Sample records for membranes regulating cell

  1. Nuclear myosin I regulates cell membrane tension

    Czech Academy of Sciences Publication Activity Database

    Venit, Tomáš; Kalendová, Alžběta; Petr, Martin; Dzijak, Rastislav; Pastorek, Lukáš; Rohožková, Jana; Malohlava, M.; Hozák, Pavel


    Roč. 6, AUG 2 (2016), č. článku 30864. ISSN 2045-2322 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0109; GA ČR GAP305/11/2232; GA MŠk(CZ) LO1304 Institutional support: RVO:68378050 Keywords : neuronal growth cone * rna-polymerase-ii * cancer cells * phosphatidylinositol 4,5-bisphosphate * myo1c * actin * transcription * complex * motor * afm Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.259, year: 2016

  2. Key factors regulating the mass delivery of macromolecules to model cell membranes

    DEFF Research Database (Denmark)

    Campbell, Richard A.; Watkins, Erik B.; Jagalski, Vivien


    We show that both gravity and electrostatics are key factors regulating interactions between model cell membranes and self-assembled liquid crystalline aggregates of dendrimers and phospholipids. The system is a proxy for the trafficking of reservoirs of therapeutic drugs to cell membranes for sl...... of the aggregates to activate endocytosis pathways on specific cell types is discussed in the context of targeted drug delivery applications.......We show that both gravity and electrostatics are key factors regulating interactions between model cell membranes and self-assembled liquid crystalline aggregates of dendrimers and phospholipids. The system is a proxy for the trafficking of reservoirs of therapeutic drugs to cell membranes for slow...

  3. Vaginal epithelial cells regulate membrane adhesiveness to co-ordinate bacterial adhesion. (United States)

    Younes, Jessica A; Klappe, Karin; Kok, Jan Willem; Busscher, Henk J; Reid, Gregor; van der Mei, Henny C


    Vaginal epithelium is colonized by different bacterial strains and species. The bacterial composition of vaginal biofilms controls the balance between health and disease. Little is known about the relative contribution of the epithelial and bacterial cell surfaces to bacterial adhesion and whether and how adhesion is regulated over cell membrane regions. Here, we show that bacterial adhesion forces with cell membrane regions not located above the nucleus are stronger than with regions above the nucleus both for vaginal pathogens and different commensal and probiotic lactobacillus strains involved in health. Importantly, adhesion force ratios over membrane regions away from and above the nucleus coincided with the ratios between numbers of adhering bacteria over both regions. Bacterial adhesion forces were dramatically decreased by depleting the epithelial cell membrane of cholesterol or sub-membrane cortical actin. Thus, epithelial cells can regulate membrane regions to which bacterial adhesion is discouraged, possibly to protect the nucleus. © 2015 John Wiley & Sons Ltd.

  4. Membrane mechanisms and intracellular signalling in cell volume regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay; Dunham, Philip B.


    Volume regulation, Signal transduction, Calcium-calmodulin, Stretch-activated channels, Eicosanoids, Macromolecular crowding, Cytoskeleton, Protein phosphorylation, dephosphorylation.......Volume regulation, Signal transduction, Calcium-calmodulin, Stretch-activated channels, Eicosanoids, Macromolecular crowding, Cytoskeleton, Protein phosphorylation, dephosphorylation....

  5. Peptide-modified PELCL electrospun membranes for regulation of vascular endothelial cells

    International Nuclear Information System (INIS)

    Zhou, Fang; Jia, Xiaoling; Yang, Yang; Yang, Qingmao; Gao, Chao; Zhao, Yunhui; Fan, Yubo; Yuan, Xiaoyan


    The efficiency of biomaterials used in small vascular repair depends greatly on their ability to interact with vascular endothelial cells (VECs). Rapid endothelialization of the vascular grafts is a promising way to prevent thrombosis and intimal hyperplasia. In this work, modification of electrospun membranes of poly(ethylene glycol)-b-poly(L-lactide-co-ε-caprolactone) (PELCL) by three different peptides for regulation of VECs were studied in order to obtain ideal bioactive biomaterials as small diameter vascular grafts. QK (a mimetic peptide to vascular endothelial growth factor), Arg-Glu-Asp-Val (REDV, a specific adhesive peptide to VECs) and Val-Ala-Pro-Gly (VAPG, a specific adhesive peptide to vascular smooth muscle cells) were investigated. Surface properties of the modified membranes and the response of VECs were verified. It was found that protein adsorption and platelet adhesion were effectively suppressed with the introduction of QK, REDV or VAPG peptides on the PELCL electrospun membranes. Both QK- and REDV-modified electrospun membranes could accelerate the proliferation of VECs in the first 9 days, and the QK-modified electrospun membrane promoted cell proliferation more significantly than the REDV-modified one. The REDV-modified PELCL membrane was the most favorable for VECs adhesion than QK- and VAPG-modified membranes. It was suggested that QK- or REDV-modified PELCL electrospun membranes may have great potential applications in cardiovascular biomaterials for rapid endothelialization in situ. - Highlights: • A series of peptide-modified PELCL electrospun membranes were prepared. • Hemocompatibility of the membranes was greatly improved by the modification. • QK-modified PELCL membrane promoted VECs proliferation more significantly. • REDV-modified PELCL membrane was the most favorable for VEC adhesion.

  6. Cell cycle-regulated membrane binding of NuMA contributes to efficient anaphase chromosome separation. (United States)

    Zheng, Zhen; Wan, Qingwen; Meixiong, Gerry; Du, Quansheng


    Accurate and efficient separation of sister chromatids during anaphase is critical for faithful cell division. It has been proposed that cortical dynein-generated pulling forces on astral microtubules contribute to anaphase spindle elongation and chromosome separation. In mammalian cells, however, definitive evidence for the involvement of cortical dynein in chromosome separation is missing. It is believed that dynein is recruited and anchored at the cell cortex during mitosis by the α subunit of heterotrimeric G protein (Gα)/mammalian homologue of Drosophila Partner of Inscuteable/nuclear mitotic apparatus (NuMA) ternary complex. Here we uncover a Gα/LGN-independent lipid- and membrane-binding domain at the C-terminus of NuMA. We show that the membrane binding of NuMA is cell cycle regulated-it is inhibited during prophase and metaphase by cyclin-dependent kinase 1 (CDK1)-mediated phosphorylation and only occurs after anaphase onset when CDK1 activity is down-regulated. Further studies indicate that cell cycle-regulated membrane association of NuMA underlies anaphase-specific enhancement of cortical NuMA and dynein. By replacing endogenous NuMA with membrane-binding-deficient NuMA, we can specifically reduce the cortical accumulation of NuMA and dynein during anaphase and demonstrate that cortical NuMA and dynein contribute to efficient chromosome separation in mammalian cells.

  7. Host Cell Plasma Membrane Phosphatidylserine Regulates the Assembly and Budding of Ebola Virus. (United States)

    Adu-Gyamfi, Emmanuel; Johnson, Kristen A; Fraser, Mark E; Scott, Jordan L; Soni, Smita P; Jones, Keaton R; Digman, Michelle A; Gratton, Enrico; Tessier, Charles R; Stahelin, Robert V


    Lipid-enveloped viruses replicate and bud from the host cell where they acquire their lipid coat. Ebola virus, which buds from the plasma membrane of the host cell, causes viral hemorrhagic fever and has a high fatality rate. To date, little has been known about how budding and egress of Ebola virus are mediated at the plasma membrane. We have found that the lipid phosphatidylserine (PS) regulates the assembly of Ebola virus matrix protein VP40. VP40 binds PS-containing membranes with nanomolar affinity, and binding of PS regulates VP40 localization and oligomerization on the plasma membrane inner leaflet. Further, alteration of PS levels in mammalian cells inhibits assembly and egress of VP40. Notably, interactions of VP40 with the plasma membrane induced exposure of PS on the outer leaflet of the plasma membrane at sites of egress, whereas PS is typically found only on the inner leaflet. Taking the data together, we present a model accounting for the role of plasma membrane PS in assembly of Ebola virus-like particles. The lipid-enveloped Ebola virus causes severe infection with a high mortality rate and currently lacks FDA-approved therapeutics or vaccines. Ebola virus harbors just seven genes in its genome, and there is a critical requirement for acquisition of its lipid envelope from the plasma membrane of the human cell that it infects during the replication process. There is, however, a dearth of information available on the required contents of this envelope for egress and subsequent attachment and entry. Here we demonstrate that plasma membrane phosphatidylserine is critical for Ebola virus budding from the host cell plasma membrane. This report, to our knowledge, is the first to highlight the role of lipids in human cell membranes in the Ebola virus replication cycle and draws a clear link between selective binding and transport of a lipid across the membrane of the human cell and use of that lipid for subsequent viral entry. Copyright © 2015, American

  8. Electrophoresis of cell membrane heparan sulfate regulates galvanotaxis in glial cells. (United States)

    Huang, Yu-Ja; Schiapparelli, Paula; Kozielski, Kristen; Green, Jordan; Lavell, Emily; Guerrero-Cazares, Hugo; Quinones-Hinojosa, Alfredo; Searson, Peter


    Endogenous electric fields modulate many physiological processes by promoting directional migration, a process known as galvanotaxis. Despite the importance of galvanotaxis in development and disease, the mechanism by which cells sense and migrate directionally in an electric field remains unknown. Here, we show that electrophoresis of cell surface heparan sulfate (HS) critically regulates this process. HS was found to be localized at the anode-facing side in fetal neural progenitor cells (fNPCs), fNPC-derived astrocytes and brain tumor-initiating cells (BTICs), regardless of their direction of galvanotaxis. Enzymatic removal of HS and other sulfated glycosaminoglycans significantly abolished or reversed the cathodic response seen in fNPCs and BTICs. Furthermore, Slit2, a chemorepulsive ligand, was identified to be colocalized with HS in forming a ligand gradient across cellular membranes. Using both imaging and genetic modification, we propose a novel mechanism for galvanotaxis in which electrophoretic localization of HS establishes cell polarity by functioning as a co-receptor and provides repulsive guidance through Slit-Robo signaling. © 2017. Published by The Company of Biologists Ltd.

  9. Ankyrin regulates KATP channel membrane trafficking and gating in excitable cells (United States)

    Kline, Crystal F.; Hund, Thomas J.; Mohler, Peter J.


    K(ATP) channels play critical roles in many cellular functions by coupling cell metabolic status to electrical activity. First discovered in cardiomyocytes,1 KATP channels (comprised of Kir6.x and SUR subunits) have since been found in many other tissues, including pancreatic beta cells, skeletal muscle, smooth muscle, brain, pituitary and kidney. By linking cellular metabolic state with membrane potential, KATP channels are able to regulate a number of cellular functions such as hormone secretion, vascular tone and excitability. Specifically, a reduction in metabolism causes a decrease in the ATP:ADP ratio, opening of KATP channels, K+ efflux, membrane hyperpolarization, and suppression of electrical activity. Conversely, increased cellular metabolism causes an increase in the ATP:ADP ratio that leads to closure of the KATP channel, membrane depolarization, and stimulation of cell electrical activity. PMID:19901534

  10. Phosphorylation of SNAP-23 regulates its dynamic membrane association during mast cell exocytosis

    Directory of Open Access Journals (Sweden)

    Pieu Naskar


    Full Text Available Upon allergen challenge, mast cells (MCs respond by releasing pre-stored mediators from their secretory granules by the transient mechanism of porosome-mediated cell secretion. The target SNARE SNAP-23 has been shown to be important for MC exocytosis, and our previous studies revealed the presence of one basal (Thr102 and two induced (Ser95 and Ser120 phosphorylation sites in its linker region. To study the role of SNAP-23 phosphorylation in the regulation of exocytosis, green fluorescence protein-tagged wild-type SNAP-23 (GFP-SNAP-23 and its phosphorylation mutants were transfected into rat basophilic leukemia (RBL-2H3 MCs. Studies on GFP-SNAP-23 transfected MCs revealed some dynamic changes in SNAP-23 membrane association. SNAP-23 was associated with plasma membrane in resting MCs, however, on activation a portion of it translocated to cytosol and internal membranes. These internal locations were secretory granule membranes. This dynamic change in the membrane association of SNAP-23 in MCs may be important for mediating internal granule-granule fusions in compound exocytosis. Further studies with SNAP-23 phosphorylation mutants revealed an important role for the phosphorylation at Thr102 in its initial membrane association, and of induced phosphorylation at Ser95 and Ser120 in its internal membrane association, during MC exocytosis.

  11. Calcium regulates cell death in cancer: Roles of the mitochondria and mitochondria-associated membranes (MAMs). (United States)

    Danese, Alberto; Patergnani, Simone; Bonora, Massimo; Wieckowski, Mariusz R; Previati, Maurizio; Giorgi, Carlotta; Pinton, Paolo


    Until 1972, the term 'apoptosis' was used to differentiate the programmed cell death that naturally occurs in organismal development from the acute tissue death referred to as necrosis. Many studies on cell death and programmed cell death have been published and most are, at least to some degree, related to cancer. Some key proteins and molecular pathways implicated in cell death have been analyzed, whereas others are still being actively researched; therefore, an increasing number of cellular compartments and organelles are being implicated in cell death and cancer. Here, we discuss the mitochondria and subdomains of the endoplasmic reticulum (ER) that interact with mitochondria, the mitochondria-associated membranes (MAMs), which have been identified as critical hubs in the regulation of cell death and tumor growth. MAMs-dependent calcium (Ca 2+ ) release from the ER allows selective Ca 2+ uptake by the mitochondria. The perturbation of Ca 2+ homeostasis in cancer cells is correlated with sustained cell proliferation and the inhibition of cell death through the modulation of Ca 2+ signaling. This article is part of a Special Issue entitled Mitochondria in Cancer, edited by Giuseppe Gasparre, Rodrigue Rossignol and Pierre Sonveaux. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Hemorrhagic shock impairs myocardial cell volume regulation and membrane integrity in dogs

    International Nuclear Information System (INIS)

    Horton, J.W.


