, but in HepG2 the neutral lipids contained the majority of CLA. The desaturation index (18:1/18:0 and 16:1/16:0) was reduced by CLA-A, but increased by CLA-B, the effects being apparent mostly in raft lipids. The Δ9 desaturase activity was inhibited by CLAs A and C. Unlike LA, which was mostly found in the sn-2 position of phospholipids, most CLAs were also incorporated significantly into the sn-1 position in both cell types. These studies show that the incorporation profiles of CLA isomers differ significantly from that of LA, and this could lead to alterations in membrane function, especially in the raft-associated proteins. PMID:20920595

  3. Mass Spectrometry of Polymer Electrolyte Membrane Fuel Cells

    PubMed Central

    Ostroverkh, Anna; Fiala, Roman; Rednyk, Andrii; Matolín, Vladimír

    2016-01-01

    The chemical analysis of processes inside fuel cells under operating conditions in either direct or inverted (electrolysis) mode and their correlation with potentiostatic measurements is a crucial part of understanding fuel cell electrochemistry. We present a relatively simple yet powerful experimental setup for online monitoring of the fuel cell exhaust (of either cathode or anode side) downstream by mass spectrometry. The influence of a variety of parameters (composition of the catalyst, fuel type or its concentration, cell temperature, level of humidification, mass flow rate, power load, cell potential, etc.) on the fuel cell operation can be easily investigated separately or in a combined fashion. We demonstrate the application of this technique on a few examples of low-temperature (70°C herein) polymer electrolyte membrane fuel cells (both alcohol- and hydrogen-fed) subjected to a wide range of conditions. PMID:28042492

  4. Mass Spectrometry of Polymer Electrolyte Membrane Fuel Cells.

    PubMed

    Johánek, Viktor; Ostroverkh, Anna; Fiala, Roman; Rednyk, Andrii; Matolín, Vladimír

    2016-01-01

    The chemical analysis of processes inside fuel cells under operating conditions in either direct or inverted (electrolysis) mode and their correlation with potentiostatic measurements is a crucial part of understanding fuel cell electrochemistry. We present a relatively simple yet powerful experimental setup for online monitoring of the fuel cell exhaust (of either cathode or anode side) downstream by mass spectrometry. The influence of a variety of parameters (composition of the catalyst, fuel type or its concentration, cell temperature, level of humidification, mass flow rate, power load, cell potential, etc.) on the fuel cell operation can be easily investigated separately or in a combined fashion. We demonstrate the application of this technique on a few examples of low-temperature (70°C herein) polymer electrolyte membrane fuel cells (both alcohol- and hydrogen-fed) subjected to a wide range of conditions.

  5. Morphological features (defects) in fuel cell membrane electrode assemblies

    NASA Astrophysics Data System (ADS)

    Kundu, S.; Fowler, M. W.; Simon, L. C.; Grot, S.

    Reliability and durability issues in fuel cells are becoming more important as the technology and the industry matures. Although research in this area has increased, systematic failure analysis, such as a failure modes and effects analysis (FMEA), are very limited in the literature. This paper presents a categorization scheme of causes, modes, and effects related to fuel cell degradation and failure, with particular focus on the role of component quality, that can be used in FMEAs for polymer electrolyte membrane (PEM) fuel cells. The work also identifies component defects imparted on catalyst-coated membranes (CCM) by manufacturing and proposes mechanisms by which they can influence overall degradation and reliability. Six major defects have been identified on fresh CCM materials, i.e., cracks, orientation, delamination, electrolyte clusters, platinum clusters, and thickness variations.

  6. Gold Nanoparticles-Enhanced Proton Exchange Membrane (PEM) Fuel Cell

    NASA Astrophysics Data System (ADS)

    Li, Hongfei; Pan, Cheng; Liu, Ping; Zhu, Yimei; Adzic, Radoslav; Rafailovich, Miriam

    Proton exchange membrane fuel cells have drawn great attention and been taken as a promising alternated energy source. One of the reasons hamper the wider application of PEM fuel cell is the catalytic poison effect from the impurity of the gas flow. Haruta has predicted that gold nanoparticles that are platelet shaped and have direct contact with the metal oxide substrate to be the perfect catalysts of the CO oxidization, yet the synthesis method is difficult to apply in the Fuel Cell. In our approach, thiol-functionalized gold nanoparticles were synthesized through two-phase method developed by Brust et al. We deposit these Au particles with stepped surface directly onto the Nafion membrane in the PEM fuel cell by Langmuir-Blodgett method, resulting in over 50% enhancement of the efficiency of the fuel cell. DFT calculations were conducted to understand the theory of this kind of enhancement. The results indicated that only when the particles were in direct surface contact with the membrane, where AuNPs attached at the end of the Nafion side chains, it could reduce the energy barrier for the CO oxidation that could happen at T<300K.

  7. The properties and extracellular location of 5'-nucleotidase of the rat fat-cell plasma membrane.

    PubMed Central

    Newby, A C; Luzio, J P; Hales, C N

    1975-01-01

    1. A phosphohydrolase specific for 5'-nucleotides was characterized by using a particulate fraction from isolated fat-cells. 2. The activity of intact cells towards 5'-AMP was studied. 3. The activity in either situation had the same KM for AMP (45 muM) and was inhibited by low concentrations of ATP (less than 50 muM), but less potently by the ATP analogues AMP-P(CH2)P(adenylyl (beta gamma-methylene)diphosphonate) and AMP-P)NH)P (adenylylimidodiphosphate). 4. Homogenization of intact fat-cells caused no increase in activity and at least 85% of the activity was recovered in the particulate preparation. 5. The preparation of fat-cells used in this work was not freely permeable to AMP. 6. The ability of intact fat-cells to hydrolyse AMP implies that 5'-nucleotidase is an ectoenzyme in fat-cells. 7. Concentrations of ATP 100 times lower than intracellular concentrations inhibit the enzyme when added extracellularly to intact fat-cells, implying that this effect is also medicated at the extracellular face of the membrane. 8. Antibodies raised to whole liver cells and whole fat-cells inhibit 5'-nucleotidase in intact cells. 9. Incubation of intact fat-cells with adrenaline (1 mug/ml) or insulin (50 mui.u./ml) failed to alter the KM or Vmax. of the enzyme. PMID:167725

  8. Monitoring the fractionation of a whey protein isolate during dead-end membrane filtration using fluorescence and chemometric methods.

    PubMed

    Elshereef, Rand; Budman, Hector; Moresoli, Christine; Legge, Raymond L

    2010-01-01

    During membrane-based separation of proteins, changes in protein concentration of the permeate and retentate streams occurs over time. The current work proposes a new approach for monitoring the changes in concentrations of proteins in both permeate and retentate by making use of data collected using fluorescence spectroscopy and intrinsic protein fluorescence analyzed by multivariate statistical techniques. Whey protein isolate consists mainly of alpha-lactalbumin (alpha-LA), beta-lactoglobulin (beta-LG), and small proportion of bovine serum albumin (BSA) and was used as a model system in this study. A fiber optic probe (FOP) was used to acquire multiwavelength fluorescence spectra for permeate and retentate streams at different times during UF-based separation of the components from a multicomponent solution. Multivariate regression models were developed for predicting the concentrations of alpha-LA, beta-LG, and BSA by establishing a calibration model between data acquired using the FOP and the corresponding protein concentration levels measured by size-exclusion chromatography. The model was validated using FOP data that were not previously used for calibration of the regression models. This comparison showed that concentrations of alpha-LA, beta-LG, and BSA could be predicted directly from FOP data within reasonable accuracy by making use of multivariate calibration tools. This approach has several attractive features including that it is nondestructive, fast, and relatively simple to perform. This technique has potential practical applications as it could offer the opportunity for in situ monitoring of membrane filtration processes by tracking individual protein transmission and selectivity of fractionation.

  9. Proteomic analysis of the sarcosine-insoluble outer membrane fraction of Pseudomonas aeruginosa responding to ampicilin, kanamycin, and tetracycline resistance.

    PubMed

    Peng, Xuanxian; Xu, Changxin; Ren, Haixia; Lin, Xiangmin; Wu, Lina; Wang, Sanying

    2005-01-01

    Nosocomial wound infections by antibiotic-resistant Pseudomonas aeruginosa strains have increasing importance in hospitals. Outer membrane proteins of the bacterium have strong influence on its resistance to antibiotics. In the current study, a parallel proteomic approach was applied to analysis of sarcosine-insoluble outer membrane fraction of P. aeruginosa responding to ampicilin, kanamycin and tetracycline resistances. Eleven differential proteins with 15 spots were determined and then identified by MALDI-TOF/MS, in which four with increased OprF, MexA, OmpH, and decreased hypothetical protein (NCBI No. 15599856), six with increased OprF, OmpH, hypothetical protein (NCBI No. 15599183) and decreased OprG, MexA, conserved hypothetical protein (NCBI No. 15600371), and eight with increased OprF, MexA, OprL, probable Omp (NCBI No. 15599856), probable secretion protein (NCBI No. 15600167), OprD and decreased OprG, conserved hypothetical protein (NCBI No. 15600371) responded to ampicilin, kanamycin, and tetracycline resistances, respectively. With the exception of OprF, the other differential proteins did not show the same behaviors against the three antibiotic resistances. Compared with our previous report on E. coli Omps responding to ampicilin and tetracycline resistances, which was only a protein difference in quality between the two antibiotics, P. aeruginosa showed significant diversity against the three antibiotics. Our findings might provide valuable data for an understanding of antibiotic-resistant difference between different species of bacteria. Meanwhile, these proteins shared by different bacteria or a bacterium against different antibiotics may provide universal targets for the development of new drugs that control antibiotic-resistant bacteria.