    An in vitro myocardial slice technique was used to quantitate alterations in cell volume regulation and membrane integrity after 2 h or hemorrhagic shock. After in vitro incubation in Krebs-Ringer-phosphate medium containing trace [ 14 C]inulin, values (ml H 2 O/g dry wt) for control nonshocked myocardial slices were 4.03 /plus minus/ 0.11 (SE) for total water, 2.16 /plus minus/ 0.07 for inulin impermeable space, and 1.76 /plus minus/ 0.15 for inulin diffusible space. Shocked myocardial slices showed impaired response to cold incubation. After 2 h of in vivo shock, total tissue water, inulin diffusible space, and inulin impermeable space increased significantly for subendocardium, whereas changes in subepicardium parameters were minimal. Shock-induced cellular swelling was accompanied by an increased total tissue sodium, but no change in tissue potassium. Calcium entry blockade in vivo significantly reduced subendocardial total tissue water as compared with shock-untreated dogs. In addition, calcium entry blockade reduced shock-induced increases in inulin diffusible space. In vitro myocardial slice studies confirm alterations in subendocardial membrane integrity after 2 h of in vivo hemorrhagic shock. Shock-induced abnormalities in myocardial cell volume regulation are reduced by calcium entry blockade in vivo

  13. Oncostatin M-stimulated apical plasma membrane biogenesis requires p27(Kip1)-regulated cell cycle dynamics

    NARCIS (Netherlands)

    Van IJzendoorn, Sven C D; Théard, Delphine; Van Der Wouden, Johanna M; Visser, Willy; Wojtal, Kacper A; Hoekstra, Dick

    Oncostatin M regulates membrane traffic and stimulates apicalization of the cell surface in hepatoma cells in a protein kinase A-dependent manner. Here, we show that oncostatin M enhances the expression of the cyclin-dependent kinase (cdk)2 inhibitor p27(Kip1), which inhibits G(1)-S-phase

  14. Membrane cholesterol regulates lysosome-plasma membrane fusion events and modulates Trypanosoma cruzi invasion of host cells.

    Directory of Open Access Journals (Sweden)

    Bárbara Hissa

    Full Text Available BACKGROUND: Trypomastigotes of Trypanosoma cruzi are able to invade several types of non-phagocytic cells through a lysosomal dependent mechanism. It has been shown that, during invasion, parasites trigger host cell lysosome exocytosis, which initially occurs at the parasite-host contact site. Acid sphingomyelinase released from lysosomes then induces endocytosis and parasite internalization. Lysosomes continue to fuse with the newly formed parasitophorous vacuole until the parasite is completely enclosed by lysosomal membrane, a process indispensable for a stable infection. Previous work has shown that host membrane cholesterol is also important for the T. cruzi invasion process in both professional (macrophages and non-professional (epithelial phagocytic cells. However, the mechanism by which cholesterol-enriched microdomains participate in this process has remained unclear. METHODOLOGY/PRINCIPAL FINDING: In the present work we show that cardiomyocytes treated with MβCD, a drug able to sequester cholesterol from cell membranes, leads to a 50% reduction in invasion by T. cruzi trypomastigotes, as well as a decrease in the number of recently internalized parasites co-localizing with lysosomal markers. Cholesterol depletion from host membranes was accompanied by a decrease in the labeling of host membrane lipid rafts, as well as excessive lysosome exocytic events during the earlier stages of treatment. Precocious lysosomal exocytosis in MβCD treated cells led to a change in lysosomal distribution, with a reduction in the number of these organelles at the cell periphery, and probably compromises the intracellular pool of lysosomes necessary for T. cruzi invasion. CONCLUSION/SIGNIFICANCE: Based on these results, we propose that cholesterol depletion leads to unregulated exocytic events, reducing lysosome availability at the cell cortex and consequently compromise T. cruzi entry into host cells. The results also suggest that two different pools of

  15. Actin filaments regulate the adhesion between the plasma membrane and the cell wall of tobacco guard cells. (United States)

    Yu, Qin; Ren, Jing-Jing; Kong, Lan-Jing; Wang, Xiu-Ling


    During the opening and closing of stomata, guard cells undergo rapid and reversible changes in their volume and shape, which affects the adhesion of the plasma membrane (PM) to the cell wall (CW). The dynamics of actin filaments in guard cells are involved in stomatal movement by regulating structural changes and intracellular signaling. However, it is unclear whether actin dynamics regulate the adhesion of the PM to the CW. In this study, we investigated the relationship between actin dynamics and PM-CW adhesion by the hyperosmotic-induced plasmolysis of tobacco guard cells. We found that actin filaments in guard cells were depolymerized during mannitol-induced plasmolysis. The inhibition of actin dynamics by treatment with latrunculin B or jasplakinolide and the disruption of the adhesion between the PM and the CW by treatment with RGDS peptide (Arg-Gly-Asp-Ser) enhanced guard cell plasmolysis. However, treatment with latrunculin B alleviated the RGDS peptide-induced plasmolysis and endocytosis. Our results reveal that the actin depolymerization is involved in the regulation of the PW-CW adhesion during hyperosmotic-induced plasmolysis in tobacco guard cells.

  16. Regulation of B cell differentiation by intracellular membrane associated proteins and microRNAs: role in the antibody response

    Directory of Open Access Journals (Sweden)

    Zheng eLou


    Full Text Available B cells are central to adaptive immunity and their functions in antibody responses are exquisitely regulated. As suggested by recent findings, B cell differentiation is mediated by intracellular membrane structures (including endosomes, lysosomes and autophagosomes and protein factors specifically associated with these membranes, including Rab7, Atg5 and Atg7. These factors participate in vesicle formation/trafficking, signal transduction and induction of gene expression to promote antigen presentation, CSR/SHM, and generation/maintenance of plasma cells and memory B cells. Their expression is induced in B cells activated to differentiate and further fine-tuned by immune-modulating microRNAs, which coordinates CSR/SHM, plasma cell differentiation and memory B cell differentiation. These short non-coding RNAs would individually target multiple factors associated with the same intracellular membrane compartments and collaboratively target a single factor in addition to regulate AID and Blimp-1. These, together with regulation of microRNA biogenesis and activities by endosomes and autophagosomes, show that intracellular membranes and microRNAs, two broadly relevant cell constituents, play important roles in balancing gene expression to specify B cell differentiation processes for optimal antibody responses.

  17. F-BAR family proteins, emerging regulators for cell membrane dynamic changes-from structure to human diseases. (United States)

    Liu, Suxuan; Xiong, Xinyu; Zhao, Xianxian; Yang, Xiaofeng; Wang, Hong


    Eukaryotic cell membrane dynamics change in curvature during physiological and pathological processes. In the past ten years, a novel protein family, Fes/CIP4 homology-Bin/Amphiphysin/Rvs (F-BAR) domain proteins, has been identified to be the most important coordinators in membrane curvature regulation. The F-BAR domain family is a member of the Bin/Amphiphysin/Rvs (BAR) domain superfamily that is associated with dynamic changes in cell membrane. However, the molecular basis in membrane structure regulation and the biological functions of F-BAR protein are unclear. The pathophysiological role of F-BAR protein is unknown. This review summarizes the current understanding of structure and function in the BAR domain superfamily, classifies F-BAR family proteins into nine subfamilies based on domain structure, and characterizes F-BAR protein structure, domain interaction, and functional relevance. In general, F-BAR protein binds to cell membrane via F-BAR domain association with membrane phospholipids and initiates membrane curvature and scission via Src homology-3 (SH3) domain interaction with its partner proteins. This process causes membrane dynamic changes and leads to seven important cellular biological functions, which include endocytosis, phagocytosis, filopodium, lamellipodium, cytokinesis, adhesion, and podosome formation, via distinct signaling pathways determined by specific domain-binding partners. These cellular functions play important roles in many physiological and pathophysiological processes. We further summarize F-BAR protein expression and mutation changes observed in various diseases and developmental disorders. Considering the structure feature and functional implication of F-BAR proteins, we anticipate that F-BAR proteins modulate physiological and pathophysiological processes via transferring extracellular materials, regulating cell trafficking and mobility, presenting antigens, mediating extracellular matrix degradation, and transmitting

  18. Wrapping of a deformable nanoparticle by the cell membrane: Insights into the flexibility-regulated nanoparticle-membrane interaction (United States)

    Tang, Huayuan; Zhang, Hongwu; Ye, Hongfei; Zheng, Yonggang


    Although many researches have been conducted on the interaction of the cell membrane with the rigid nanoparticle (NP), relatively little is known about the interaction of the membrane with the deformable NP, which is a promising kind of drug delivery carrier. In this paper, we investigate the wrapping of a deformable NP by the membrane, with particular attention paid to the location of the NP. Phase diagrams with respect to the normalized NP-membrane adhesion strength and the bending stiffness ratio between the NP and membrane are presented. The results show that the NP is easier to be fully wrapped but harder to be shallowly wrapped when the NP locates outside than inside the vesicle. For the system with an outside NP, there are three distinct stages separated by two critical bending stiffness ratios as the NP becomes softer. Moreover, the critical normalized adhesion strength required for a deformable NP to be fully wrapped is the same as that for a rigid NP when the bending stiffness ratio is higher than a critical value, which is different from the wrapping behavior by an initially flat membrane. In addition, a larger vesicle size facilitates the full wrapping configuration when the NP is inside, whereas it prohibits it when the NP is outside. These results are consistent with the previous research and can provide guidelines for the design of drug delivery systems based on the flexibility-tunable NPs.

  19. Membrane potential of cells and its regulation during aging. 2. Report: the effect of hormones on the level of the cellular plasma membrane polarization. (United States)

    Frolkis, V V; Tanin, S A; Gorban, E N; Bogatskaya, L N; Sabko, V E


    Age-dependent changes in the polarization of plasma membranes (PM) of various cell types and the mechanisms responsible for its regulation were studied in the experiments on the adult (6-8 and old (28-32 months) Wistar male rats. It was found that the effect of the hormones on the PM polarization level is altered during aging. This being related to shifts in the number and affinity of the hormonal receptors, energetic processes and protein synthesis in the cell.

  20. Identification of a novel role for dematin in regulating red cell membrane function by modulating spectrin-actin interaction. (United States)

    Koshino, Ichiro; Mohandas, Narla; Takakuwa, Yuichi


    The membrane skeleton plays a central role in maintaining the elasticity and stability of the erythrocyte membrane, two biophysical features critical for optimal functioning and survival of red cells. Many constituent proteins of the membrane skeleton are phosphorylated by various kinases, and phosphorylation of β-spectrin by casein kinase and of protein 4.1R by PKC has been documented to modulate erythrocyte membrane mechanical stability. In this study, we show that activation of endogenous PKA by cAMP decreases membrane mechanical stability and that this effect is mediated primarily by phosphorylation of dematin. Co-sedimentation assay showed that dematin facilitated interaction between spectrin and F-actin, and phosphorylation of dematin by PKA markedly diminished this activity. Quartz crystal microbalance measurement revealed that purified dematin specifically bound the tail region of the spectrin dimer in a saturable manner with a submicromolar affinity. Pulldown assay using recombinant spectrin fragments showed that dematin, but not phospho-dematin, bound to the tail region of the spectrin dimer. These findings imply that dematin contributes to the maintenance of erythrocyte membrane mechanical stability by facilitating spectrin-actin interaction and that phosphorylation of dematin by PKA can modulate these effects. In this study, we have uncovered a novel functional role for dematin in regulating erythrocyte membrane function.

  1. Identification of a Novel Role for Dematin in Regulating Red Cell Membrane Function by Modulating Spectrin-Actin Interaction* (United States)

    Koshino, Ichiro; Mohandas, Narla; Takakuwa, Yuichi


    The membrane skeleton plays a central role in maintaining the elasticity and stability of the erythrocyte membrane, two biophysical features critical for optimal functioning and survival of red cells. Many constituent proteins of the membrane skeleton are phosphorylated by various kinases, and phosphorylation of β-spectrin by casein kinase and of protein 4.1R by PKC has been documented to modulate erythrocyte membrane mechanical stability. In this study, we show that activation of endogenous PKA by cAMP decreases membrane mechanical stability and that this effect is mediated primarily by phosphorylation of dematin. Co-sedimentation assay showed that dematin facilitated interaction between spectrin and F-actin, and phosphorylation of dematin by PKA markedly diminished this activity. Quartz crystal microbalance measurement revealed that purified dematin specifically bound the tail region of the spectrin dimer in a saturable manner with a submicromolar affinity. Pulldown assay using recombinant spectrin fragments showed that dematin, but not phospho-dematin, bound to the tail region of the spectrin dimer. These findings imply that dematin contributes to the maintenance of erythrocyte membrane mechanical stability by facilitating spectrin-actin interaction and that phosphorylation of dematin by PKA can modulate these effects. In this study, we have uncovered a novel functional role for dematin in regulating erythrocyte membrane function. PMID:22927433

  2. Myosin Vb mediated plasma membrane homeostasis regulates peridermal cell size and maintains tissue homeostasis in the zebrafish epidermis. (United States)

    Sonal; Sidhaye, Jaydeep; Phatak, Mandar; Banerjee, Shamik; Mulay, Aditya; Deshpande, Ojas; Bhide, Sourabh; Jacob, Tressa; Gehring, Ines; Nuesslein-Volhard, Christiane; Sonawane, Mahendra


    The epidermis is a stratified epithelium, which forms a barrier to maintain the internal milieu in metazoans. Being the outermost tissue, growth of the epidermis has to be strictly coordinated with the growth of the embryo. The key parameters that determine tissue growth are cell number and cell size. So far, it has remained unclear how the size of epidermal cells is maintained and whether it contributes towards epidermal homeostasis. We have used genetic analysis in combination with cellular imaging to show that zebrafish goosepimples/myosin Vb regulates plasma membrane homeostasis and is involved in maintenance of cell size in the periderm, the outermost epidermal layer. The decrease in peridermal cell size in Myosin Vb deficient embryos is compensated by an increase in cell number whereas decrease in cell number results in the expansion of peridermal cells, which requires myosin Vb (myoVb) function. Inhibition of cell proliferation as well as cell size expansion results in increased lethality in larval stages suggesting that this two-way compensatory mechanism is essential for growing larvae. Our analyses unravel the importance of Myosin Vb dependent cell size regulation in epidermal homeostasis and demonstrate that the epidermis has the ability to maintain a dynamic balance between cell size and cell number.