  10. Deoxygenation affects tyrosine phosphoproteome of red cell membrane from patients with sickle cell disease.

    PubMed

    Siciliano, Angela; Turrini, Franco; Bertoldi, Mariarita; Matte, Alessandro; Pantaleo, Antonella; Olivieri, Oliviero; De Franceschi, Lucia

    2010-04-15

    Sickle cell disease (SCD) is a worldwide distributed hereditary red cell disorder related to the production of a defective form of hemoglobin, hemoglobin S (HbS). One of the hallmarks of SCD is the presence of dense, dehydrate highly adhesive sickle red blood cells (RBCs) that result from persistent membrane damage associated with HbS polymerization, abnormal activation of membrane cation transports and generation of distorted and rigid red cells with membrane perturbation and cytoskeleton dysfunction. Although modulation of phosphorylation state of the proteins from membrane and cytoskeleton networks has been proposed to participate in red cell homeostasis, much still remains to be investigated in normal and diseased red cells. Here, we report that tyrosine (Tyr-) phosphoproteome of sickle red cells was different from normal controls and was affected by deoxygenation. We found proteins, p55 and band 4.1, from the junctional complex, differently Tyr-phosphorylated in SCD RBCs compared to normal RBCs under normoxia and modulated by deoxygenation, while band 4.2 was similarly Tyr-phosphorylated in both conditions. In SCD RBCs we identified the phosphopeptides for protein 4.1R located in the protein FERM domain (Tyr-13) and for alpha-spectrin located near or in a linker region (Tyr-422 and Tyr-1498) involving protein areas crucial for their functions in the context of red cell membrane properties, suggesting that Tyr-phosphorylation may be part of the events involved in maintaining membrane mechanical stability in SCD red cells.

  11. Creating Transient Cell Membrane Pores Using a Standard Inkjet Printer

    PubMed Central

    Owczarczak, Alexander B.; Shuford, Stephen O.; Wood, Scott T.; Deitch, Sandra; Dean, Delphine

    2012-01-01

    Bioprinting has a wide range of applications and significance, including tissue engineering, direct cell application therapies, and biosensor microfabrication.1-10 Recently, thermal inkjet printing has also been used for gene transfection.8,9 The thermal inkjet printing process was shown to temporarily disrupt the cell membranes without affecting cell viability. The transient pores in the membrane can be used to introduce molecules, which would otherwise be too large to pass through the membrane, into the cell cytoplasm.8,9,11 The application being demonstrated here is the use of thermal inkjet printing for the incorporation of fluorescently labeled g-actin monomers into cells. The advantage of using thermal ink-jet printing to inject molecules into cells is that the technique is relatively benign to cells.8, 12 Cell viability after printing has been shown to be similar to standard cell plating methods1,8. In addition, inkjet printing can process thousands of cells in minutes, which is much faster than manual microinjection. The pores created by printing have been shown to close within about two hours. However, there is a limit to the size of the pore created (~10 nm) with this printing technique, which limits the technique to injecting cells with small proteins and/or particles. 8,9,11 A standard HP DeskJet 500 printer was modified to allow for cell printing.3, 5, 8 The cover of the printer was removed and the paper feed mechanism was bypassed using a mechanical lever. A stage was created to allow for placement of microscope slides and coverslips directly under the print head. Ink cartridges were opened, the ink was removed and they were cleaned prior to use with cells. The printing pattern was created using standard drawing software, which then controlled the printer through a simple print command. 3T3 fibroblasts were grown to confluence, trypsinized, and then resuspended into phosphate buffered saline with soluble fluorescently labeled g-actin monomers. The

  12. A novel unitized regenerative proton exchange membrane fuel cell

    NASA Technical Reports Server (NTRS)

    Murphy, O. J.; Cisar, A. J.; Gonzalez-Martin, A.; Salinas, C. E.; Simpson, S. F.

    1995-01-01

    A difficulty encountered in designing a unitized regenerative proton exchange membrane (PEM) fuel cell lies in the incompatibility of electrode structures and electrocatalyst materials optimized for either of the two functions (fuel cell or electrolyzer) with the needs of the other function. This difficulty is compounded in previous regenerative fuel cell designs by the fact that water, which is needed for proton conduction in the PEM during both modes of operation, is the reactant supplied to the anode in the electrolyzer mode of operation and the product formed at the cathode in the fuel cell mode. Drawbacks associated with existing regenerative fuel cells have been addressed in work performed at Lynntech. In a first innovation, electrodes function either as oxidation electrodes (hydrogen ionization or oxygen evolution) or as reduction electrodes (oxygen reduction or hydrogen evolution) in the fuel cell and electrolyzer modes, respectively. Control of liquid water within the regenerative fuel cell has been brought about by a second innovation. A novel PEM has been developed with internal channels that permit the direct access of water along the length of the membrane. Lateral diffusion of water along the polymer chains of the PEM provides the water needed at electrode/PEM interfaces. Fabrication of the novel unitized regenerative fuel cell and results obtained on testing it will be presented.

  13. Sodium channels in membrane vesicles from cultured toad bladder cells

    SciTech Connect

    Asher, C.; Moran, A.; Rossier, B.C.; Garty, H. Ben Gurion Univ., Beer-Sheva Institut de Pharmacologie de l'Universite de Lausanne )

    1988-04-01

    Electrical potential-driven {sup 22}Na{sup +} fluxes were measured in membrane vesicles prepared from TBM-18(cl23) cells (a clone of the established cell line TB-M). Fifty to seventy percent of the tracer uptake in vesicles derived from cells that were cultivated on a porous support were blocked by the diuretic amiloride. The amiloride inhibition constant was <0.1 {mu}M, indicating that this flux is mediated by the apical Na{sup +}-specific channels. Vesicles prepared from cells that were not grown on a porous support exhibited much smaller amiloride-sensitive fluxes. Two Ca{sup 2+}-dependent processes that down-regulated the channel conductance and were previously identified in native epithelia were found in the cultured cells as well. Vesicles isolated from cells that were preincubated with 5 {times} 10{sup {minus}7} M aldosterone for 16-20 h exhibited higher amiloride-sensitive conductance than vesicles derived from control, steroid-depleted cells. Thus membrane derived from TBM-18(cl23) cells can be used to characterize the epithelial Na{sup +} channel and its hormonal regulation.

  14. Membrane tether formation from outer hair cells with optical tweezers.

    PubMed Central

    Li, Zhiwei; Anvari, Bahman; Takashima, Masayoshi; Brecht, Peter; Torres, Jorge H; Brownell, William E

    2002-01-01

    Optical tweezers were used to characterize the mechanical properties of the outer hair cell (OHC) plasma membrane by pulling tethers with 4.5-microm polystyrene beads. Tether formation force and tether force were measured in static and dynamic conditions. A greater force was required for tether formations from OHC lateral wall (499 +/- 152 pN) than from OHC basal end (142 +/- 49 pN). The difference in the force required to pull tethers is consistent with an extensive cytoskeletal framework associated with the lateral wall known as the cortical lattice. The apparent plasma membrane stiffness, estimated under the static conditions by measuring tether force at different tether length, was 3.71 pN/microm for OHC lateral wall and 4.57 pN/microm for OHC basal end. The effective membrane viscosity was measured by pulling tethers at different rates while continuously recording the tether force, and estimated in the range of 2.39 to 5.25 pN x s/microm. The viscous force most likely results from the viscous interactions between plasma membrane lipids and the OHC cortical lattice and/or integral membrane proteins. The information these studies provide on the mechanical properties of the OHC lateral wall is important for understanding the mechanism of OHC electromotility. PMID:11867454

  15. Modified SPEEK membranes for direct ethanol fuel cell

    NASA Astrophysics Data System (ADS)

    Maab, Husnul; Nunes, Suzana Pereira

    Membranes with low ethanol crossover were prepared aiming their application for direct ethanol fuel cell (DEFC). They were based on (1) sulfonated poly(ether ether ketone) (SPEEK) coated with carbon molecular sieves (CMS) and (2) on SPEEK/PI homogeneous blends. The membranes were characterized concerning their water and ethanol solution uptake, water and ethanol permeability in pervaporation experiments and their performance in DEFC tests. The ethanol permeabilities for the CMS-coated (180 nm and 400 nm thick layers) SPEEK were 8.5 and 3.1 × 10 -10 kg m s -1 m -2 and for the homogeneous SPEEK/PI blends membranes with 10, 20 and 30 wt.% of PI were 4.4, 1.0 and 0.4 × 10 -10 kg m s -1 m -2 respectively, which is 2- to 50-fold lower than that for plain SPEEK (19 × 10 -10 kg m s -1 m -2). Particularly the SPEEK/PI membranes had substantially better performance than Nafion 117 ® membranes in DEFC tests at 60 °C and 90 °C.