  3. Regulation of CD4+ T-Cell Function by Membrane Cholesterol (United States)


    pathogen immune evasion or possibly when a robust CD4 T-cell response is desired in response to vaccination. A potential target for regulating T-cell...Carcinogenesis 1998;19: 287-290. Rouse BT and Sehrawat S. Immunity and immunopathology to viruses : what decided the outcome? Nat Rev Immunol 2010;10:514...the Health Sciences 4301 Jones Bridge Road, Bethesda, Maryland 20814 The CD4 T helper cells play a central role in initiating immune

  4. Mg(2+,ATP-dependent plasma membrane calcium pump of smooth muscle cells. ІІ. Regulation of activity

    Directory of Open Access Journals (Sweden)

    T. О. Veklich


    Full Text Available Plasma membrane Ca2+-pump is one of key proteins, which takes part in Ca2+ exchange in smooth muscle cells. It has a lot of diverse functions from control of basal cytoplasmal Ca2+ concentration to regulation of proteins involved in Ca2+-dependent signal pathway. Ca2+ pump function is often depen­dent on the isoform or even form of alternative splicing. Allowing for a variety of Ca2+-pump functions and properties, which were reviewed in detail in the first part of our review article cycle (Ukr. Biochem. J., 2015; 87(1, the precise control of the mentioned pump activity is very important for cell functioning­. The other part of this article is dedicated to different regulation factors of smooth muscle plasma membrane Ca2+-pump activity: endogenous and exo­genous, biotic and abiotic factors. Special attention is given to literature data and own results about design and the search of selective plasma membrane Ca2+-pump inhibitor which would allow examining its functioning in smooth muscle cells more meticulously.

  5. The influence of growth regulators on membrane permeability in cultures of winter wheat cells

    Czech Academy of Sciences Publication Activity Database

    Filek, M.; KoĽcielniak, J.; Marcińska, I.; Macháčková, Ivana; Krekule, Jan


    Roč. 59, 9/10 (2004), s. 673-678 ISSN 0939-5075 R&D Projects: GA AV ČR IBS5038352 Institutional research plan: CEZ:AV0Z5038910 Keywords : growth regulators * cells * permeability Subject RIV: GC - Agronomy Impact factor: 0.715, year: 2004

  6. Membrane Microdomains and Cytoskeleton Organization Shape and Regulate the IL-7 Receptor Signalosome in Human CD4 T-cells* (United States)

    Tamarit, Blanche; Bugault, Florence; Pillet, Anne-Hélène; Lavergne, Vincent; Bochet, Pascal; Garin, Nathalie; Schwarz, Ulf; Thèze, Jacques; Rose, Thierry


    Interleukin (IL)-7 is the main homeostatic regulator of CD4 T-lymphocytes (helper) at both central and peripheral levels. Upon activation by IL-7, several signaling pathways, mainly JAK/STAT, PI3K/Akt and MAPK, induce the expression of genes involved in T-cell differentiation, activation, and proliferation. We have analyzed the early events of CD4 T-cell activation by IL-7. We have shown that IL-7 in the first few min induces the formation of cholesterol-enriched membrane microdomains that compartmentalize its activated receptor and initiate its anchoring to the cytoskeleton, supporting the formation of the signaling complex, the signalosome, on the IL-7 receptor cytoplasmic domains. Here we describe by stimulated emission depletion microscopy the key roles played by membrane microdomains and cytoskeleton transient organization in the IL-7-regulated JAK/STAT signaling pathway. We image phospho-STAT5 and cytoskeleton components along IL-7 activation kinetics using appropriate inhibitors. We show that lipid raft inhibitors delay and reduce IL-7-induced JAK1 and JAK3 phosphorylation. Drug-induced disassembly of the cytoskeleton inhibits phospho-STAT5 formation, transport, and translocation into the nucleus that controls the transcription of genes involved in T-cell activation and proliferation. We fit together the results of these quantitative analyses and propose the following mechanism. Activated IL-7 receptors embedded in membrane microdomains induce actin-microfilament meshwork formation, anchoring microtubules that grow radially from rafted receptors to the nuclear membrane. STAT5 phosphorylated by signalosomes are loaded on kinesins and glide along the microtubules across the cytoplasm to reach the nucleus 2 min after IL-7 stimulation. Radial microtubules disappear 15 min later, while transversal microtubules, independent of phospho-STAT5 transport, begin to bud from the microtubule organization center. PMID:23329834

  7. Podoplanin, novel 43-kd membrane protein of glomerular epithelial cells, is down-regulated in puromycin nephrosis. (United States)

    Breiteneder-Geleff, S.; Matsui, K.; Soleiman, A.; Meraner, P.; Poczewski, H.; Kalt, R.; Schaffner, G.; Kerjaschki, D.


    Puromycin aminonucleoside nephrosis (PAN), a rat model of human minimal change nephropathy, is characterized by extensive flattening of glomerular epithelial cell (podocyte) foot processes and by severe proteinuria. For comparison of expression of glomerular membrane proteins of normal and PAN rats, a membrane protein fraction of isolated rat glomeruli was prepared and monoclonal antibodies were raised against it. An IgG-secreting clone designated LF3 was selected that specifically immunolabeled podocytes of normal but not of PAN rats. The antigen of LF3 IgG was identified as a 43-kd glycoprotein. Molecular cloning of its cDNA was performed in a delta gt11 expression library prepared from mRNA of isolated rat glomeruli. The predicted amino acid sequence indicated a 166-amino-acid integral membrane protein with a single membrane-spanning domain, two potential phosphorylation sites in its short cytoplasmic tail, and six potential O-glycosylation sites in the large ectodomain. High amino acid sequence identities were found to membrane glycoproteins of rat lung and bone and mouse thymus epithelial cells as well as to a phorbol-ester-induced protein in a mouse osteoblast cell line and to a canine influenza C virus receptor. In PAN, expression of this 43-kd protein was selectively reduced to < 30%, as determined by quantitative immunogold electron microscopy, immunoblotting, and Northern blotting. These data provide evidence that transcription of the 43-kd transmembrane podocyte glycoprotein is specifically down-regulated in PAN. To indicate that this protein could be associated with transformation of arborized foot processes to flat feet (Latin, pes planus) we have called it podoplanin. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 10 Figure 12 Figure 13 Figure 14 Figure 15 PMID:9327748

  8. Ion channel regulation of the dynamical instability of the resting membrane potential in saccular hair cells of the green frog (Rana esculenta)

    NARCIS (Netherlands)

    Jorgensen, F; Kroese, ABA


    Aims: We investigated the ion channel regulation of the resting membrane potential of hair cells with the aim to determine if the resting membrane potential is poised close to instability and thereby a potential cause of the spontaneous afferent spike activity. Methods: The ionic mechanism and the

  9. Ouabain sensitive Na+/K+-pump regulates other membrane transporters in the microdomain of smooth muscle cells

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Nilsson, Holger; Aalkjær, Christian

    Ouabain, a specific inhibitor of the Na+/K+-pump, has previously been shown to interfere with intercellular communication. We have demonstrated a mechanism of this action of ouabain (1). We have showed that gap junctions between vascular smooth muscle cells (SMCs) are regulated through...... an interaction between the Na+/K+-pump and the Na+/Ca2+-exchanger leading to an increase in the intracellular calcium concentration  ([Ca2+]i) in discrete areas near the plasma membrane. This regulation suggests a close association of the proteins in microdomains. We have also suggested that this Na......+/K+-pump-containing microdomain is functionally linked to KATP channels via the local ion homeostasis and that this interaction can be bidirectional (1;2). [Ca2+]i in individual SMCs was imaged simultaneously with isometric force in rat mesenteric small arteries. Paired cultured rat aortic smooth muscle cells (A7r5) were used...

  10. Decidual-secreted factors alter invasive trophoblast membrane and secreted proteins implying a role for decidual cell regulation of placentation.

    Directory of Open Access Journals (Sweden)

    Ellen Melaleuca Menkhorst

    Full Text Available Inadequate or inappropriate implantation and placentation during the establishment of human pregnancy is thought to lead to first trimester miscarriage, placental insufficiency and other obstetric complications. To create the placental blood supply, specialized cells, the 'extravillous trophoblast' (EVT invade through the differentiated uterine endometrium (the decidua to engraft and remodel uterine spiral arteries. We hypothesized that decidual factors would regulate EVT function by altering the production of EVT membrane and secreted factors. We used a proteomics approach to identify EVT membrane and secreted proteins regulated by decidual cell factors. Human endometrial stromal cells were decidualized in vitro by treatment with estradiol (10(-8 M, medroxyprogesterone acetate (10(-7 M and cAMP (0.5 mM for 14 days. Conditioned media (CM was collected on day 2 (non-decidualized CM and 14 (decidualized CM of treatment. Isolated primary EVT cultured on Matrigel™ were treated with media control, non-decidualized or decidualized CM for 16 h. EVT CM was fractionated for proteins <30 kDa using size-exclusion affinity nanoparticles (SEAN before trypsin digestion and HPLC-MS/MS. 43 proteins produced by EVT were identified; 14 not previously known to be expressed in the placenta and 12 which had previously been associated with diseases of pregnancy including preeclampsia. Profilin 1, lysosome associated membrane glycoprotein 1 (LAMP1, dipeptidyl peptidase 1 (DPP1/cathepsin C and annexin A2 expression by interstitial EVT in vivo was validated by immunhistochemistry. Decidual CM regulation in vitro was validated by western blotting: decidualized CM upregulated profilin 1 in EVT CM and non-decidualized CM upregulated annexin A2 in EVT CM and pro-DPP1 in EVT cell lysate. Here, non-decidualized factors induced protease expression by EVT suggesting that non-decidualized factors may induce a pro-inflammatory cascade. Preeclampsia is a pro

  11. Membrane potential of cells and its regulation during aging. 1. Report: the role of energetic metabolism and plasma membrane phospholipid contents. (United States)

    Frolkis, V V; Tanin, S A; Gorban, E N; Bogatskaya, L N; Sabko, V E


    Age-dependent changes in the polarization of the plasma membranes (PM) of various cell types and the mechanisms responsible for its regulation were studied in the experiments on the adult (6-8 months) and old (28-32 months) Wistar male rats. The relationship was found between the specificity of cellular function and the pattern of changes in resting potential (RP) and action potential (AP). In senility, changes occur in the ratio of different pathways of energy provision of the ionic transport: the contribution made by enzymic reactions to the mechanism of maintenance of the PM polarization level is diminished. Changes in the PM phospholipid composition influences considerably the character of the PM electric reactions in aging. An anti-oxidant, dibunol (butilated hydroxytoluene, BHT) has appeared to restore many reactions of the cellular PM in the old animals.

  12. Cooperation of decay-accelerating factor and membrane cofactor protein in regulating survival of human cervical cancer cells

    International Nuclear Information System (INIS)

    Gao, Ling-Juan; Guo, Shu-Yu; Cai, You-Qun; Gu, Ping-Qing; Su, Ya-Juan; Gong, Hui; Liu, Yun; Chen, Chen


    Decay-accelerating factor (DAF) and membrane cofactor protein (MCP) are the key molecules involved in cell protection against autologus complement, which restricts the action of complement at critical stages of the cascade reaction. The cooperative effect of DAF and MCP on the survival of human cervical cancer cell (ME180) has not been demonstrated. In this study we applied, for the first time, short hairpin RNA (shRNA) to knock down the expression of the DAF and MCP with the aim of exploiting complement more effectively for tumor cell damage. Meanwhile, we investigated the cooperative effects of DAF and MCP on the viability and migration, moreover the proliferation of ME180 cell. The results showed that shRNA inhibition of DAF and MCP expression enhanced complement-dependent cytolysis (CDC) up to 39% for MCP and up to 36% for DAF, and the combined inhibition of both regulators yielded further additive effects in ME180 cells. Thus, the activities of DAF and MCP, when present together, are greater than the sum of the two protein individually. These data indicated that combined DAF and MCP shRNA described in this study may offer an additional alternative to improve the efficacy of antibody-and complement-based cancer immunotherapy

  13. Membrane Cells for Brine Electrolysis. (United States)

    Tingle, M.


    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)

  14. Deciphering Cell-to-Cell Communication in Acquisition of Cancer Traits: Extracellular Membrane Vesicles Are Regulators of Tissue Biomechanics. (United States)

    Pokharel, Deep; Wijesinghe, Philip; Oenarto, Vici; Lu, Jamie F; Sampson, David D; Kennedy, Brendan F; Wallace, Vincent P; Bebawy, Mary


    Deciphering the role of cell-to-cell communication in acquisition of cancer traits such as metastasis is one of the key challenges of integrative biology and clinical oncology. In this context, extracellular vesicles (EVs) are important vectors in cell-to-cell communication and serve as conduits in the transfer of cellular constituents required for cell function and for the establishment of cellular phenotypes. In the case of malignancy, they have been shown to support the acquisition of common traits defined as constituting the hallmarks of cancer. Cellular biophysics has contributed to our understanding of some of these central traits with changes in tissue biomechanics reflective of cell state. Indeed, much is known about stiffness of the tissue scaffold in the context of cell invasion and migration. This article advances this knowledge frontier by showing for the first time that EVs are mediators of tissue biomechanical properties and, importantly, demonstrates a link between the acquisition of cancer multidrug resistance and increased tissue stiffness of the malignant mass. The methodology used in the study employed optical coherence elastography and atomic force microscopy on breast cancer cell monolayers and tumor spheroids. Specifically, we show here that the acquired changes in tissue stiffness can be attributed to the intracellular transfer of a protein complex comprising ezrin, radixin, moesin, CD44, and P-glycoprotein. This has important implications in facilitating mechano-transduced signaling cascades that regulate the acquisition of cancer traits, such as invasion and metastasis. Finally, this study also introduces novel targets and strategies for diagnostic and therapeutic innovation in oncology, with a view to prevention of metastatic spread and personalized medicine in cancer treatment.

  15. Fuel cell membrane humidification (United States)

    Wilson, Mahlon S.


    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.

  16. Cell Membrane Coating Nanotechnology. (United States)

    Fang, Ronnie H; Kroll, Ashley V; Gao, Weiwei; Zhang, Liangfang


    Nanoparticle-based therapeutic, prevention, and detection modalities have the potential to greatly impact how diseases are diagnosed and managed in the clinic. With the wide range of nanomaterials available, the rational design of nanocarriers on an application-specific basis has become increasingly commonplace. Here, a comprehensive overview is provided on an emerging platform: cell-membrane-coating nanotechnology. As a fundamental unit of biology, cells carry out a wide range of functions, including the remarkable ability to interface and interact with their surrounding environment. Instead of attempting to replicate such functions via synthetic techniques, researchers are now directly leveraging naturally derived cell membranes as a means of bestowing nanoparticles with enhanced biointerfacing capabilities. This top-down technique is facile, highly generalizable, and has the potential to greatly augment existing nanocarriers. Further, the introduction of a natural membrane substrate onto nanoparticles surfaces has enabled additional applications beyond those traditionally associated with nanomedicine. Despite its relative youth, there exists an impressive body of literature on cell membrane coating, which is covered here in detail. Overall, there is still significant room for development, as researchers continue to refine existing workflows while finding new and exciting applications that can take advantage of this developing technology. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Vaginal epithelial cells regulate membrane adhesiveness to co-ordinate bacterial adhesion

    NARCIS (Netherlands)

    Younes, Jessica A.; Klappe, Karin; Kok, Jan Willem; Busscher, Henk J.; Reid, Gregor; van der Mei, Henny C.

    Vaginal epithelium is colonized by different bacterial strains and species. The bacterial composition of vaginal biofilms controls the balance between health and disease. Little is known about the relative contribution of the epithelial and bacterial cell surfaces to bacterial adhesion and whether

  18. A role for protein kinase C in the regulation of membrane fluidity and Ca²(+) flux at the endoplasmic reticulum and plasma membranes of HEK293 and Jurkat cells. (United States)

    Chen, Lihong; Meng, Qingli; Jing, Xian; Xu, Pingxiang; Luo, Dali


    Protein kinase C (PKC) plays a prominent role in the regulation of a variety of cellular functions, including Ca²(+) signalling. In HEK293 and Jurkat cells, the Ca²(+) release and Ca²(+) uptake stimulated by several different activators were attenuated by activation of PKC with phorbol myristate acetate (PMA) or 1-oleoyl-2-acetyl-sn-glycerol (OAG) and potentiated by PKC inhibition with Gö6983 or knockdown of PKCα or PKCβ using shRNA. Immunostaining and Western blotting analyses revealed that PKCα and PKCβII accumulated at the plasma membrane (PM) and that these isoforms, along with PKCβI, also translocated to the endoplasmic reticulum (ER) upon activation with PMA. Measurements of membrane fluidity showed that, like the cell membrane stabilizers bovine serum albumin (BSA) and ursodeoxycholate (UDCA), PMA and OAG significantly reduced the fluidity of both the PM and ER membranes; these effects were blocked in PKC-knockdown cells. Interestingly, both BSA and UDCA inhibited the Ca²(+) responses to agonists to the same extent as PMA, whereas Tween 20, which increases membrane fluidity, raised the internal Ca²(+) concentration. Thus, activation of PKC induces both translocation of PKC to the PM and ER membranes and downregulation of membrane fluidity, thereby negatively modulating Ca²(+) flux. Copyright © 2010 Elsevier Inc. All rights reserved.

  19. Membrane-Type 1 Matrix Metal loproteinase Is Regulated by Sp1 through the Differential Activation of AKT, JNK, and ERK Pathways in Human Prostate Tumor Cells

    Directory of Open Access Journals (Sweden)

    Isis C. Sroka


    Full Text Available We and other investigators have previously shown that membrane-type 1 matrix metalloproteinase (MT1-MMP is overexpressed in invasive prostate cancer cells. However, the mechanism for this expression is not known. Here, we show that MT1-MMP is minimally expressed in nonmalignant primary prostate cells, moderately expressed in DU-145 cells, and highly expressed in invasive PC-3 and PC-3N cells. Using human MT1-MMP promoter reporter plasmids and mobility shift assays, we show that Spi regulates MT1-MMP expression in DU-145, PC-3, and PC-3N cells and in PC3-N cells using chromatin immunoprecipitation analysis and silencing RNA. Investigation of signaling pathway showed that DU-145 cells express constitutively phosphorylated extracellular stress-regulated kinase (ERK, whereas PC-3 and PC-3N cells express constitutively phosphorylated AKT/PKB and c-Jun NH2 terminal kinase (JNK. We show that MT1-MMP and Spi levels are decreased in PC-3 and PC-3N cells when phosphatidylinositol-3 kinase and JNK are inhibited, and that MT1-MMP levels are decreased in DU-145 cells when MEK is inhibited. Transient transfection of PC-3 and PC-3N cells with a dominant-negative JNK or p85, and of DU-145 cells with a dominant negative ERK, reduces MT1-MMP promoter activity. These results indicate differential signaling control of Spi-mediated transcriptional regulation of MT1-MMP in prostate cancer cell lines.

  20. Role of the Na+/K+-ATPase in regulating the membrane potential in rat peritoneal mast cells

    DEFF Research Database (Denmark)

    Friis, U G; Praetorius, Birger Hans; Knudsen, T


    of Sylgard-coated patch pipettes (3-6 M[omega]). High-resolution membrane currents were recorded with an EPC-9 patch-clamp amplifier controlled by the 'E9SCREEN' software. In addition, a charting programme on another computer synchronously recorded at low resolution (2 Hz) membrane potential and holding...... current (low-pass filtered at 500 Hz). 3. Na+/K+-ATPase activity was measured as the ouabain-sensitive change in the zero-current potential. The zero-current potential in rat peritoneal mast cells measured 2 min after obtaining whole-cell configuration amounted to 1.7 +/- 2.5 mV (n = 21). Ouabain (5 m...... preincubated in nominal calcium-free external solution for 12 +/- 1.6 min before whole-cell configuration, the membrane potential amounted to -53.7 +/- 9.8 mV (n = 8). A subsequent superfusion with ouabain (5 mM) depolarized the membrane potential (ouabain-sensitive hyperpolarization (delta mV): 23.0 +/- 8.4 m...

  1. Proton exchange membrane fuel cells

    CERN Document Server

    Qi, Zhigang


    Preface Proton Exchange Membrane Fuel CellsFuel CellsTypes of Fuel CellsAdvantages of Fuel CellsProton Exchange Membrane Fuel CellsMembraneCatalystCatalyst LayerGas Diffusion MediumMicroporous LayerMembrane Electrode AssemblyPlateSingle CellStackSystemCell Voltage Monitoring Module (CVM)Fuel Supply Module (FSM)Air Supply Module (ASM)Exhaust Management Module (EMM)Heat Management Module (HMM)Water Management Module (WMM)Internal Power Supply Module (IPM)Power Conditioning Module (PCM)Communications Module (COM)Controls Module (CM)SummaryThermodynamics and KineticsTheoretical EfficiencyVoltagePo

  2. Dissecting the role of conformational change and membrane binding by the bacterial cell division regulator MinE in the stimulation of MinD ATPase activity. (United States)

    Ayed, Saud H; Cloutier, Adam D; McLeod, Laura J; Foo, Alexander C Y; Damry, Adam M; Goto, Natalie K


    The bacterial cell division regulators MinD and MinE together with the division inhibitor MinC localize to the membrane in concentrated zones undergoing coordinated pole-to-pole oscillation to help ensure that the cytokinetic division septum forms only at the mid-cell position. This dynamic localization is driven by MinD-catalyzed ATP hydrolysis, stimulated by interactions with MinE's anti-MinCD domain. This domain is buried in the 6-β-stranded MinE "closed" structure, but is liberated for interactions with MinD, giving rise to a 4-β-stranded "open" structure through an unknown mechanism. Here we show that MinE-membrane interactions induce a structural change into a state resembling the open conformation. However, MinE mutants lacking the MinE membrane-targeting sequence stimulated higher ATP hydrolysis rates than the full-length protein, indicating that binding to MinD is sufficient to trigger this conformational transition in MinE. In contrast, conformational change between the open and closed states did not affect stimulation of ATP hydrolysis rates in the absence of membrane binding, although the MinD-binding residue Ile-25 is critical for this conformational transition. We therefore propose an updated model where MinE is brought to the membrane through interactions with MinD. After stimulation of ATP hydrolysis, MinE remains bound to the membrane in a state that does not catalyze additional rounds of ATP hydrolysis. Although the molecular basis for this inhibited state is unknown, previous observations of higher-order MinE self-association may explain this inhibition. Overall, our findings have general implications for Min protein oscillation cycles, including those that regulate cell division in bacterial pathogens. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Efficient Overproduction of Membrane Proteins in Lactococcus lactis Requires the Cell Envelope Stress Sensor/Regulator Couple CesSR (United States)

    Pinto, Joao P. C.; Kuipers, Oscar P.; Marreddy, Ravi K. R.; Poolman, Bert; Kok, Jan


    Background Membrane proteins comprise an important class of molecules whose study is largely frustrated by several intrinsic constraints, such as their hydrophobicity and added requirements for correct folding. Additionally, the complexity of the cellular mechanisms that are required to insert membrane proteins functionally in the membrane and to monitor their folding state makes it difficult to foresee the yields at which one can obtain them or to predict which would be the optimal production host for a given protein. Methods and Findings We describe a rational design approach to improve the lactic acid bacterium Lactococcus lactis as a producer of membrane proteins. Our transcriptome data shows that the two-component system CesSR, which senses cell envelope stresses of different origins, is one of the major players when L. lactis is forced to overproduce the endogenous membrane protein BcaP, a branched-chain amino acid permease. Growth of the BcaP-producing L. lactis strain and its capability to produce membrane proteins are severely hampered when the CesSR system itself or particular members of the CesSR regulon are knocked out, notably the genes ftsH, oxaA2, llmg_2163 and rmaB. Overexpressing cesSR reduced the growth defect, thus directly improving the production yield of BcaP. Applying this rationale to eukaryotic proteins, some of which are notoriously more difficult to produce, such as the medically-important presenilin complex, we were able to significantly diminish the growth defect seen in the wild-type strain and improve the production yield of the presenilin variant PS1Δ9-H6 more than 4-fold. Conclusions The results shed light into a key, and perhaps central, membrane protein quality control mechanism in L. lactis. Modulating the expression of CesSR benefited the production yields of membrane proteins from different origins. These findings reinforce L. lactis as a legitimate alternative host for the production of membrane proteins. PMID:21818275

  4. Plasma membrane Ca2+-ATPase isoforms composition regulates cellular pH homeostasis in differentiating PC12 cells in a manner dependent on cytosolic Ca2+ elevations

    DEFF Research Database (Denmark)

    Boczek, Tomasz; Lisek, Malwina; Ferenc, Bozena


    cellular acidification during KCl-stimulated Ca2+ influx. Because SERCA and NCX modulated cellular pH response in neglectable manner, and all conditions used to inhibit PMCA prevented KCl-induced pH drop, we considered PMCA2 and PMCA3 as mainly responsible for transport of protons to intracellular milieu......Plasma membrane Ca2+-ATPase (PMCA) by extruding Ca2+ outside the cell, actively participates in the regulation of intracellular Ca2+ concentration. Acting as Ca2+/H+ counter-transporter, PMCA transports large quantities of protons which may affect organellar pH homeostasis. PMCA exists in four......+-driven opening of mitochondrial permeability transition pore as putative underlying mechanism. The findings presented here demonstrate a crucial role of PMCA2 and PMCA3 in regulation of cellular pH and indicate PMCA membrane composition important for preservation of electrochemical gradient...

  5. Plasma membrane changes during programmed cell deaths. (United States)

    Zhang, Yingying; Chen, Xin; Gueydan, Cyril; Han, Jiahuai


    Ruptured and intact plasma membranes are classically considered as hallmarks of necrotic and apoptotic cell death, respectively. As such, apoptosis is usually considered a non-inflammatory process while necrosis triggers inflammation. Recent studies on necroptosis and pyroptosis, two types of programmed necrosis, revealed that plasma membrane rupture is mediated by MLKL channels during necroptosis but depends on non-selective gasdermin D (GSDMD) pores during pyroptosis. Importantly, the morphology of dying cells executed by MLKL channels can be distinguished from that executed by GSDMD pores. Interestingly, it was found recently that secondary necrosis of apoptotic cells, a previously believed non-regulated form of cell lysis that occurs after apoptosis, can be programmed and executed by plasma membrane pore formation like that of pyroptosis. In addition, pyroptosis is associated with pyroptotic bodies, which have some similarities to apoptotic bodies. Therefore, different cell death programs induce distinctive reshuffling processes of the plasma membrane. Given the fact that the nature of released intracellular contents plays a crucial role in dying/dead cell-induced immunogenicity, not only membrane rupture or integrity but also the nature of plasma membrane breakdown would determine the fate of a cell as well as its ability to elicit an immune response. In this review, we will discuss recent advances in the field of apoptosis, necroptosis and pyroptosis, with an emphasis on the mechanisms underlying plasma membrane changes observed on dying cells and their implication in cell death-elicited immunogenicity.