  16. Membrane phenotypic studies in B cell lymphoproliferative disorders.

    PubMed Central

    Scott, C S; Limbert, H J; MacKarill, I D; Roberts, B E

    1985-01-01

    A total of 398 cases of B cell lymphoproliferative disease were phenotypically characterised by membrane mouse red blood cell (MRBC) receptor, surface immunoglobulin, common acute lymphoblastic leukaemia (CALLA), and FMC7 and T1 monoclonal antibody studies. Relations between chronic lymphocytic leukaemia (CLL), prolymphocytic leukaemia (PLL), and "prolymphocytoid" CLL variants were examined with particular reference to the expression of FMC7. In addition, the reactivity of TU1 monoclonal antibody with B cell disorders was established. The results suggest that despite some heterogeneity most cases may be characterised by their phenotypic patterns and that these investigations provide a reproducible basis for classification. PMID:2413082

  17. Rigid proteins and softening of biological membranes-with application to HIV-induced cell membrane softening.

    PubMed

    Agrawal, Himani; Zelisko, Matthew; Liu, Liping; Sharma, Pradeep

    2016-05-06

    A key step in the HIV-infection process is the fusion of the virion membrane with the target cell membrane and the concomitant transfer of the viral RNA. Experimental evidence suggests that the fusion is preceded by considerable elastic softening of the cell membranes due to the insertion of fusion peptide in the membrane. What are the mechanisms underpinning the elastic softening of the membrane upon peptide insertion? A broader question may be posed: insertion of rigid proteins in soft membranes ought to stiffen the membranes not soften them. However, experimental observations perplexingly appear to show that rigid proteins may either soften or harden membranes even though conventional wisdom only suggests stiffening. In this work, we argue that regarding proteins as merely non-specific rigid inclusions is flawed, and each protein has a unique mechanical signature dictated by its specific interfacial coupling to the surrounding membrane. Predicated on this hypothesis, we have carried out atomistic simulations to investigate peptide-membrane interactions. Together with a continuum model, we reconcile contrasting experimental data in the literature including the case of HIV-fusion peptide induced softening. We conclude that the structural rearrangements of the lipids around the inclusions cause the softening or stiffening of the biological membranes.

  18. Effects of motor patterns on water-soluble and membrane proteins and cholinesterase activity in subcellular fractions of rat brain tissue

    NASA Technical Reports Server (NTRS)

    Pevzner, L. Z.; Venkov, L.; Cheresharov, L.

    1980-01-01

    Albino rats were kept for a year under conditions of daily motor load or constant hypokinesia. An increase in motor activity results in a rise in the acetylcholinesterase activity determined in the synaptosomal and purified mitochondrial fractions while hypokinesia induces a pronounced decrease in this enzyme activity. The butyrylcholinesterase activity somewhat decreases in the synaptosomal fraction after hypokinesia but does not change under the motor load pattern. Motor load causes an increase in the amount of synaptosomal water-soluble proteins possessing an intermediate electrophoretic mobility and seem to correspond to the brain-specific protein 14-3-2. In the synaptosomal fraction the amount of membrane proteins with a low electrophoretic mobility and with the cholinesterase activity rises. Hypokinesia, on the contrary, decreases the amount of these membrane proteins.

  19. Effect of isosmotic removal of extracellular Na+ on cell volume and membrane potential in muscle cells.

    PubMed

    Peña-Rasgado, C; Summers, J C; McGruder, K D; DeSantiago, J; Rasgado-Flores, H

    1994-09-01

    Isosmotic removal of extracellular Na+ (Nao) is a frequently performed manipulation. With the use of isolated voltage-clamped barnacle muscle cells, the effect of this manipulation on isosmotic cell volume was studied. Replacement of Nao by tris(hydroxymethyl)aminomethane produced membrane depolarization (approximately 20 mV) and cell volume loss (approximately 14%). The membrane depolarization was verapamil insensitive but depended on extracellular Ca2+ (Cao) and was probably due to activation of intracellular Ca2+ (Cai)-dependent nonselective cation channels. The cell volume loss did not require membrane depolarization but depended on Cao. This was probably due to an increase in Cai, mediated by activation of Ca2+ influx via Na+/Ca2+ exchange. Nao replacement by Li+ also promoted membrane depolarization (approximately 20 mV) and cell volume loss (20%). Both effects were reduced (approximately 73%) but were not abolished by Cao removal. Under this condition, the remaining membrane depolarization was probably due to a higher membrane permeability of Li+ over Na+. The remaining cell volume loss was due to membrane depolarization, which probably induced Ca2+ release from intracellular stores.

  20. Vaccinia virus interactions with the cell membrane studied by new chromatic vesicle and cell sensor assays.

    PubMed

    Orynbayeva, Z; Kolusheva, S; Groysman, N; Gavrielov, N; Lobel, L; Jelinek, R

    2007-02-01

    The potential danger of cross-species viral infection points to the significance of understanding the contributions of nonspecific membrane interactions with the viral envelope compared to receptor-mediated uptake as a factor in virus internalization and infection. We present a detailed investigation of the interactions of vaccinia virus particles with lipid bilayers and with epithelial cell membranes using newly developed chromatic biomimetic membrane assays. This analytical platform comprises vesicular particles containing lipids interspersed within reporter polymer units that emit intense fluorescence following viral interactions with the lipid domains. The chromatic vesicles were employed as membrane models in cell-free solutions and were also incorporated into the membranes of epithelial cells, thereby functioning as localized membrane sensors on the cell surface. These experiments provide important insight into membrane interactions with and fusion of virions and the kinetic profiles of these processes. In particular, the data emphasize the significance of cholesterol/sphingomyelin domains (lipid rafts) as a crucial factor promoting bilayer insertion of the viral particles. Our analysis of virus interactions with polymer-labeled living cells exposed the significant role of the epidermal growth factor receptor in vaccinia virus infectivity; however, the data also demonstrated the existence of additional non-receptor-mediated mechanisms contributing to attachment of the virus to the cell surface and its internalization.

  1. Muscarinic receptor size on smooth muscle cells and membranes

    SciTech Connect

    Collins, S.M.; Jung, C.Y.; Grover, A.K.

    1986-08-01

    The loss of (/sup 3/H)quinuclidinyl benzilate ((/sup 3/H)QNB) binding following high-energy radiation was used to compare the muscarinic receptor size on single smooth muscle cells isolated by collagenase digestion from the canine stomach and on plasma membranes derived from intact gastric smooth muscle without exposure to exogenous proteolysis. Radiation inactivation of galactose oxidase (68 kdaltons), yeast alcohol dehydrogenase (160 kdaltons), and pyruvate kinase (224 kdaltons) activities were used as molecular-weight standards. Radiation inactivation of (/sup 3/H)QNB binding to rat brain membranes, which gave a target size of 86 kdaltons, served as an additional control. In isolated smooth muscle cells, the calculated size of the muscarinic receptor was 80 +/- 8 kdaltons. In contrast, in a smooth muscle enriched plasma membrane preparation, muscarinic receptor size was significantly smaller at 45 +/- 3 kdaltons. Larger molecular sizes were obtained either in the presence of protease inhibitors (62 +/- 4 kdaltons) or by using a crude membrane preparation of gastric smooth muscle 86 +/- 7 kdaltons).

  2. The organochlorine herbicide chloridazon interacts with cell membranes.

    PubMed

    Suwalsky, M; Benites, M; Villena, F; Norris, B; Quevedo, L

    1998-07-01

    Chloridazon is a widely used organochlorine herbicide. In order to evaluate its perturbing effect on cell membranes it was made to interact with human erythrocytes, frog adrenergic neuroepithelial synapse and molecular models. These consisted in multilayers of dimyristoylphosphatidylethanolamine (DMPE) and of dimyristoylphosphatidyltidylcholine (DMPC), representative of phospholipid classes located in the inner and outer monolayers of the erythrocyte membrane, respectively. X-ray diffraction showed that chloridazon interacted preferentially with DMPC multilayers. Scanning electron microscopy revealed that 0.1 mM chloridazon induced erythrocyte crenation. According to the bilayer couple hypothesis, this is due to the preferential insertion of chloridazon in the phosphatidylcholine-rich external moiety of the red cell membrane. Electrophysiological measurements showed that nerve stimulation was followed immediately by a transient increase in short-circuit current (SCC) and in the potential difference (PD) of the neuroepithelial synapse. Increasing concentrations of chloridazon caused a dose-dependent and reversible decrease of the responses of both parameters to 76% of their control values. The pesticide induced a similar (28%) significant time-dependent decrease in the basal values of the SCC and of PD. These results are in accordance with a perturbing effect of chloridazon on the phospholipid moiety of the nerve fibre membrane leading to interference with total ion transport across the nerve skin junction.

  3. Membrane electrolytic cell for minimizing hypochlorite and chlorate formation

    SciTech Connect

    Fair, D. L.; Justice, D. D.; Woodard Jr., K. E.