  6. Cell invasion through basement membrane


    Morrissey, Meghan A; Hagedorn, Elliott J; Sherwood, David R


    Cell invasion through basement membrane is an essential part of normal development and physiology, and occurs during the pathological progression of human inflammatory diseases and cancer. F-actin-rich membrane protrusions, called invadopodia, have been hypothesized to be the “drill bits” of invasive cells, mediating invasion through the dense, highly cross-linked basement membrane matrix. Though studied in vitro for over 30 y, invadopodia function in vivo has remained elusive. We have recent...

  7. Model cell membranes

    DEFF Research Database (Denmark)

    Günther-Pomorski, Thomas; Nylander, Tommy; Cardenas Gomez, Marite


    The high complexity of biological membranes has motivated the development and application of a wide range of model membrane systems to study biochemical and biophysical aspects of membranes in situ under well defined conditions. The aim is to provide fundamental understanding of processes control...


    DEFF Research Database (Denmark)


    A method for preparing polybenzimidazole or polybenzimidazole blend membranes and fabricating gas diffusion electrodes and membrane-electrode assemblies is provided for a high temperature polymer electrolyte membrane fuel cell. Blend polymer electrolyte membranes based on PBI and various thermopl......A method for preparing polybenzimidazole or polybenzimidazole blend membranes and fabricating gas diffusion electrodes and membrane-electrode assemblies is provided for a high temperature polymer electrolyte membrane fuel cell. Blend polymer electrolyte membranes based on PBI and various...... thermoplastic polymers for high temperature polymer electrolyte fuel cells have also been developed. Miscible blends are used for solution casting of polymer membranes (solid electrolytes). High conductivity and enhanced mechanical strength were obtained for the blend polymer solid electrolytes...... electrolyte membrane by hot-press. The fuel cell can operate at temperatures up to at least 200 °C with hydrogen-rich fuel containing high ratios of carbon monoxide such as 3 vol% carbon monoxide or more, compared to the carbon monoxide tolerance of 10-20 ppm level for Nafion$m(3)-based polymer electrolyte...

  9. APC/β-catenin-rich complexes at membrane protrusions regulate mammary tumor cell migration and mesenchymal morphology

    International Nuclear Information System (INIS)

    Odenwald, Matthew A; Prosperi, Jenifer R; Goss, Kathleen H


    The APC tumor suppressor is mutated or downregulated in many tumor types, and is prominently localized to punctate clusters at protrusion tips in migratory cells, such as in astrocytes where it has been implicated in directed cell motility. Although APC loss is considered an initiating event in colorectal cancer, for example, it is less clear what role APC plays in tumor cell motility and whether loss of APC might be an important promoter of tumor progression in addition to initiation. The localization of APC and β-catenin was analyzed in multiple cell lines, including non-transformed epithelial lines treated with a proteasome inhibitor or TGFβ to induce an epithelial-to-mesenchymal transition (EMT), as well as several breast cancer lines, by immunofluorescence. APC expression was knocked down in 4T07 mammary tumor cells using lentiviral-mediated delivery of APC-specific short-hairpin (sh) RNAs, and assessed using quantitative (q) reverse-transcriptase (RT)-PCR and western blotting. Tumor cell motility was analyzed by performing wound-filling assays, and morphology via immunofluorescence (IF) and phase-contrast microscopy. Additionally, proliferation was measured using BrdU incorporation, and TCF reporter assays were performed to determine β-catenin/TCF-mediated transcriptional activity. APC/β-catenin-rich complexes were observed at protrusion ends of migratory epithelial cells treated with a proteasome inhibitor or when EMT has been induced and in tumor cells with a mesenchymal, spindle-like morphology. 4T07 tumor cells with reduced APC levels were significantly less motile and had a more rounded morphology; yet, they did not differ significantly in proliferation or β-catenin/TCF transcriptional activity. Furthermore, we found that APC/β-catenin-rich complexes at protrusion ends were dependent upon an intact microtubule cytoskeleton. These findings indicate that membrane protrusions with APC/β-catenin-containing puncta control the migratory potential and

  10. Homeoviscous adaptation and the regulation of membrane lipids

    DEFF Research Database (Denmark)

    Ernst, Robert; Ejsing, Christer S; Antonny, Bruno


    Biological membranes are complex and dynamic assemblies of lipids and proteins. Poikilothermic organisms including bacteria, fungi, reptiles, and fish do not control their body temperature and must adapt their membrane lipid composition in order to maintain membrane fluidity in the cold. This ada......Biological membranes are complex and dynamic assemblies of lipids and proteins. Poikilothermic organisms including bacteria, fungi, reptiles, and fish do not control their body temperature and must adapt their membrane lipid composition in order to maintain membrane fluidity in the cold...... such as neurons maintain unique lipid compositions with specific physicochemical properties. To date little is known about the sensory mechanisms regulating the acyl chain profile in such specialized cells or during adaptive responses. Here we summarize our current understanding of lipid metabolic networks...

  11. Hybrid membranes for fuel cells (United States)

    Bochkareva, S. S.; Shashkina, S. S.


    Fuel cells are a very efficient, reliable, durable, and environmentally friendly energy source. Membranes for fuel cells were developed based on nitrogen-containing high-molecular compounds and organic–inorganic composites. Their electrical conductivities were measured. The influence of a silicon block of composites on the proton exchange properties of membranes was proved.The comparative characterization of the studied materials was performed.

  12. Cell-free reconstitution of vacuole membrane fragmentation reveals regulation of vacuole size and number by TORC1 (United States)

    Michaillat, Lydie; Baars, Tonie Luise; Mayer, Andreas


    Size and copy number of organelles are influenced by an equilibrium of membrane fusion and fission. We studied this equilibrium on vacuoles—the lysosomes of yeast. Vacuole fusion can readily be reconstituted and quantified in vitro, but it had not been possible to study fission of the organelle in a similar way. Here we present a cell-free system that reconstitutes fragmentation of purified yeast vacuoles (lysosomes) into smaller vesicles. Fragmentation in vitro reproduces physiological aspects. It requires the dynamin-like GTPase Vps1p, V-ATPase pump activity, cytosolic proteins, and ATP and GTP hydrolysis. We used the in vitro system to show that the vacuole-associated TOR complex 1 (TORC1) stimulates vacuole fragmentation but not the opposing reaction of vacuole fusion. Under nutrient restriction, TORC1 is inactivated, and the continuing fusion activity then dominates the fusion/fission equilibrium, decreasing the copy number and increasing the volume of the vacuolar compartment. This result can explain why nutrient restriction not only induces autophagy and a massive buildup of vacuolar/lysosomal hydrolases, but also leads to a concomitant increase in volume of the vacuolar compartment by coalescence of the organelles into a single large compartment. PMID:22238359

  13. Expression of progesterone receptor membrane component-2 within the immature rat ovary and its role in regulating mitosis and apoptosis of spontaneously immortalized granulosa cells. (United States)

    Griffin, Daniel; Liu, Xiufang; Pru, Cindy; Pru, James K; Peluso, John J


    Progesterone receptor membrane component 2 (Pgrmc2) mRNA was detected in the immature rat ovary. By 48 h after eCG, Pgrmc2 mRNA levels decreased by 40% and were maintained at 48 h post-hCG. Immunohistochemical studies detected PGRMC2 in oocytes and ovarian surface epithelial, interstitial, thecal, granulosa, and luteal cells. PGRMC2 was also present in spontaneously immortalized granulosa cells, localizing to the cytoplasm of interphase cells and apparently to the mitotic spindle of cells in metaphase. Interestingly, PGRMC2 levels appeared to decrease during the G1 stage of the cell cycle. Moreover, overexpression of PGRMC2 suppressed entry into the cell cycle, possibly by binding the p58 form of cyclin dependent kinase 11b. Conversely, Pgrmc2 small interfering RNA (siRNA) treatment increased the percentage of cells in G1 and M stage but did not increase the number of cells, which was likely due to an increase in apoptosis. Depleting PGRMC2 did not inhibit cellular (3)H-progesterone binding, but attenuated the ability of progesterone to suppress mitosis and apoptosis. Taken together these studies suggest that PGRMC2 affects granulosa cell mitosis by acting at two specific stages of the cell cycle. First, PGRMC2 regulates the progression from the G0 into the G1 stage of the cell cycle. Second, PGRMC2 appears to localize to the mitotic spindle, where it likely promotes the final stages of mitosis. Finally, siRNA knockdown studies indicate that PGRMC2 is required for progesterone to slow the rate of granulosa cell mitosis and apoptosis. These findings support a role for PGRMC2 in ovarian follicle development. © 2014 by the Society for the Study of Reproduction, Inc.

  14. Cell invasion through basement membrane (United States)

    Morrissey, Meghan A; Hagedorn, Elliott J; Sherwood, David R


    Cell invasion through basement membrane is an essential part of normal development and physiology, and occurs during the pathological progression of human inflammatory diseases and cancer. F-actin-rich membrane protrusions, called invadopodia, have been hypothesized to be the “drill bits” of invasive cells, mediating invasion through the dense, highly cross-linked basement membrane matrix. Though studied in vitro for over 30 y, invadopodia function in vivo has remained elusive. We have recently discovered that invadopodia breach basement membrane during anchor cell invasion in C. elegans, a genetically and visually tractable in vivo invasion event. Further, we found that the netrin receptor DCC localizes to the initial site of basement membrane breach and directs invasion through a single gap in the matrix. In this commentary, we examine how the dynamics and structure of AC-invadopodia compare with in vitro invadopodia and how the netrin receptor guides invasion through a single basement membrane breach. We end with a discussion of our surprising result that the anchor cell pushes the basement membrane aside, instead of completely dissolving it through proteolysis, and provide some ideas for how proteases and physical displacement may work together to ensure efficient and robust invasion. PMID:24778942

  15. Formation and Regulation of Mitochondrial Membranes

    Directory of Open Access Journals (Sweden)

    Laila Cigana Schenkel


    Full Text Available Mitochondrial membrane phospholipids are essential for the mitochondrial architecture, the activity of respiratory proteins, and the transport of proteins into the mitochondria. The accumulation of phospholipids within mitochondria depends on a coordinate synthesis, degradation, and trafficking of phospholipids between the endoplasmic reticulum (ER and mitochondria as well as intramitochondrial lipid trafficking. Several studies highlight the contribution of dietary fatty acids to the remodeling of phospholipids and mitochondrial membrane homeostasis. Understanding the role of phospholipids in the mitochondrial membrane and their metabolism will shed light on the molecular mechanisms involved in the regulation of mitochondrial function and in the mitochondrial-related diseases.

  16. Efficient overproduction of membrane proteins in Lactococcus lactis requires the cell envelope stress sensor/regulator couple CesSR

    NARCIS (Netherlands)

    Pinto, Joao P C; Kuipers, Oscar P; Marreddy, Ravi K R; Poolman, Bert; Kok, Jan


    BACKGROUND: Membrane proteins comprise an important class of molecules whose study is largely frustrated by several intrinsic constraints, such as their hydrophobicity and added requirements for correct folding. Additionally, the complexity of the cellular mechanisms that are required to insert

  17. Autoinhibitory Regulation of Plasma Membrane H+-ATPases

    DEFF Research Database (Denmark)

    Pedersen, Jesper Torbøl

    Electrochemical gradients across cell membranes are essential for nutrient uptake. In plant and fungal cells the electrochemical gradient across the plasma membrane (PM) can build much higher than in mammalian cells. The protein responsible for this gradient is the essential PM H+-ATPase that uses...... a huge amount of energy in form of ATP, to pump out protons. To avoid complete energy depletion in the cells, tight regulation of the PM H+-ATPase is a necessity. The proteins two terminal domains have been identified as autoinhibitory domains that regulate the pumping activity, but due to lack of a high...... in mammalian cells and it has been speculated if they have a similar function in plants. In this thesis we show, that plant PM H+-ATPases are receptors for lysophospholipids and the autoinhibitory terminal inhibition is released upon lysophospholipid binding. Finally, we have used a group of stabilizing...

  18. Intracellular cholesterol level regulates sensitivity of glioblastoma cells against temozolomide-induced cell death by modulation of caspase-8 activation via death receptor 5-accumulation and activation in the plasma membrane lipid raft. (United States)

    Yamamoto, Yutaro; Tomiyama, Arata; Sasaki, Nobuyoshi; Yamaguchi, Hideki; Shirakihara, Takuya; Nakashima, Katsuhiko; Kumagai, Kosuke; Takeuchi, Satoru; Toyooka, Terushige; Otani, Naoki; Wada, Kojiro; Narita, Yoshitaka; Ichimura, Koichi; Sakai, Ryuichi; Namba, Hiroki; Mori, Kentaro


    Development of resistance against temozolomide (TMZ) in glioblastoma (GBM) after continuous treatment with TMZ is one of the critical problems in clinical GBM therapy. Intracellular cholesterol regulates cancer cell biology, but whether intracellular cholesterol is involved in TMZ resistance of GBM cells remains unclear. The involvement of intracellular cholesterol in acquired resistance against TMZ in GBM cells was investigated. Intracellular cholesterol levels were measured in human U251 MG cells with acquired TMZ resistance (U251-R cells) and TMZ-sensitive control U251 MG cells (U251-Con cells), and found that the intracellular cholesterol level was significantly lower in U251-R cells than in U251-Con cells. In addition, treatment by intracellular cholesterol remover, methyl-beta cyclodextrin (MβCD), or intracellular cholesterol inducer, soluble cholesterol (Chol), regulated TMZ-induced U251-Con cell death in line with changes in intracellular cholesterol level. Involvement of death receptor 5 (DR5), a death receptor localized in the plasma membrane, was evaluated. TMZ without or with MβCD and/or Chol caused accumulation of DR5 into the plasma membrane lipid raft and formed a complex with caspase-8, an extrinsic caspase cascade inducer, reflected in the induction of cell death. In addition, treatment with caspase-8 inhibitor or knockdown of DR5 dramatically suppressed U251-Con cell death induced by combination treatment with TMZ, MβCD, and Chol. Combined treatment of Chol with TMZ reversed the TMZ resistance of U251-R cells and another GBM cell model with acquired TMZ resistance, whereas clinical antihypercholesterolemia agents at physiological concentrations suppressed TMZ-induced cell death of U251-Con cells. These findings suggest that intracellular cholesterol level affects TMZ treatment of GBM mediated via a DR5-caspase-8 mechanism. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Endoplasmic reticulum membrane reorganization is regulated by ionic homeostasis. (United States)

    Varadarajan, Shankar; Tanaka, Kayoko; Smalley, Joshua L; Bampton, Edward T W; Pellecchia, Maurizio; Dinsdale, David; Willars, Gary B; Cohen, Gerald M


    Recently we described a new, evolutionarily conserved cellular stress response characterized by a reversible reorganization of endoplasmic reticulum (ER) membranes that is distinct from canonical ER stress and the unfolded protein response (UPR). Apogossypol, a putative broad spectrum BCL-2 family antagonist, was the prototype compound used to induce this ER membrane reorganization. Following microarray analysis of cells treated with apogossypol, we used connectivity mapping to identify a wide range of structurally diverse chemicals from different pharmacological classes and established their ability to induce ER membrane reorganization. Such structural diversity suggests that the mechanisms initiating ER membrane reorganization are also diverse and a major objective of the present study was to identify potentially common features of these mechanisms. In order to explore this, we used hierarchical clustering of transcription profiles for a number of chemicals that induce membrane reorganization and discovered two distinct clusters. One cluster contained chemicals with known effects on Ca(2+) homeostasis. Support for this was provided by the findings that ER membrane reorganization was induced by agents that either deplete ER Ca(2+) (thapsigargin) or cause an alteration in cellular Ca(2+) handling (calmodulin antagonists). Furthermore, overexpression of the ER luminal Ca(2+) sensor, STIM1, also evoked ER membrane reorganization. Although perturbation of Ca(2+) homeostasis was clearly one mechanism by which some agents induced ER membrane reorganization, influx of extracellular Na(+) but not Ca(2+) was required for ER membrane reorganization induced by apogossypol and the related BCL-2 family antagonist, TW37, in both human and yeast cells. Not only is this novel, non-canonical ER stress response evolutionary conserved but so also are aspects of the mechanism of formation of ER membrane aggregates. Thus perturbation of ionic homeostasis is important in the regulation

  20. The regulated expression, intracellular trafficking, and membrane recycling of the P2Y-like receptor GPR17 in Oli-neu oligodendroglial cells. (United States)

    Fratangeli, Alessandra; Parmigiani, Elena; Fumagalli, Marta; Lecca, Davide; Benfante, Roberta; Passafaro, Maria; Buffo, Annalisa; Abbracchio, Maria P; Rosa, Patrizia


    GPR17 is a G-protein-coupled receptor that is activated by two classes of molecules: uracil-nucleotides and cysteinyl-leukotrienes. GPR17 is required for initiating the differentiation of oligodendrocyte precursors but has to be down-regulated to allow cells to undergo terminal maturation. Although a great deal has been learned about GPR17 expression and signaling, no information is currently available about the trafficking of native receptors after the exposure of differentiating oligodendrocytes to endogenous agonists. Here, we demonstrate that neuron-conditioned medium induces the transcriptionally mediated, time-regulated expression of GPR17 in Oli-neu, an oligodendrocyte precursor cell line, making these cells suitable for studying the endocytic traffic of the native receptor. Agonist-induced internalization, intracellular trafficking, and membrane recycling of GPR17 were analyzed by biochemical and immunofluorescence assays using an ad hoc-developed antibody against the extracellular N-terminal of GPR17. Both UDP-glucose and LTD(4) increased GPR17 internalization, although with different efficiency. At early time points, internalized GPR17 co-localized with transferrin receptor, whereas at later times it partially co-localized with the lysosomal marker Lamp1, suggesting that a portion of GPR17 is targeted to lysosomes upon ligand binding. An analysis of receptor recycling and degradation demonstrated that a significant aliquot of GPR17 is recycled to the cell surface. Furthermore, internalized GPR17 displayed a co-localization with the marker of the "short loop" recycling endosomes, Rab4, while showing very minor co-localization with the "long loop" recycling marker, Rab11. Our results provide the first data on the agonist-induced trafficking of native GPR17 in oligodendroglial cells and may have implications for both physiological and pathological myelination.

  1. Cell membrane structures during exocytosis. (United States)

    Savigny, Pascale; Evans, John; McGrath, Kathryn M


    Exocytosis is a key biological process that controls the neurotransmission and release of hormones from cells. In endocrine cells, hormones are packed into secretory vesicles and released into the extracellular environment via openings in the plasma membrane, a few hundred nanometers wide, which form as a result of fusion of the membranes of the granule and cell. The complex processes and dynamics that result in the formation of the fusion pore, as well as its structure, remain scantly understood. A number of different exocytosis mechanisms have been postulated. Furthermore, the possibility exists that several mechanisms occur simultaneously. We present here an investigation of the cell membrane dynamics during exocytosis in anterior pituitary cells, especially gonadotropes, which secrete LH, a hormone central to ovulation. Gonadotrope enrichment was achieved using immunolabeled magnetic nanobeads. Three complementary imaging techniques were used to realize a fine structure study of the dynamics of the exocytosis-like sites occurring during secretion. Living pituitary and gonadotrope-enriched cells were imaged with atomic force microscopy, as well as cells that had been fixed to obtain better resolution. Atomic force microscopy, along with scanning and transmission electron microscopy, studies of these cells revealed that there are at least two different site configurations: simple single fusion pores and a complex association of pores consisting of a simple primary site combined with secondary attachments.

  2. Corrugated Membrane Fuel Cell Structures

    Energy Technology Data Exchange (ETDEWEB)

    Grot, Stephen [President, Ion Power Inc.


    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.

  3. Negative regulators of cell proliferation (United States)

    Johnson, T. C.; Spooner, B. S. (Principal Investigator)


    Cell proliferation is governed by the influence of both mitogens and inhibitors. Although cell contact has long been thought to play a fundamental role in cell cycling regulation, and negative regulators have long been suspected to exist, their isolation and purification has been complicated by a variety of technical difficulties. Nevertheless, over recent years an ever-expanding list of putative negative regulators have emerged. In many cases, their biological inhibitory activities are consistent with density-dependent growth inhibition. Most likely their interactions with mitogenic agents, at an intracellular level, are responsible for either mitotic arrest or continued cell cycling. A review of naturally occurring cell growth inhibitors is presented with an emphasis on those factors shown to be residents of the cell surface membrane. Particular attention is focused on a cell surface sialoglycopeptide, isolated from intact bovine cerebral cortex cells, which has been shown to inhibit the proliferation of an unusually wide range of target cells. The glycopeptide arrest cells obtained from diverse species, both fibroblasts and epithelial cells, and a broad variety of transformed cells. Signal transduction events and a limited spectrum of cells that are refractory to the sialoglycopeptide have provided insight into the molecular events mediated by this cell surface inhibitor.

  4. Molecular machines open cell membranes. (United States)

    García-López, Víctor; Chen, Fang; Nilewski, Lizanne G; Duret, Guillaume; Aliyan, Amir; Kolomeisky, Anatoly B; Robinson, Jacob T; Wang, Gufeng; Pal, Robert; Tour, James M


    Beyond the more common chemical delivery strategies, several physical techniques are used to open the lipid bilayers of cellular membranes. These include using electric and magnetic fields, temperature, ultrasound or light to introduce compounds into cells, to release molecular species from cells or to selectively induce programmed cell death (apoptosis) or uncontrolled cell death (necrosis). More recently, molecular motors and switches that can change their conformation in a controlled manner in response to external stimuli have been used to produce mechanical actions on tissue for biomedical applications. Here we show that molecular machines can drill through cellular bilayers using their molecular-scale actuation, specifically nanomechanical action. Upon physical adsorption of the molecular motors onto lipid bilayers and subsequent activation of the motors using ultraviolet light, holes are drilled in the cell membranes. We designed molecular motors and complementary experimental protocols that use nanomechanical action to induce the diffusion of chemical species out of synthetic vesicles, to enhance the diffusion of traceable molecular machines into and within live cells, to induce necrosis and to introduce chemical species into live cells. We also show that, by using molecular machines that bear short peptide addends, nanomechanical action can selectively target specific cell-surface recognition sites. Beyond the in vitro applications demonstrated here, we expect that molecular machines could also be used in vivo, especially as their design progresses to allow two-photon, near-infrared and radio-frequency activation.

  5. Molecular machines open cell membranes (United States)

    García-López, Víctor; Chen, Fang; Nilewski, Lizanne G.; Duret, Guillaume; Aliyan, Amir; Kolomeisky, Anatoly B.; Robinson, Jacob T.; Wang, Gufeng; Pal, Robert; Tour, James M.


    Beyond the more common chemical delivery strategies, several physical techniques are used to open the lipid bilayers of cellular membranes. These include using electric and magnetic fields, temperature, ultrasound or light to introduce compounds into cells, to release molecular species from cells or to selectively induce programmed cell death (apoptosis) or uncontrolled cell death (necrosis). More recently, molecular motors and switches that can change their conformation in a controlled manner in response to external stimuli have been used to produce mechanical actions on tissue for biomedical applications. Here we show that molecular machines can drill through cellular bilayers using their molecular-scale actuation, specifically nanomechanical action. Upon physical adsorption of the molecular motors onto lipid bilayers and subsequent activation of the motors using ultraviolet light, holes are drilled in the cell membranes. We designed molecular motors and complementary experimental protocols that use nanomechanical action to induce the diffusion of chemical species out of synthetic vesicles, to enhance the diffusion of traceable molecular machines into and within live cells, to induce necrosis and to introduce chemical species into live cells. We also show that, by using molecular machines that bear short peptide addends, nanomechanical action can selectively target specific cell-surface recognition sites. Beyond the in vitro applications demonstrated here, we expect that molecular machines could also be used in vivo, especially as their design progresses to allow two-photon, near-infrared and radio-frequency activation.


    NARCIS (Netherlands)



    The effect of alpha-1-adrenoceptor stimulation on inositol phosphates (InsPs), intracellular Ca2+ and membrane current was measured in vas deferens DDT1 MF-2 cells. The InsPs were analyzed after labelling the cells with [H-3]myo-inositol using high performance liquid chromatography and the internal

  7. The effect of membrane-regulated actin polymerization on a two-phase flow model for cell motility

    KAUST Repository

    Kimpton, L. S.


    Two-phase flow models have been widely used to model cell motility and we have previously demonstrated that even the simplest, stripped-down, 1D model displays many observed features of cell motility [Kimpton, L.S., Whiteley, J.P., Waters, S.L., King, J.R. & Oliver, J.M. (2013) Multiple travelling-wave solutions in a minimal model for cell motility. Math. Med. Biol. 30, 241 - 272]. In this paper, we address a limitation of the previous model.We show that the two-phase flow framework can exhibit travelling-wave solutions with biologically plausible actin network profiles in two simple models that enforce polymerization or depolymerization of the actin network at the ends of the travelling, 1D strip of cytoplasm. © 2014 The authors 2014. Published by Oxford University Press on behalf of the Institute of Mathematics and its Applications. All rights reserved.

  8. Roles of membrane trafficking in plant cell wall dynamics

    Directory of Open Access Journals (Sweden)

    Kazuo eEbine


    Full Text Available 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.

  9. Matrix metalloproteinase 2 and membrane type 1 matrix metalloproteinase co-regulate axonal outgrowth of mouse retinal ganglion cells

    DEFF Research Database (Denmark)

    Gaublomme, Djoere; Buyens, Tom; De Groef, Lies


    , we were able to show that broad-spectrum MMP inhibition reduces axon outgrowth of mouse retinal ganglion cells (RGCs), implicating MMPs as beneficial factors in axonal regeneration. Additional studies, using more specific MMP inhibitors and MMP-deficient mice, disclosed that both MMP-2 and MT1-MMP......, but not MMP-9, are involved in this process. Furthermore, administration of a novel antibody to MT1-MMP that selectively blocks pro-MMP-2 activation revealed a functional co-involvement of these proteinases in determining RGC axon outgrowth. Subsequent immunostainings showed expression of both MMP-2 and MT1......-MMP in RGC axons and glial cells. Finally, results from combined inhibition of MMP-2 and β1-integrin were suggestive for a functional interaction between these molecules. Overall, our data indicate MMP-2 and MT1-MMP as promising axonal outgrowth-promoting molecules. Axonal regeneration in the central...

  10. Membrane tension and cytoskeleton organization in cell motility. (United States)

    Sens, Pierre; Plastino, Julie


    Cell membrane shape changes are important for many aspects of normal biological function, such as tissue development, wound healing and cell division and motility. Various disease states are associated with deregulation of how cells move and change shape, including notably tumor initiation and cancer cell metastasis. Cell motility is powered, in large part, by the controlled assembly and disassembly of the actin cytoskeleton. Much of this dynamic happens in close proximity to the plasma membrane due to the fact that actin assembly factors are membrane-bound, and thus actin filaments are generally oriented such that their growth occurs against or near the membrane. For a long time, the membrane was viewed as a relatively passive scaffold for signaling. However, results from the last five years show that this is not the whole picture, and that the dynamics of the actin cytoskeleton are intimately linked to the mechanics of the cell membrane. In this review, we summarize recent findings concerning the role of plasma membrane mechanics in cell cytoskeleton dynamics and architecture, showing that the cell membrane is not just an envelope or a barrier for actin assembly, but is a master regulator controlling cytoskeleton dynamics and cell polarity.