    1985-07-09

    An electrolytic cell for the electrolysis of an alkali metal chloride brine is comprised of an anode compartment and a cathode compartment separated by a cation exchange membrane. The anode is comprised of an unflattened expanded structure of a valve metal selected from the group consisting of titanium, tantalum, niobium, and alloys thereof. At least one side of the anode has as the electrochemically active surface an electrodeposited layer of a valve metal oxide. A plurality of cracks traverse the electrodeposited layer and a coating of a platinum metal group oxide covers the electrodeposited layer and substantially fills the cracks. The cationic exchange membrane is comprised of a laminated structure having a first surface adapted to contact an anolyte in which the ion exchange groups are predominately sulfonic acid groups. The first surface is also in contact with the electrochemically active surface of the anode. A second surface of the cation exchange membrane, adapted to contact a catholyte, has ion exchange groups which are predominately carboxylic acid groups. The cathode positioned in the cathode compartment is spaced apart from the cation exchange membrane. The cell operates with both a low chlorine overvoltage and a low oxygen overvoltage. During electrolysis of alkali metal chloride brines, the formation of hypochlorite and chlorate ions is minimized and the alkali metal hydroxides produced have low chlorate concentrations and are suitable for use without further treatment in chlorate-sensitive applications. Spent brine treatment is simplified and at reduced costs.

  4. Arabidopsis Membrane Steroid Binding Protein 1 Is Involved in Inhibition of Cell ElongationW⃞

    PubMed Central

    Yang, Xiao-Hua; Xu, Zhi-Hong; Xue, Hong-Wei

    2005-01-01

    A putative Membrane Steroid Binding Protein (designated MSBP1) was identified and functionally characterized as a negative regulator of cell elongation in Arabidopsis thaliana. The MSBP1 gene encodes a 220–amino acid protein that can bind to progesterone, 5-dihydrotestosterone, 24-epi-brassinolide (24-eBL), and stigmasterol with different affinities in vitro. Transgenic plants overexpressing MSBP1 showed short hypocotyl phenotype and increased steroid binding capacity in membrane fractions, whereas antisense MSBP1 transgenic plants showed long hypocotyl phenotypes and reduced steroid binding capacity, indicating that MSBP1 negatively regulates hypocotyl elongation. The reduced cell elongation of MSBP1-overexpressing plants was correlated with altered expression of genes involved in cell elongation, such as expansins and extensins, indicating that enhanced MSBP1 affected a regulatory pathway for cell elongation. Suppression or overexpression of MSBP1 resulted in enhanced or reduced sensitivities, respectively, to exogenous progesterone and 24-eBL, suggesting a negative role of MSBP1 in steroid signaling. Expression of MSBP1 in hypocotyls is suppressed by darkness and activated by light, suggesting that MSBP1, as a negative regulator of cell elongation, plays a role in plant photomorphogenesis. This study demonstrates the functional roles of a steroid binding protein in growth regulation in higher plants. PMID:15608331

  5. CLN3 Loss Disturbs Membrane Microdomain Properties and Protein Transport in Brain Endothelial Cells

    PubMed Central

    Tecedor, Luis; Stein, Colleen S.; Schultz, Mark L.; Farwanah, Hany; Sandhoff, Konrad

    2013-01-01

    Juvenile neuronal ceroid lipofuscinosis (JNCL) is a fatal childhood-onset neurodegenerative disorder caused by mutations in ceroid lipofuscinosis neuronal-3 (CLN3), a hydrophobic transmembrane protein of unresolved function. Previous studies indicate blood–brain barrier (BBB) defects in JNCL, and our earlier report showed prominent Cln3 expression in mouse brain endothelium. Here we find that CLN3 is necessary for normal trafficking of the microdomain-associated proteins caveolin-1, syntaxin-6, and multidrug resistance protein 1 (MDR1) in brain endothelial cells. Correspondingly, CLN3-null cells have reduced caveolae, and impaired caveolae- and MDR1-related functions including endocytosis, drug efflux, and cell volume regulation. We also detected an abnormal blood–brain barrier response to osmotic stress in vivo. Evaluation of the plasma membrane with fluorescent sphingolipid probes suggests microdomain destabilization and enhanced fluidity in CLN3-null cells. In further work we found that application of the glycosphingolipid lactosylceramide to CLN3-deficient cells rescues protein transport and caveolar endocytosis. Last, we show that CLN3 localizes to the trans-Golgi network (TGN) and partitions with buoyant microdomain fractions. We propose that CLN3 facilitates TGN-to-plasma membrane transport of microdomain-associated proteins. Insult to this pathway may underlie BBB dysfunction and contribute to JNCL pathogenesis. PMID:24227717

  6. Osteogenic cell fractions isolated from mouse tongue muscle.

    PubMed

    Harada, Koji; Harada, Toyoko; Ferdous, Tarannum; Takenawa, Takanori; Ueyama, Yoshiya

    2015-07-01

    The use of stem cells represents a promising approach for the treatment of bone defects. However, successful treatments rely upon the availability of cells that are easily obtained and that appropriately differentiate into osteoblasts. The tongue potentially represents a source of autologous cells for such purposes. In the present study, the ability of stem cell antigen-1 (Sca-1) positive cells derived from tongue muscle to differentiate into osteoblasts was investigated. The tongue muscles were excised from Jcl-ICR mice and tongue muscle-derived Sca-1-positive cells (TDSCs) were isolated from the tongue muscle using a magnetic cell separation system with microbeads. TDSCs were cultured in plastic dishes or gelatin sponges of β-tricalcium phosphate (β-TCP) with bone differentiation-inducing medium. The expression of osteogenic markers (Runx2, osterix, alkaline phosphatase, fibronectin, osteocalcin, osteonectin and osteopontin) was investigated in cultured TDSCs by western blot analysis. The formation of mineralized matrices was examined using alizarin red S and Von Kossa staining. Bone formation was investigated in cultured TDSCs by hematoxylin-eosin staining and immunohistochemistry. In the present study, the expression of Sca-1 in mouse tongue muscle was demonstrated and TDSCs were isolated at high purity. TDSCs differentiated into cells of osteoblast lineage, as demonstrated by the upregulation of osteoblastic marker expression. The formation of mineralized matrices was confirmed by alizarin red S or Von Kossa staining in vitro. Bone formation was observed in the gelatin sponges of β-TCP, which were subsequently implanted under the skin of the backs of nude mice. These results suggested that TDSCs retain their osteogenic differentiation potential and therefore the tongue muscle may be used as a source of stem cells for bone regeneration.

  7. Sterol-Rich Membrane Domains Define Fission Yeast Cell Polarity.

    PubMed

    Makushok, Tatyana; Alves, Paulo; Huisman, Stephen Michiel; Kijowski, Adam Rafal; Brunner, Damian

    2016-05-19

    Cell polarization is crucial for the functioning of all organisms. The cytoskeleton is central to the process but its role in symmetry breaking is poorly understood. We study cell polarization when fission yeast cells exit starvation. We show that the basis of polarity generation is de novo sterol biosynthesis, cell surface delivery of sterols, and their recruitment to the cell poles. This involves four phases occurring independent of the polarity factor cdc42p. Initially, multiple, randomly distributed sterol-rich membrane (SRM) domains form at the plasma membrane, independent of the cytoskeleton and cell growth. These domains provide platforms on which the growth and polarity machinery assembles. SRM domains are then polarized by the microtubule-dependent polarity factor tea1p, which prepares for monopolar growth initiation and later switching to bipolar growth. SRM polarization requires F-actin but not the F-actin organizing polarity factors for3p and bud6p. We conclude that SRMs are key to cell polarization.

  8. Proton exchange membrane fuel cell technology for transportation applications

    SciTech Connect

    Swathirajan, S.

    1996-04-01

    Proton Exchange Membrane (PEM) fuel cells are extremely promising as future power plants in the transportation sector to achieve an increase in energy efficiency and eliminate environmental pollution due to vehicles. GM is currently involved in a multiphase program with the US Department of Energy for developing a proof-of-concept hybrid vehicle based on a PEM fuel cell power plant and a methanol fuel processor. Other participants in the program are Los Alamos National Labs, Dow Chemical Co., Ballard Power Systems and DuPont Co., In the just completed phase 1 of the program, a 10 kW PEM fuel cell power plant was built and tested to demonstrate the feasibility of integrating a methanol fuel processor with a PEM fuel cell stack. However, the fuel cell power plant must overcome stiff technical and economic challenges before it can be commercialized for light duty vehicle applications. Progress achieved in phase I on the use of monolithic catalyst reactors in the fuel processor, managing CO impurity in the fuel cell stack, low-cost electrode-membrane assembles, and on the integration of the fuel processor with a Ballard PEM fuel cell stack will be presented.

  9. Voltage- and Tension-Dependent Lipid Mobility in the Outer Hair Cell Plasma Membrane

    NASA Astrophysics Data System (ADS)

    Oghalai, John S.; Zhao, Hong-Bo; Kutz, J. Walter; Brownell, William E.

    2000-01-01

    The mechanism responsible for electromotility of outer hair cells in the ear is unknown but is thought to reside within the plasma membrane. Lipid lateral diffusion in the outer hair cell plasma membrane is a sigmoidal function of transmembrane potential and bathing media osmolality. Cell depolarization or hyposmotic challenge shorten the cell and reduce membrane fluidity by half. Changing the membrane tension with amphipathic drugs results in similar reductions. These dynamic changes in membrane fluidity represent the modulation of membrane tension by lipid-protein interactions. The voltage dependence may be associated with the force-generating motors that contribute to the exquisite sensitivity of mammalian hearing.