  11. Substrate curvature regulates cell migration. (United States)

    He, Xiuxiu; Jiang, Yi


    Cell migration is essential in many aspects of biology. Many basic migration processes, including adhesion, membrane protrusion and tension, cytoskeletal polymerization, and contraction, have to act in concert to regulate cell migration. At the same time, substrate topography modulates these processes. In this work, we study how substrate curvature at micrometer scale regulates cell motility. We have developed a 3D mechanical model of single cell migration and simulated migration on curved substrates with different curvatures. The simulation results show that cell migration is more persistent on concave surfaces than on convex surfaces. We have further calculated analytically the cell shape and protrusion force for cells on curved substrates. We have shown that while cells spread out more on convex surfaces than on concave ones, the protrusion force magnitude in the direction of migration is larger on concave surfaces than on convex ones. These results offer a novel biomechanical explanation to substrate curvature regulation of cell migration: geometric constrains bias the direction of the protrusion force and facilitates persistent migration on concave surfaces.

  12. Fuel-Cell Structure Prevents Membrane Drying (United States)

    Mcelroy, J.


    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.

  13. A novel approach to regulate cell membrane permeability for ATP and NADH formation in Saccharomyces cerevisiae induced by air cold plasma (United States)

    Dong, Xiaoyu; Liu, Tingting; Xiong, Yuqin


    Air cold plasma has been used as a novel method for enhancing microbial fermentation. The aim of this work was to explore the effect of plasma on membrane permeability and the formation of ATP and NADH in Saccharomyces cerevisiae, so as to provide valuable information for large-scale application of plasma in the fermentation industry. Suspensions of S. cerevisiae cells were exposed to air cold plasma for 0, 1, 2, 3, 4 and 5 min, and then subjected to various analyses prior to fermentation (0 h) and at the 9 and 21 h stages of fermentation. Compared with non-exposed cells, cells exposed to plasma for 1 min exhibited a marked increase in cytoplasmic free Ca2+ concentration as a result of the significant increase in membrane potential prior to fermentation. At the same time, the ATP level in the cell suspension decreased by about 40%, resulting in a reduction of about 60% in NADH prior to culturing. However, the levels of ATP and NADH in the culture at the 9 and 21 h fermentation stages were different from the level at 0 h. Taken together, the results indicated that exposure of S. cerevisiae to air cold plasma could increase its cytoplasmic free Ca2+ concentration by improving the cell membrane potential, consequently leading to changes in ATP and NADH levels. Supported by National Natural Science Foundation of China (Nos. 21246012, 21306015 and 21476032).

  14. In-membrane micro fuel cell

    Energy Technology Data Exchange (ETDEWEB)

    Omosebi, Ayokunle; Besser, Ronald


    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.

  15. Regulation of Trafficking, Membrane Retention and Turnover of the Na+, HCO3- Co-Transporter NBCn1 in Epithelial Cells

    DEFF Research Database (Denmark)

    Olesen, Christina Wilkens

    of cell types. It contributes importantly to the normal physiological function of many human organs, and its dysfunction has been linked to several pathologies including cardiovascular diseases and breast cancer. Expression of a truncated constitutive active HER2 receptor, p95HER2, is associated with poor...... and mechanisms of NBCn1 trafficking, membrane retention and degradation in MCF-7 cells and other epithelial cell types. In Paper-I we review existing evidence regarding the possible roles of the SLC4 and SLC26 families of HCO3 - transporters in breast, colon and lung cancer. Bioinformatic analysis of publicly...

  16. D120 and K152 within the PH Domain of T Cell Adapter SKAP55 Regulate Plasma Membrane Targeting of SKAP55 and LFA-1 Affinity Modulation in Human T Lymphocytes. (United States)

    Witte, Amelie; Meineke, Bernhard; Sticht, Jana; Philipsen, Lars; Kuropka, Benno; Müller, Andreas J; Freund, Christian; Schraven, Burkhart; Kliche, Stefanie


    The β2-integrin lymphocyte function-associated antigen 1 (LFA-1) is needed for the T cell receptor (TCR)-induced activation of LFA-1 to promote T cell adhesion and interaction with antigen-presenting cells (APCs). LFA-1-mediated cell-cell interactions are critical for proper T cell differentiation and proliferation. The Src kinase-associated phosphoprotein of 55 kDa (SKAP55) is a key regulator of TCR-mediated LFA-1 signaling (inside-out/outside-in signaling). To gain an understanding of how SKAP55 controls TCR-mediated LFA-1 activation, we assessed the functional role of its pleckstrin homology (PH) domain. We identified two critical amino acid residues within the PH domain of SKAP55, aspartic acid 120 (D120) and lysine 152 (K152). D120 facilitates the retention of SKAP55 in the cytoplasm of nonstimulated T cells, while K152 promotes SKAP55 membrane recruitment via actin binding upon TCR triggering. Importantly, the K152-dependent interaction of the PH domain with actin promotes the binding of talin to LFA-1, thus facilitating LFA-1 activation. These data suggest that K152 and D120 within the PH domain of SKAP55 regulate plasma membrane targeting and TCR-mediated activation of LFA-1. Copyright © 2017 American Society for Microbiology.

  17. Fuel cell and membrane therefore

    Energy Technology Data Exchange (ETDEWEB)

    Aindow, Tai-Tsui


    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.

  18. Regulation of transport processes across the tonoplast membrane

    Directory of Open Access Journals (Sweden)

    Oliver eTrentmann


    Full Text Available In plants, the vacuole builds up the cellular turgor and represents an important component in cellular responses to diverse stress stimuli. Rapid volume changes of cells, particularly of motor cells, like guard cells, are caused by variation of osmolytes and consequently of the water contents in the vacuole. Moreover, directed solute uptake into or release out of the large central vacuole allows adaptation of cytosolic metabolite levels according to the current physiological requirements and specific cellular demands. Therefore, solute passage across the vacuolar membrane, the tonoplast, has to be tightly regulated. Important principles in vacuolar transport regulation are changes of tonoplast transport protein abundances by differential expression of genes or changes of their activities, e.g. due to post-translational modification or by interacting proteins. Because vacuolar transport is in most cases driven by an electro-chemical gradient altered activities of tonoplast proton pumps significantly influence vacuolar transport capacities. Intense studies on individual tonoplast proteins but also unbiased system biological approaches have provided important insights into the regulation of vacuolar transport. This short review refers to selected examples of tonoplast proteins and their regulation, with special focus on protein phosphorylation.

  19. Anion permselective membrane. [For redox fuel cells

    Energy Technology Data Exchange (ETDEWEB)

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


    Experimental anion permeselective membranes were improved and characterized for use as separators in a chemical redox, power storage cell being developed at the NASA Lewis Research Center. The goal of minimal Fe/sup +3/ ion transfer was achieved for each candidate membrane system. Minimal membrane resistivity was demonstrated by reduction of film thickness using synthetic backing materials but usefulness of thin membranes was limited by the scarcity of compatible fabrics. The most durable and useful backing fabrics were modacrylics. One membrane, a copolymer of 4 vinylpyridine and vinyl benzylchloride was outstanding in overall electrochemical and physical properties. Long term (1000 hrs) membrane chemical and thermal durability in redox environment was shown by three candidate polymers and two membranes. The remainder had good durability at ambient temperature. Manufacturing capability was demonstrated for large scale production of membrane sheets 5.5 ft/sup 2/ in area for two candidate systems.

  20. Membrane-Initiated Estradiol Signaling Regulating Sexual Receptivity (United States)

    Micevych, Paul E.; Dewing, Phoebe


    Estradiol has profound actions on the structure and function of the nervous system. In addition to nuclear actions that directly modulate gene expression, the idea that estradiol can rapidly activate cell signaling by binding to membrane estrogen receptors (mERs) has emerged. Even the regulation of sexual receptivity, an action previously thought to be completely regulated by nuclear ERs, has been shown to have a membrane-initiated estradiol signaling (MIES) component. This highlighted the question of the nature of mERs. Several candidates have been proposed, ERα, ERβ, ER-X, GPR30 (G protein coupled estrogen receptor), and a receptor activated by a diphenylacrylamide compound, STX. Although each of these receptors has been shown to be active in specific assays, we present evidence for and against their participation in sexual receptivity by acting in the lordosis-regulating circuit. The initial MIES that activates the circuit is in the arcuate nucleus of the hypothalamus (ARH). Using both activation of μ-opioid receptors (MOR) in the medial preoptic nucleus and lordosis behavior, we document that both ERα and the STX-receptor participate in the required MIES. ERα and the STX-receptor activation of cell signaling are dependent on the transactivation of type 1 metabotropic glutamate receptors (mGluR1a) that augment progesterone synthesis in astrocytes and protein kinase C (PKC) in ARH neurons. While estradiol-induced sexual receptivity does not depend on neuroprogesterone, proceptive behaviors do. Moreover, the ERα and the STX-receptor activation of medial preoptic MORs and augmentation of lordosis were sensitive to mGluR1a blockade. These observations suggest a common mechanism through which mERs are coupled to intracellular signaling cascades, not just in regulating reproduction, but in actions throughout the neuraxis including the cortex, hippocampus, striatum, and dorsal root ganglias. PMID:22649369

  1. NCAM regulates cell motility

    DEFF Research Database (Denmark)

    Prag, Søren; Lepekhin, Eugene A; Kolkova, Kateryna


    Cell migration is required during development of the nervous system. The regulatory mechanisms for this process, however, are poorly elucidated. We show here that expression of or exposure to the neural cell adhesion molecule (NCAM) strongly affected the motile behaviour of glioma cells...... independently of homophilic NCAM interactions. Expression of the transmembrane 140 kDa isoform of NCAM (NCAM-140) caused a significant reduction in cellular motility, probably through interference with factors regulating cellular attachment, as NCAM-140-expressing cells exhibited a decreased attachment...... to a fibronectin substratum compared with NCAM-negative cells. Ectopic expression of the cytoplasmic part of NCAM-140 also inhibited cell motility, presumably via the non-receptor tyrosine kinase p59(fyn) with which NCAM-140 interacts. Furthermore, we showed that the extracellular part of NCAM acted as a paracrine...

  2. Diffuse Charge Effects in Fuel Cell Membranes

    NARCIS (Netherlands)

    Biesheuvel, P.M.; Franco, A.A.; Bazant, M.Z.


    It is commonly assumed that electrolyte membranes in fuel cells are electrically neutral, except in unsteady situations, when the double-layer capacitance is heuristically included in equivalent circuit calculations. Indeed, the standard model for electron transfer kinetics at the membrane/electrode

  3. Direct Cytoskeleton Forces Cause Membrane Softening in Red Blood Cells (United States)

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


    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

  4. Advanced membrane electrode assemblies for fuel cells (United States)

    Kim, Yu Seung; Pivovar, Bryan S.


    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.

  5. Cell volume and membrane stretch independently control K+ channel activity

    DEFF Research Database (Denmark)

    Bomholtz, Sofia Hammami; Willumsen, Niels J; Olsen, Hervør L


    A number of potassium channels including members of the KCNQ family and the Ca(2+) activated IK and SK, but not BK, are strongly and reversibly regulated by small changes in cell volume. It has been argued that this general regulation is mediated through sensitivity to changes in membrane stretch....... To test this hypothesis we have studied the regulation of KCNQ1 and BK channels after expression in Xenopus oocytes. Results from cell-attached patch clamp studies (approximately 50 microm(2) macropatches) in oocytes expressing BK channels demonstrate that the macroscopic volume-insensitive BK current...... that stretch and volume sensitivity can be considered two independent regulatory mechanisms....

  6. ABBA regulates plasma-membrane and actin dynamics to promote radial glia extension. (United States)

    Saarikangas, Juha; Hakanen, Janne; Mattila, Pieta K; Grumet, Martin; Salminen, Marjo; Lappalainen, Pekka


    Radial glia play key roles in neuronal migration, axon guidance, and neurogenesis during development of the central nervous system. However, the molecular mechanisms regulating growth and morphology of these extended cells are unknown. We show that ABBA, a novel member of the IRSp53-MIM protein family, is enriched in different types of radial glia. ABBA binds ATP-actin monomers with high affinity and deforms PtdIns(4,5)P(2)-rich membranes in vitro through its WH2 and IM domains, respectively. In radial-glia-like C6-R cells, ABBA localises to the interface between the actin cytoskeleton and plasma membrane, and its depletion by RNAi led to defects in lamellipodial dynamics and process extension. Together, this study identifies ABBA as a novel regulator of actin and plasma membrane dynamics in radial glial cells, and provides evidence that membrane binding and deformation activity is critical for the cellular functions of IRSp53-MIM-ABBA family proteins.

  7. A novel bioactive membrane by cell electrospinning. (United States)

    Chen, Haiping; Liu, Yuanyuan; Hu, Qingxi


    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. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. Cell-geometry-dependent changes in plasma membrane order direct stem cell signalling and fate. (United States)

    von Erlach, Thomas C; Bertazzo, Sergio; Wozniak, Michele A; Horejs, Christine-Maria; Maynard, Stephanie A; Attwood, Simon; Robinson, Benjamin K; Autefage, Hélène; Kallepitis, Charalambos; Del Río Hernández, Armando; Chen, Christopher S; Goldoni, Silvia; Stevens, Molly M


    Cell size and shape affect cellular processes such as cell survival, growth and differentiation 1-4 , thus establishing cell geometry as a fundamental regulator of cell physiology. The contributions of the cytoskeleton, specifically actomyosin tension, to these effects have been described, but the exact biophysical mechanisms that translate changes in cell geometry to changes in cell behaviour remain mostly unresolved. Using a variety of innovative materials techniques, we demonstrate that the nanostructure and lipid assembly within the cell plasma membrane are regulated by cell geometry in a ligand-independent manner. These biophysical changes trigger signalling events involving the serine/threonine kinase Akt/protein kinase B (PKB) that direct cell-geometry-dependent mesenchymal stem cell differentiation. Our study defines a central regulatory role by plasma membrane ordered lipid raft microdomains in modulating stem cell differentiation with potential translational applications.