  10. Characterization of plasma membrane proteins from ovarian cancer cells using mass spectrometry.

    PubMed

    Springer, David L; Auberry, Deanna L; Ahram, Mamoun; Adkins, Joshua N; Feldhaus, Jane M; Wahl, Jon H; Wunschel, David S; Rodland, Karin D

    To determine how the repertoire of plasma membrane proteins change with disease state, specifically related to cancer, several methods for preparation of plasma membrane proteins were evaluated. Cultured cells derived from stage IV ovarian tumors were grown to 90% confluence and harvested in buffer containing CHAPS detergent. This preparation was centrifuged at low speed to remove insoluble cellular debris resulting in a crude homogenate. Glycosylated proteins in the crude homogenate were selectively enriched using lectin affinity chromatography. The crude homogenate and the lectin purified sample were prepared for mass spectrometric evaluation. The general procedure for protein identification began with trypsin digestion of protein fractions followed by separation by reversed phase liquid chromatography that was coupled directly to a conventional tandem mass spectrometer (i.e. LCQ ion trap). Mass and fragmentation data for the peptides were searched against a human proteome data base using the informatics program SEQUEST. Using this procedure 398 proteins were identified with high confidence, including receptors, membrane-associated ligands, proteases, phosphatases, as well as structural and adhesion proteins. Results indicate that lectin chromatography provides a select subset of proteins and that the number and quality of the identifications improve as does the confidence of the protein identifications for this subset. These results represent the first step in development of methods to separate and successfully identify plasma membrane proteins from advanced ovarian cancer cells. Further characterization of plasma membrane proteins will contribute to our understanding of the mechanisms underlying progression of this deadly disease and may lead to new targeted interventions as well as new biomarkers for diagnosis.

  11. Characterization of Plasma Membrane Proteins from Ovarian Cancer Cells Using Mass Spectrometry

    SciTech Connect

    Springer, David L.; Auberry, Deanna L.; Ahram, Mamoun; Adkins, Joshua N.; Feldhaus, Jane M.; Wahl, Jon H.; Wunsch, David M.; Rodland, Karin D.

    2003-01-01

    To determine how the repertoire of plasma membrane proteins change with disease state, specifically related to cancer, several methods for preparation of plasma membrane proteins were evaluated. Cultured cells derived from stage IV ovarian tumors were grown to 90% confluence and harvested in buffer containing CHAPS detergent. This preparation was centrifuged at low speed to remove insoluble cellular debris resulting in a crude homogenate. Glycosylated proteins in the crude homogenate were selectively enriched using lectin affinity chromatography. The crude homogenate and the lectin purified sample were prepared for mass spectrometric evaluation. The general procedure for protein identification began with trypsin digestion of protein fractions followed by separation by reversed phase liquid chromatography that was coupled directly to a conventional tandem mass spectrometer (i.e. LCQ ion trap). Mass and fragmentation data for the peptides were searched against a human proteome data base using the informatics program SEQUEST. Using this procedure 398 proteins were identified with high confidence, including receptors, membrane-associated ligands, proteases, phosphatases, as well as structural and adhesion proteins. Results indicate that lectin chromatography provides a select subset of proteins and that the number and quality of the identifications improve as does the confidence of the protein identifications for this subset. These results represent the first step in development of methods to separate and successfully identify plasma membrane proteins from advanced ovarian cancer cells. Further characterization of plasma membrane proteins will contribute to our understanding of the mechanisms underlying progression of this deadly disease and may lead to new targeted interventions as well as new biomarkers for diagnosis.

  12. Characterization of Plasma Membrane Proteins from Ovarian Cancer Cells Using Mass Spectrometry

    DOE PAGES

    Springer, David L.; Auberry, Deanna L.; Ahram, Mamoun; ...

    2004-01-01

    To determine how the repertoire of plasma membrane proteins change with disease state, specifically related to cancer, several methods for preparation of plasma membrane proteins were evaluated. Cultured cells derived from stage IV ovarian tumors were grown to 90% confluence and harvested in buffer containing CHAPS detergent. This preparation was centrifuged at low speed to remove insoluble cellular debris resulting in a crude homogenate. Glycosylated proteins in the crude homogenate were selectively enriched using lectin affinity chromatography. The crude homogenate and the lectin purified sample were prepared for mass spectrometric evaluation. The general procedure for protein identification began with trypsinmore » digestion of protein fractions followed by separation by reversed phase liquid chromatography that was coupled directly to a conventional tandem mass spectrometer (i.e. LCQ ion trap). Mass and fragmentation data for the peptides were searched against a human proteome data base using the informatics program SEQUEST. Using this procedure 398 proteins were identified with high confidence, including receptors, membrane-associated ligands, proteases, phosphatases, as well as structural and adhesion proteins. Results indicate that lectin chromatography provides a select subset of proteins and that the number and quality of the identifications improve as does the confidence of the protein identifications for this subset. These results represent the first step in development of methods to separate and successfully identify plasma membrane proteins from advanced ovarian cancer cells. Further characterization of plasma membrane proteins will contribute to our understanding of the mechanisms underlying progression of this deadly disease and may lead to new targeted interventions as well as new biomarkers for diagnosis.« less

  13. Evaluation of stem cell components in retrocorneal membranes.

    PubMed

    Lee, Seok Hyun; Kim, Kyoung Woo; Kim, Mi Kyung; Chun, Yeoun Sook; Kim, Jae Chan

    2014-06-01

    The purpose of this study was to elucidate the origin and cellular composition of retrocorneal membranes (RCMs) associated with chemical burns using immunohistochemical staining for primitive cell markers. Six cases of RCMs were collected during penetrating keratoplasty. We examined RCMs with hematoxylin and eosin (H&E), periodic acid-Schiff (PAS) staining and immunohistochemical analysis using monoclonal antibodies against hematopoietic stem cells (CD34, CD133, c-kit), mesenchymal stem cells (beta-1-integrin, TGF-β, vimentin, hSTRO-1), fibroblasts (FGF-β, α-smooth muscle actin), and corneal endothelial cells (type IV collagen, CD133, VEGF, VEGFR1). Histologic analysis of RCMs revealed an organized assembly of spindle-shaped cells, pigment-laden cells, and thin collagenous matrix structures. RCMs were positive for markers of mesenchymal stem cells including beta-1-integrin, TGF-β, vimentin, and hSTRO-1. Fibroblast markers were also positive, including FGF-β and α-smooth muscle actin (SMA). In contrast, immunohistochemical staining was negative for hematopoietic stem cell markers including CD34, CD133 and c-kit as well as corneal endothelial cell markers such as type IV collagen, CD133 except VEGF and VEGFR1. Pigment-laden cells did not stain with any antibodies. The results of this study suggest that RCMs consist of a thin collagen matrix and fibroblast-like cells and may be a possible neogenetic structure produced from a lineage of bone marrow-derived mesenchymal stem cells.

  14. Interaction of Boron Nitride Nanosheets with Model Cell Membranes.

    PubMed

    Hilder, Tamsyn A; Gaston, Nicola

    2016-06-03

    Boron nitride nanomaterials have attracted attention for biomedical applications, due to their improved biocompatibility when compared with carbon nanomaterials. Recently, graphene and graphene oxide nanosheets have been shown, both experimentally and computationally, to destructively extract phospholipids from Escherichia coli. Boron nitride nanosheets (BNNSs) have exciting potential biological and environmental applications, for example the ability to remove oil from water. These applications are likely to increase the exposure of prokaryotes and eukaryotes to BNNSs. Yet, despite their promise, the interaction between BNNSs and cell membranes has not yet been investigated. Here, all-atom molecular dynamics simulations were used to demonstrate that BNNSs are spontaneously attracted to the polar headgroups of the lipid bilayer. The BNNSs do not passively cross the lipid bilayer, most likely due to the large forces experienced by the BNNSs. This study provides insight into the interaction of BNNSs with cell membranes and may aid our understanding of their improved biocompatibility.

  15. Collaboration between primitive cell membranes and soluble catalysts.

    PubMed

    Adamala, Katarzyna P; Engelhart, Aaron E; Szostak, Jack W

    2016-03-21

    One widely held model of early life suggests primitive cells consisted of simple RNA-based catalysts within lipid compartments. One possible selective advantage conferred by an encapsulated catalyst is stabilization of the compartment, resulting from catalyst-promoted synthesis of key membrane components. Here we show model protocell vesicles containing an encapsulated enzyme that promotes the synthesis of simple fatty acid derivatives become stabilized to Mg(2+), which is required for ribozyme activity and RNA synthesis. Thus, protocells capable of such catalytic transformations would have enjoyed a selective advantage over other protocells in high Mg(2+) environments. The synthetic transformation requires both the catalyst and vesicles that solubilize the water-insoluble precursor lipid. We suggest that similar modified lipids could have played a key role in early life, and that primitive lipid membranes and encapsulated catalysts, such as ribozymes, may have acted in conjunction with each other, enabling otherwise-impossible chemical transformations within primordial cells.

  16. Patterns of Nonelectrolyte Permeability in Human Red Blood Cell Membrane

    PubMed Central

    Naccache, P.; Sha'afi, R. I.

    1973-01-01

    The permeability of human red cell membrane to 90 different molecules has been measured. These solutes cover a wide spectrum of nonelectrolytes with varying chemical structure, chain length, lipid solubility, chemical reactive group, ability to form hydrogen bonds, and other properties. In general, the present study suggests that the permeability of red cell membrane to a large solute is determined by lipid solubility, its molecular size, and its hydrogen-bonding ability. The permeability coefficient increases with increasing lipid solubility and decreasing ability to form hydrogen bonds, whereas it decreases with increasing molecular size. In the case of small solutes, the predominant diffusion factor is steric hindrance augmented by lipid solubility. It is also found that replacement of a hydroxyl group by a carbonyl group or an ether linkage tends to increase permeability. On the other hand, replacement of a hydroxyl group by an amide group tends to decrease the permeability coefficient. PMID:4804758

  17. Quantitative understanding of cell signaling: The importance of membrane organization

    PubMed Central

    Radhakrishnan, Krishnan; Halász, Ádám; Vlachos, Dion; Edwards, Jeremy S.

    2010-01-01

    Systems biology modeling of signal transduction pathways traditionally employs ordinary differential equations, deterministic models based on assumptions of spatial homogeneity. However, this can be a poor approximation for certain aspects of signal transduction, especially its initial steps: the cell membrane exhibits significant spatial organization, with diffusion rates approximately two orders of magnitude slower than those in the cytosol. Thus, to unravel the complexities of signaling pathways, quantitative models must consider spatial organization as an important feature of cell signaling. Furthermore, spatial separation limits the number of molecules that can physically interact, requiring stochastic simulation methods that account for individual molecules. Herein, we discuss the need for mathematical models and experiments that appreciate the importance of spatial organization in the membrane. PMID:20829029

  18. Collaboration between primitive cell membranes and soluble catalysts

    PubMed Central

    Adamala, Katarzyna P.; Engelhart, Aaron E.; Szostak, Jack W.

    2016-01-01

    One widely held model of early life suggests primitive cells consisted of simple RNA-based catalysts within lipid compartments. One possible selective advantage conferred by an encapsulated catalyst is stabilization of the compartment, resulting from catalyst-promoted synthesis of key membrane components. Here we show model protocell vesicles containing an encapsulated enzyme that promotes the synthesis of simple fatty acid derivatives become stabilized to Mg2+, which is required for ribozyme activity and RNA synthesis. Thus, protocells capable of such catalytic transformations would have enjoyed a selective advantage over other protocells in high Mg2+ environments. The synthetic transformation requires both the catalyst and vesicles that solubilize the water-insoluble precursor lipid. We suggest that similar modified lipids could have played a key role in early life, and that primitive lipid membranes and encapsulated catalysts, such as ribozymes, may have acted in conjunction with each other, enabling otherwise-impossible chemical transformations within primordial cells. PMID:26996603

  19. Membrane associated qualitative differences in cell ultrastructure of chemically and high pressure cryofixed plant cells.

    PubMed

    Zechmann, Bernd; Müller, Maria; Zellnig, Günther

    2007-06-01

    Membrane contrast can sometimes be poor in biological samples after high pressure freezing (HPF) and freeze substitution (FS). The addition of water to the FS-medium has been shown to improve membrane contrast in animal tissue and yeast. In the present study we tested the effects of 1% and 5% water added to the FS-medium (2% osmium with 0.2% uranyl acetate in anhydrous acetone) on the quality and visibility of membranes in high pressure frozen leaf samples of Cucurbita pepo L. plants and compared them to chemically fixed cells (3% glutaraldehyde post-fixed with 1% osmium tetroxide). The addition of water to the FS-medium drastically decreased the amounts of well preserved cells and did not significantly improve the quality nor visibility of membranes. In samples that were freeze substituted in FS-media containing 1% and 5% water the width of thylakoid membranes was found to be significantly increased of about 20% and the perinuclear space was up to 76% wider in comparison to what was found in samples which were freeze substituted without water. No differences were found in the thickness of membranes between chemically and cryofixed cells that were freeze substituted in the FS-medium without water. Nevertheless, in chemically fixed cells the intrathylakoidal space was about 120% wider than in cryofixed cells that were freeze substituted with or without water. The present results demonstrate that the addition of water to the FS-medium does not improve membrane contrast but changes the width of thylakoid membranes and the perinuclear space in the present plant material. The addition of water to the FS-medium is therefore not as essential for improved membrane contrast in the investigated plant samples as it was observed in cells of animal tissues and yeast cells.

  20. The insecticide DDT decreases membrane potential and cell input resistance of cultured human liver cells.

    PubMed

    Schefczik, K; Buff, K

    1984-10-03

    The resting membrane potential, Em, and the cell input resistance, Rinp, of cultured human Chang liver cells were measured using the single electrode 'double-pulse' current clamp technique, following exposure of the cells to the insecticide DDT (20 microM). In control (unexposed) cells, the mean Em was -24 mV, and the mean Rinp was 30 M omega. Neither parameter was significantly impaired after 1 h of cell exposure to DDT. But after 7 and 48 h, the Em was depolarized by 15 and 25 mV, respectively, in parallel with a decrease of the cell input resistance. The strongly time-delayed effect of DDT on Chang liver cell membranes may indicate a mode of interaction different from excitable membranes.

  1. Do heavy ions cause microlesions in cell membranes?

    NASA Technical Reports Server (NTRS)

    Koniarek, Jan P.; Worgul, Basil V.

    1992-01-01

    The microlesion question is investigated by monitoring the electrical potential difference across the endothelium of rat corneas in vitro before, during, and after irradiation. When the corneas were exposed to 1 Gy of Fe-56 ions (450 and 600 MeV/a.m.u.), no effect was detected on this parameter. These results suggest that direct physical damage to cell membranes, as predicted by the microlesion theory, does not take place.

  2. Interferometric tomography of fuel cells for monitoring membrane water content.

    PubMed

    Waller, Laura; Kim, Jungik; Shao-Horn, Yang; Barbastathis, George

    2009-08-17

    We have developed a system that uses two 1D interferometric phase projections for reconstruction of 2D water content changes over time in situ in a proton exchange membrane (PEM) fuel cell system. By modifying the filtered backprojection tomographic algorithm, we are able to incorporate a priori information about the object distribution into a fast reconstruction algorithm which is suitable for real-time monitoring.

  3. Pfaffosidic Fraction from Hebanthe paniculata Induces Cell Cycle Arrest and Caspase-3-Induced Apoptosis in HepG2 Cells

    PubMed Central

    da Silva, Tereza Cristina; Cogliati, Bruno; Latorre, Andréia Oliveira; Akisue, Gokithi; Nagamine, Márcia Kazumi; Haraguchi, Mitsue; Hansen, Daiane; Sanches, Daniel Soares; Dagli, Maria Lúcia Zaidan

    2015-01-01

    Hebanthe paniculata roots (formerly Pfaffia paniculata and popularly known as Brazilian ginseng) show antineoplastic, chemopreventive, and antiproliferative properties. Functional properties of these roots and their extracts are usually attributed to the pfaffosidic fraction, which is composed mainly by pfaffosides A–F. However, the therapeutic potential of this fraction in cancer cells is not yet entirely understood. This study aimed to analyze the antitumoral effects of the purified pfaffosidic fraction or saponinic fraction on the human hepatocellular carcinoma HepG2 cell line. Cellular viability, proliferation, and apoptosis were evaluated, respectively, by MTT assay, BrdU incorporation, activated caspase-3 immunocytochemistry, and DNA fragmentation assay. Cell cycle was analyzed by flow cytometry and the cell cycle-related proteins were analyzed by quantitative PCR and Western blot. The cells exposed to pfaffosidic fraction had reduced viability and cellular growth, induced G2/M at 48 h or S at 72 h arrest, and increased sub-G1 cell population via cyclin E downregulation, p27KIP1 overexpression, and caspase-3-induced apoptosis, without affecting the DNA integrity. Antitumoral effects of pfaffosidic fraction from H. paniculata in HepG2 cells originated by multimechanisms of action might be associated with cell cycle arrest in the S phase, by CDK2 and cyclin E downregulation and p27KIP1 overexpression, besides induction of apoptosis through caspase-3 activation. PMID:26075002

  4. A micromechanic study of cell polarity and plasma membrane cell body coupling in Dictyostelium.

    PubMed Central

    Merkel, R; Simson, R; Simson, D A; Hohenadl, M; Boulbitch, A; Wallraff, E; Sackmann, E

    2000-01-01

    We used micropipettes to aspirate leading and trailing edges of wild-type and mutant cells of Dictyostelium discoideum. Mutants were lacking either myosin II or talin, or both proteins simultaneously. Talin is a plasma membrane-associated protein important for the coupling between membrane and actin cortex, whereas myosin II is a cytoplasmic motor protein essential for the locomotion of Dictyostelium cells. Aspiration into the pipette occurred above a threshold pressure only. For all cells containing talin this threshold was significantly lower at the leading edge of an advancing cell as compared to its rear end, whereas we found no such difference in cells lacking talin. Wild-type and talin-deficient cells were able to retract from the pipette against an applied suction pressure. In these cells, retraction was preceded by an accumulation of myosin II in the tip of the aspirated cell lobe. Mutants lacking myosin II could not retract, even if the suction pressures were removed after aspiration. We interpreted the initial instability and the subsequent plastic deformation of the cell surface during aspiration in terms of a fracture between the cell plasma membrane and the cell body, which may involve destruction of part of the cortex. Models are presented that characterize the coupling strength between membrane and cell body by a surface energy sigma. We find sigma approximately 0.6(1.6) mJ/m(2) at the leading (trailing) edge of wild-type cells. PMID:10920005

  5. Polymer Electrolyte Membrane (PEM) Fuel Cells Modeling and Optimization

    NASA Astrophysics Data System (ADS)

    Zhang, Zhuqian; Wang, Xia; Shi, Zhongying; Zhang, Xinxin; Yu, Fan

    2006-11-01

    Performance of polymer electrolyte membrane (PEM) fuel cells is dependent on operating parameters and designing parameters. Operating parameters mainly include temperature, pressure, humidity and the flow rate of the inlet reactants. Designing parameters include reactants distributor patterns and dimensions, electrodes dimensions, and electrodes properties such as porosity, permeability and so on. This work aims to investigate the effects of various designing parameters on the performance of PEM fuel cells, and the optimum values will be determined under a given operating condition.A three-dimensional steady-state electrochemical mathematical model was established where the mass, fluid and thermal transport processes are considered as well as the electrochemical reaction. A Powell multivariable optimization algorithm will be applied to investigate the optimum values of designing parameters. The objective function is defined as the maximum potential of the electrolyte fluid phase at the membrane/cathode interface at a typical value of the cell voltage. The robustness of the optimum design of the fuel cell under different cell potentials will be investigated using a statistical sensitivity analysis. By comparing with the reference case, the results obtained here provide useful tools for a better design of fuel cells.

  6. Proton electrolyte membrane properties and direct methanol fuel cell performance. II. Fuel cell performance and membrane properties effects

    NASA Astrophysics Data System (ADS)

    Silva, V. S.; Schirmer, J.; Reissner, R.; Ruffmann, B.; Silva, H.; Mendes, A.; Madeira, L. M.; Nunes, S. P.

    In order to study the relationship between the properties of proton electrolyte membranes (PEMs), obtained through standard characterization methods, and the direct methanol fuel cell (DMFC) performance, inorganic-organic hybrid membranes, modified via in situ hydrolysis, were used in a membrane electrolyte assembly (MEA) for DMFC application. The membranes, the characterization of which was performed in the previous paper of this series, were based on sulfonated poly(ether ether ketone) (sPEEK) with a sulfonation degree (SD) of 87% and were loaded with different amounts of zirconium oxide (5.0, 7.5, 10.0, 12.5 wt.%). The standard characterization methods applied were impedance spectroscopy (proton conductivity), water uptake, and pervaporation (permeability to methanol). The MEAs were characterized investigating the DMFC current-voltage polarization curves, constant voltage current (CV, 35 mV), and open-circuit voltage (OCV). The fuel cell ohmic resistance (null phase angle impedance, NPAI) and CO 2 concentration in the cathode outlet were also measured. The characterization results show that the incorporation of the inorganic oxide in the polymer network decreases the DMFC current density for CV experiments, CO 2 concentration in the cathode outlet for both OCV and CV experiments and, finally, the maximum power density output. The opposite effect was verified in terms of the NPAI (ohmic resistance) for both OCV and CV experiments. A good agreement was found between the studied DMFC performance parameters and the characterization results evaluated by impedance spectroscopy, water uptake and pervaporation experiments.

  7. Latent progenitor cells as potential regulators for tympanic membrane regeneration

    NASA Astrophysics Data System (ADS)

    Kim, Seung Won; Kim, Jangho; Seonwoo, Hoon; Jang, Kyung-Jin; Kim, Yeon Ju; Lim, Hye Jin; Lim, Ki-Taek; Tian, Chunjie; Chung, Jong Hoon; Choung, Yun-Hoon

    2015-06-01

    Tympanic membrane (TM) perforation, in particular chronic otitis media, is one of the most common clinical problems in the world and can present with sensorineural healing loss. Here, we explored an approach for TM regeneration where the latent progenitor or stem cells within TM epithelial layers may play an important regulatory role. We showed that potential TM stem cells present highly positive staining for epithelial stem cell markers in all areas of normal TM tissue. Additionally, they are present at high levels in perforated TMs, especially in proximity to the holes, regardless of acute or chronic status, suggesting that TM stem cells may be a potential factor for TM regeneration. Our study suggests that latent TM stem cells could be potential regulators of regeneration, which provides a new insight into this clinically important process and a potential target for new therapies for chronic otitis media and other eardrum injuries.

  8. MECHANISM OF GLUCOSE TRANSPORT ACROSS THE YEAST CELL MEMBRANE

    PubMed Central

    Cirillo, Vincent P.

    1962-01-01

    Cirillo, Vincent P. (Seton Hall College of Medicine and Dentistry, Jersey City, N.J.). Mechanism of glucose transport across the yeast cell membrane. J. Bacteriol. 84:485–491. 1962.—The kinetics of d-glucose and l-sorbose transport was studied in Saccharomyces cerevisiae inhibited with iodoacetic acid under nitrogen to prevent glucose metabolism. d-Glucose was found to compete with l-sorbose for a common membrane transport system with an apparent affinity greater than 25 times that of sorbose. A comparison of the net rate of glucose and sorbose transport at 50 and 500 mm external concentration showed that glucose transport is greater than that of sorbose from the lower concentration, but sorbose transport is greater than glucose at the higher concentration. This reversal of transport rate of two sugars with markedly different affinities is predicted by the membrane carrier theory. A further prediction of carrier theory was confirmed by the demonstration that the rate of glucose transport into fructose-loaded cells is greater than into unloaded cells. PMID:14021412

  9. Laser-photophoretic migration and fractionation of human blood cells.

    PubMed

    Monjushiro, Hideaki; Tanahashi, Yuko; Watarai, Hitoshi

    2013-05-13

    Laser photophoretic migration behavior of human blood cells in saline solution was investigated under the irradiation of Nd:YAG laser beam (532 nm) in the absence and the presence of the flow in a fused silica capillary. Red blood cells (RBC) were migrated faster than white blood cells (WBC) and blood pellets to the direction of propagation of laser light. The observed photophoretic velocity of RBC was about 11 times faster than those of others. This was understood from the larger photophoretic efficiency of RBC than that of WBC, which was simulated based on the Mie scattering theory. Furthermore, it was found that, during the photophoretic migration, RBCs spontaneously orientated parallel to the migration direction so as to reduce the drag force. Finally, it was demonstrated that RBC and WBC were separated in a micro-channel flow system by the laser photophoresis.

  10. Analysis of Nuclear RNA Interference (RNAi) in Human Cells by Subcellular Fractionation and Argonaute Loading

    PubMed Central

    Gagnon, Keith T.; Li, Liande; Janowski, Bethany A.; Corey, David R.

    2014-01-01

    RNA interference (RNAi) is well known for its ability to regulate gene expression in the cytoplasm of mammalian cells. In mammalian cell nuclei, however, the impact of RNAi has remained more controversial. A key technical hurdle has been a lack of optimized protocols for the isolation and analysis of cell nuclei. Here we describe a simplified protocol for nuclei isolation from cultured cells that incorporates a method for obtaining nucleoplasmic and chromatin fractions and removing cytoplasmic contamination. Cell fractions can then be used to detect the presence and activity of RNAi factors in the nucleus. We present a protocol for investigating an early step in RNAi, Argonaute protein loading with small RNAs, which is enabled by our improved extract preparations. These protocols facilitate characterization of nuclear RNAi and can be applied to the analysis of other nuclear proteins and pathways. From cellular fractionation to analysis of Argonaute loading results, this protocol takes 4–6 d to complete. PMID:25079428

  11. Ethyl ether fraction of Gastrodia elata Blume protects amyloid beta peptide-induced cell death.

    PubMed

    Kim, Hyeon-Ju; Moon, Kwang-Deog; Lee, Dong-Seok; Lee, Sang-Han

    2003-01-01

    Alzheimer's disease is the most common cause of dementia in the elderly. Recently, it has been reported that Alzheimer's disease is associated with cell death in neuronal cells including the hippocampus. Amyloid beta-peptide stimulates neuronal cell death, but the underlying signaling pathways are poorly understood. In order to develop anti-dementia agents with potential therapeutic value, we examined the effect of the herbal compound Gastrodia elata Blume (GEB) on neuronal cell death induced by amyloid beta-peptide in IMR-32 neuroblastoma cells. The fractionation of GEB was carried out in various solvents. The hydroxyl radical scavenging effect of the ethyl ether fraction was more potent than any other fractions. In cells treated with amyloid beta-peptide, the neuroprotective effect of the ethyl ether, chloroform, and butanol fractions was 92, 44, and 39%, respectively, compared with control. Taken together, these results suggest that the ethyl ether fraction of GEB contains one or more compounds that dramatically reduce amyloid beta-peptide induced neuronal cell death in vitro.

  12. 16K Fractionation of 3T3-L1 Adipocytes to Produce a Crude GLUT4-Containing Vesicle Fraction.

    PubMed

    Sadler, Jessica B A; Lamb, Christopher A; Gould, Gwyn W; Bryant, Nia J

    2016-03-01

    The insulin-sensitive pool of glucose transporter isoform 4 (GLUT4) can be isolated from total cell membranes using the 16K fractionation protocol, described here. This method produces a light membrane-containing supernatant that includes the insulin-sensitive pool of GLUT4 in GLUT4 storage vesicles. The 16K pellet fraction contains the heavy membranes (including the plasma membrane, mitochondria, nuclei, Golgi apparatus, and endoplasmic reticulum). The distribution of proteins between the two fractions is determined via immunoblotting. By subjecting insulin-stimulated versus unstimulated cells to this protocol, the mobilization of proteins out of the insulin-sensitive GLUT4 pool can be assessed.

  13. Graphene-doped electrospun nanofiber membrane electrodes and proton exchange membrane fuel cell performance

    NASA Astrophysics Data System (ADS)

    Wei, Meng; Jiang, Min; Liu, Xiaobo; Wang, Min; Mu, Shichun

    2016-09-01

    A rational electrode structure can allow proton exchange membrane (PEM) fuel cells own high performance with a low noble metal loading and an optimal transport pathway for reaction species. In this study, we develop a graphene doped polyacrylonitile (PAN)/polyvinylident fluoride (PVDF) (GPP) electrospun nanofiber electrode with improved electrical conductivity and high porosity, which could enhance the triple reaction boundary and promote gas and water transport throughout the porous electrode. Thus the increased electrochemical active surface area (ECSA) of Pt catalysts and fuel cell performance can be expected. As results, the ECSA of hot-pressed electrospun electrodes with 2 wt% graphene oxide (GO) is up to 84.3 m2/g, which is greatly larger than that of the conventional electrode (59.5 m2/g). Significantly, the GPP nanofiber electrospun electrode with Pt loading of 0.2 mg/cm2 exhibits higher fuel cell voltage output and stability than the conventional electrode.

  14. Quantitative analysis of cell surface membrane proteins using membrane-impermeable chemical probe coupled with 18O labeling.

    PubMed

    Zhang, Haizhen; Brown, Roslyn N; Qian, Wei-Jun; Monroe, Matthew E; Purvine, Samuel O; Moore, Ronald J; Gritsenko, Marina A; Shi, Liang; Romine, Margaret F; Fredrickson, James K; Pasa-Tolić, Ljiljana; Smith, Richard D; Lipton, Mary S

    2010-05-07

    We report a mass spectrometry-based strategy for quantitative analysis of cell surface membrane proteome changes. The strategy includes enrichment of surface membrane proteins using a membrane-impermeable chemical probe followed by stable isotope (18)O labeling and LC-MS analysis. We applied this strategy for enriching membrane proteins expressed by Shewanella oneidensis MR-1, a Gram-negative bacterium with known metal-reduction capability via extracellular electron transfer between outer membrane proteins and extracellular electron receptors. LC/MS/MS analysis resulted in the identification of about 400 proteins with 79% of them being predicted to be membrane localized. Quantitative aspects of the membrane enrichment were shown by peptide level (16)O and (18)O labeling of proteins from wild-type and mutant cells (generated from deletion of a type II secretion protein, GspD) prior to LC-MS analysis. Using a chemical probe labeled pure protein as an internal standard for normalization, the quantitative data revealed reduced abundances in Delta gspD mutant cells of many outer membrane proteins including the outer membrane c-type cytochromes OmcA and MtrC, in agreement with a previous report that these proteins are substrates of the type II secretion system.

  15. A review of polymer electrolyte membranes for direct methanol fuel cells

    NASA Astrophysics Data System (ADS)

    Neburchilov, Vladimir; Martin, Jonathan; Wang, Haijiang; Zhang, Jiujun

    This review describes the polymer electrolyte membranes (PEM) that are both under development and commercialized for direct methanol fuel cells (DMFC). Unlike the membranes for hydrogen fuelled PEM fuel cells, among which perfluorosulfonic acid based membranes show complete domination, the membranes for DMFC have numerous variations, each has its advantages and disadvantages. No single membrane is emerging as absolutely superior to others. This review outlines the prospects of the currently known membranes for DMFC. The membranes are evaluated according to various properties, including: methanol crossover, proton conductivity, durability, thermal stability and maximum power density. Hydrocarbon and composite fluorinated membranes currently show the most potential for low cost membranes with low methanol permeability and high durability. Some of these membranes are already beginning to impact the portable fuel cell market.

  16. Fascin, an Actin-bundling Protein, Induces Membrane Protrusions and Increases Cell Motility of Epithelial Cells

    PubMed Central

    Yamashiro, Shigeko; Yamakita, Yoshihiko; Ono, Shoichiro; Matsumura, Fumio

    1998-01-01

    Fascin is an actin-bundling protein that is found in membrane ruffles, microspikes, and stress fibers. The expression of fascin is greatly increased in many transformed cells, as well as in specialized normal cells including neuronal cells and antigen-presenting dendritic cells. A morphological characteristic common to these cells expressing high levels of fascin is the development of many membrane protrusions in which fascin is predominantly present. To examine whether fascin contributes to the alterations in microfilament organization at the cell periphery, we have expressed fascin in LLC-PK1 epithelial cells to levels as high as those found in transformed cells and in specialized normal cells. Expression of fascin results in large changes in morphology, the actin cytoskeleton, and cell motility: fascin-transfected cells form an increased number of longer and thicker microvilli on apical surfaces, extend lamellipodia-like structures at basolateral surfaces, and show disorganization of cell–cell contacts. Cell migration activity is increased by 8–17 times when assayed by modified Boyden chamber. Microinjection of a fascin protein into LLC-PK1 cells causes similar morphological alterations including the induction of lamellipodia at basolateral surfaces and formation of an increased number of microvilli on apical surfaces. Furthermore, microinjection of fascin into REF-52 cells, normal fibroblasts, induces the formation of many lamellipodia at all regions of cell periphery. These results together suggest that fascin is directly responsible for membrane protrusions through reorganization of the microfilament cytoskeleton at the cell periphery. PMID:9571235

  17. Production of membrane proteins through the wheat-germ cell-free technology.

    PubMed

    Nozawa, Akira; Nanamiya, Hideaki; Tozawa, Yuzuru

    2010-01-01

    Membrane proteins play crucial roles in various processes. However, biochemical characterization of the membrane proteins remains challenging due to the difficulty in producing membrane proteins in a functional state. Here, we describe a novel method for the production of functional membrane proteins based on a wheat germ cell-free translation system. Using this method, functional membrane proteins are successfully synthesized in the presence of liposomes and a detergent. In addition, the synthesized membrane proteins are easily purified from the cell-free translation mixture as proteoliposomes by sucrose density gradient ultracentrifugation. These advantages over conventional approaches are very helpful for the clarification of the function of membrane proteins.

  18. Self-humidified proton exchange membrane fuel cells: Operation of larger cells and fuel cell stacks

    SciTech Connect

    Dhar, H.P.; Lee, J.H.; Lewinski, K.A.

    1996-12-31

    The PEM fuel cell is promising as the power source for use in mobile and stationary applications primarily because of its high power density, all solid components, and simplicity of operation. For wide acceptability of this power source, its cost has to be competitive with the presently available energy sources. The fuel cell requires continuous humidification during operation as a power source. The humidification unit however, increases fuel cell volume, weight, and therefore decreases its overall power density. Great advantages in terms of further fuel cell simplification can be achieved if the humidification process can be eliminated or minimized. In addition, cost reductions are associated with the case of manufacturing and operation. At BCS Technology we have developed a technology of self-humidified operation of PEM fuel cells based on the mass balance of the reactants and products and the ability of membrane electrode assembly (MEA) to retain water necessary for humidification under the cell operating conditions. The reactants enter the fuel cell chambers without carrying any form of water, whether in liquid or vapor form. Basic principles of self-humidified operation of fuel cells as practiced by BCS Technology, Inc. have been presented previously in literature. Here, we report the operation of larger self-humidified single cells and fuel cell stacks. Fuel cells of areas Up to 100 cm{sup 2} have been operated. We also show the self-humidified operation of fuel cell stacks of 50 and 100 cm{sup 2} electrode areas.

  19. Nanofiber Composite Membranes for Alkaline Fuel Cells: Generation of Compositional, Morphological, and Functional Property Relationships

    DTIC Science & Technology

    2015-12-01

    properties of nanofiber composite anion-exchange membranes for alkaline fuel cells. A new membrane fabrication strategy, utilizing polymer fiber...electrospinning, will be employed to make hydroxide-conducting membranes with an entirely new morphology, where one electrospun polymer provides pathways...for ion conductivity and the second electrospun polymer restricts ionomer swelling and imparts mechanical strength to the membrane. The functional

  20. Traversal of cells by radiation and absorbed fraction estimates for electrons and alpha particles

    SciTech Connect

    Eckerman, K.F.; Ryman, J.C.; Taner, A.C.; Kerr, G.D.

    1985-01-01

    Consideration of the pathlength which radiation traverses in a cell is central to algorithms for estimating energy deposition on a cellular level. Distinct pathlength distributions occur for radionuclides: (1) uniformly distributed in space about the cell (referred to as -randomness); (2) uniformly distributed on the surface of the cell (S-randomness); and (3) uniformly distributed within the cell volume (I-randomness). For a spherical cell of diameter d, the mean pathlengths are 2/3d, 1/2d, and 3/4d, respectively, for these distributions. Algorithms for simulating the path of radiation through a cell are presented and the absorbed fraction in the cell and its nucleus are tabulated for low energy electrons and alpha particles emitted on the surface of spherical cells. The algorithms and absorbed fraction data should be of interest to those concerned with the dosimetry of radionuclide-labeled monoclonal antibodies. 8 refs., 3 figs., 2 tabs.