  9. A palmitoylation switch mechanism regulates Rac1 function and membrane organization (United States)

    Navarro-Lérida, Inmaculada; Sánchez-Perales, Sara; Calvo, María; Rentero, Carles; Zheng, Yi; Enrich, Carlos; Del Pozo, Miguel A


    The small GTPase Rac1 plays important roles in many processes, including cytoskeletal reorganization, cell migration, cell-cycle progression and gene expression. The initiation of Rac1 signalling requires at least two mechanisms: GTP loading via the guanosine triphosphate (GTP)/guanosine diphosphate (GDP) cycle, and targeting to cholesterol-rich liquid-ordered plasma membrane microdomains. Little is known about the molecular mechanisms governing this specific compartmentalization. We show that Rac1 can incorporate palmitate at cysteine 178 and that this post-translational modification targets Rac1 for stabilization at actin cytoskeleton-linked ordered membrane regions. Palmitoylation of Rac1 requires its prior prenylation and the intact C-terminal polybasic region and is regulated by the triproline-rich motif. Non-palmitoylated Rac1 shows decreased GTP loading and lower association with detergent-resistant (liquid-ordered) membranes (DRMs). Cells expressing no Rac1 or a palmitoylation-deficient mutant have an increased content of disordered membrane domains, and markers of ordered membranes isolated from Rac1-deficient cells do not correctly partition in DRMs. Importantly, cells lacking Rac1 palmitoylation show spreading and migration defects. These data identify palmitoylation as a mechanism for Rac1 function in actin cytoskeleton remodelling by controlling its membrane partitioning, which in turn regulates membrane organization. PMID:22157745

  10. Plasma Membrane H(+)-ATPase Regulation in the Center of Plant Physiology. (United States)

    Falhof, Janus; Pedersen, Jesper Torbøl; Fuglsang, Anja Thoe; Palmgren, Michael


    The plasma membrane (PM) H(+)-ATPase is an important ion pump in the plant cell membrane. By extruding protons from the cell and generating a membrane potential, this pump energizes the PM, which is a prerequisite for growth. Modification of the autoinhibitory terminal domains activates PM H(+)-ATPase activity, and on this basis it has been hypothesized that these regulatory termini are targets for physiological factors that activate or inhibit proton pumping. In this review, we focus on the posttranslational regulation of the PM H(+)-ATPase and place regulation of the pump in an evolutionary and physiological context. The emerging picture is that multiple signals regulating plant growth interfere with the posttranslational regulation of the PM H(+)-ATPase. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

  11. Cooperative tumour cell membrane targeted phototherapy (United States)

    Kim, Heegon; Lee, Junsung; Oh, Chanhee; Park, Ji-Ho


    The targeted delivery of therapeutics using antibodies or nanomaterials has improved the precision and safety of cancer therapy. However, the paucity and heterogeneity of identified molecular targets within tumours have resulted in poor and uneven distribution of targeted agents, thus compromising treatment outcomes. Here, we construct a cooperative targeting system in which synthetic and biological nanocomponents participate together in the tumour cell membrane-selective localization of synthetic receptor-lipid conjugates (SR-lipids) to amplify the subsequent targeting of therapeutics. The SR-lipids are first delivered selectively to tumour cell membranes in the perivascular region using fusogenic liposomes. By hitchhiking with extracellular vesicles secreted by the cells, the SR-lipids are transferred to neighbouring cells and further spread throughout the tumour tissues where the molecular targets are limited. We show that this tumour cell membrane-targeted delivery of SR-lipids leads to uniform distribution and enhanced phototherapeutic efficacy of the targeted photosensitizer.

  12. Indole prevents Escherichia coli cell division by modulating membrane potential (United States)

    Chimerel, Catalin; Field, Christopher M.; Piñero-Fernandez, Silvia; Keyser, Ulrich F.; Summers, David K.


    Indole is a bacterial signalling molecule that blocks E. coli cell division at concentrations of 3–5 mM. We have shown that indole is a proton ionophore and that this activity is key to the inhibition of division. By reducing the electrochemical potential across the cytoplasmic membrane of E. coli, indole deactivates MinCD oscillation and prevents formation of the FtsZ ring that is a prerequisite for division. This is the first example of a natural ionophore regulating a key biological process. Our findings have implications for our understanding of membrane biology, bacterial cell cycle control and potentially for the design of antibiotics that target the cell membrane. PMID:22387460

  13. Regulation of membrane peptides by the Pseudomonas plasmid alk regulon. (United States)

    Benson, S; Oppici, M; Shapiro, J; Fennewald, M


    Pseudomonas putida strains carrying the plasmid alk genes will grow on n-alkanes. Induced alk+ strains contain membrane activities for alkane hydroxylation and dehydrogenation of aliphatic primary alcohols. P. putida cytoplasmic and outer membranes can be separated by sucrose gradient centrifugation after disruption of cells by either mild detergent lysis or passage through a French press. Both the membrane component of alkane hydroxylase and membrane alcohol dehydrogenase fractionated with the cytoplasmic membrane. Induction of the alk regulon resulted in the appearance of at least three new plasmid-determined cytoplasmic membrane peptides of about 59,000 (59K), 47,000 (47K), and 40,000 (40K) daltons as well as the disappearance of a pair of chromosomally encoded outer membrane peptides of about 43,000 daltons. The 40K peptide is the membrane component of alkane hydroxylase and the product of the plasmid alkB gene because the alkB1029 mutation altered the properties of alkane hydroxylase in whole cells, reduced its thermal stability in cell extracts, and led to increased electrophoretic mobility of the inducible 40K peptide. These results are consistent with a model for vectorial oxidation of n-alkanes in the cytoplasmic membrane of P. putida.

  14. Fuel cell subassemblies incorporating subgasketed thrifted membranes (United States)

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


    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.

  15. Role of phosphatidylinositol 4,5-bisphosphate in regulating EHD2 plasma membrane localization.

    Directory of Open Access Journals (Sweden)

    Laura C Simone

    Full Text Available The four mammalian C-terminal Eps15 homology domain-containing proteins (EHD1-EHD4 play pivotal roles in endocytic membrane trafficking. While EHD1, EHD3 and EHD4 associate with intracellular tubular/vesicular membranes, EHD2 localizes to the inner leaflet of the plasma membrane. Currently, little is known about the regulation of EHD2. Thus, we sought to define the factors responsible for EHD2's association with the plasma membrane. The subcellular localization of endogenous EHD2 was examined in HeLa cells using confocal microscopy. Although EHD partner proteins typically mediate EHD membrane recruitment, EHD2 was targeted to the plasma membrane independent of two well-characterized binding proteins, syndapin2 and EHBP1. Additionally, the EH domain of EHD2, which facilitates canonical EHD protein interactions, was not required to direct overexpressed EHD2 to the cell surface. On the other hand, several lines of evidence indicate that the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2 plays a crucial role in regulating EHD2 subcellular localization. Pharmacologic perturbation of PIP2 metabolism altered PIP2 plasma membrane distribution (as assessed by confocal microscopy, and caused EHD2 to redistribute away from the plasma membrane. Furthermore, overexpressed EHD2 localized to PIP2-enriched vacuoles generated by active Arf6. Finally, we show that although cytochalasin D caused actin microfilaments to collapse, EHD2 was nevertheless maintained at the plasma membrane. Intriguingly, cytochalasin D induced relocalization of both PIP2 and EHD2 to actin aggregates, supporting a role of PIP2 in controlling EHD2 subcellular localization. Altogether, these studies emphasize the significance of membrane lipid composition for EHD2 subcellular distribution and offer new insights into the regulation of this important endocytic protein.

  16. Omega 3 fatty acids chemosensitize multidrug resistant colon cancer cells by down-regulating cholesterol synthesis and altering detergent resistant membranes composition.


    Gelsomino, G; Gazzano, E


    Background The activity of P-glycoprotein (Pgp) and multidrug resistance related protein 1 (MRP1), two membrane transporters involved in multidrug resistance of colon cancer, is increased by high amounts of cholesterol in plasma membrane and detergent resistant membranes (DRMs). It has never been investigated whether omega 3 polyunsatured fatty acids (PUFAs), which modulate cholesterol homeostasis in dyslipidemic syndromes and have chemopreventive effects in colon cancer, may affect the respo...

  17. Cell membrane softening in human breast and cervical cancer cells (United States)

    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.


    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.

  18. A practical guide for the identification of membrane and plasma membrane proteins in human embryonic stem cells and human embryonal carcinoma cells.

    NARCIS (Netherlands)

    Dormeyer, W.; van Hoof, D.; Mummery, C.L.; Krijgsveld, J.; Heck, A.


    The identification of (plasma) membrane proteins in cells can provide valuable insights into the regulation of their biological processes. Pluripotent cells such as human embryonic stem cells and embryonal carcinoma cells are capable of unlimited self-renewal and share many of the biological

  19. Structure and properties of cell membranes. Volume 3: Methodology and properties of membranes

    International Nuclear Information System (INIS)

    Benga, G.


    This book covers the topics: Quantum chemical approach to study the mechanisms of proton translocation across membranes through protein molecules; monomolecular films as biomembrane models; planar lipid bilayers in relation to biomembranes; relation of liposomes to cell membranes; reconstitution of membrane transport systems; structure-function relationships in cell membranes as revealed by X-ray techniques; structure-function relationships in cell membranes as revealed by spin labeling ESR; structure and dynamics of cell membranes as revealed by NMR techniques; the effect of dietary lipids on the composition and properties of biological membranes and index

  20. Hematopoietic protein-1 regulates the actin membrane skeleton and membrane stability in murine erythrocytes.

    Directory of Open Access Journals (Sweden)

    Maia M Chan

    Full Text Available Hematopoietic protein-1 (Hem-1 is a hematopoietic cell specific member of the WAVE (Wiskott-Aldrich syndrome verprolin-homologous protein complex, which regulates filamentous actin (F-actin polymerization in many cell types including immune cells. However, the roles of Hem-1 and the WAVE complex in erythrocyte biology are not known. In this study, we utilized mice lacking Hem-1 expression due to a non-coding point mutation in the Hem1 gene to show that absence of Hem-1 results in microcytic, hypochromic anemia characterized by abnormally shaped erythrocytes with aberrant F-actin foci and decreased lifespan. We find that Hem-1 and members of the associated WAVE complex are normally expressed in wildtype erythrocyte progenitors and mature erythrocytes. Using mass spectrometry and global proteomics, Coomassie staining, and immunoblotting, we find that the absence of Hem-1 results in decreased representation of essential erythrocyte membrane skeletal proteins including α- and β- spectrin, dematin, p55, adducin, ankyrin, tropomodulin 1, band 3, and band 4.1. Hem1⁻/⁻ erythrocytes exhibit increased protein kinase C-dependent phosphorylation of adducin at Ser724, which targets adducin family members for dissociation from spectrin and actin, and subsequent proteolysis. Increased adducin Ser724 phosphorylation in Hem1⁻/⁻ erythrocytes correlates with decreased protein expression of the regulatory subunit of protein phosphatase 2A (PP2A, which is required for PP2A-dependent dephosphorylation of PKC targets. These results reveal a novel, critical role for Hem-1 in the homeostasis of structural proteins required for formation and stability of the actin membrane skeleton in erythrocytes.

  1. Focus on Membrane Differentiation and Membrane Domains in the Prokaryotic Cell

    NARCIS (Netherlands)

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


    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

  2. Alternate Fuel Cell Membranes for Energy Independence

    Energy Technology Data Exchange (ETDEWEB)

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


    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

  3. Membrane phosphorylation and nerve cell function

    International Nuclear Information System (INIS)

    Baer, P.R.


    This thesis deals with the phosphorylation of membrane components. In part I a series of experiments is described using the hippocampal slice as a model system. In part II a different model system - cultured hybrid cells - is used to study protein and lipid phosphorylation, influenced by incubation with neuropeptides. In part III in vivo and in vitro studies are combined to study protein phosphorylation after neuroanatomical lesions. In a section of part II (Page 81-90) labelling experiments of the membrane inositol-phospholipids are described. 32 P-ATP was used to label phospholipids in intact hybrid cells, and short incubations were found to be the most favourable. (C.F.)

  4. Remodelling of primary human CD4+ T cell plasma membrane order by n-3 PUFA. (United States)

    Fan, Yang-Yi; Fuentes, Natividad R; Hou, Tim Y; Barhoumi, Rola; Li, Xian C; Deutz, Nicolaas E P; Engelen, Marielle P K J; McMurray, David N; Chapkin, Robert S


    Cell membrane fatty acids influence fundamental properties of the plasma membrane, including membrane fluidity, protein functionality, and lipid raft signalling. Evidence suggests that dietary n-3 PUFA may target the plasma membrane of immune cells by altering plasma membrane lipid dynamics, thereby regulating the attenuation of immune cell activation and suppression of inflammation. As lipid-based immunotherapy might be a promising new clinical strategy for the treatment of inflammatory disorders, we conducted in vitro and in vivo experiments to examine the effects of n-3 PUFA on CD4+ T cell membrane order, mitochondrial bioenergetics and lymphoproliferation. n-3 PUFA were incorporated into human primary CD4+ T cells phospholipids in vitro in a dose-dependent manner, resulting in a reduction in whole cell membrane order, oxidative phosphorylation and proliferation. At higher doses, n-3 PUFA induced unique phase separation in T cell-derived giant plasma membrane vesicles. Similarly, in a short-term human pilot study, supplementation of fish oil (4 g n-3 PUFA/d) for 6 weeks in healthy subjects significantly elevated EPA (20 : 5n-3) levels in CD4+ T cell membrane phospholipids, and reduced membrane lipid order. These results demonstrate that the dynamic reshaping of human CD4+ T cell plasma membrane organisation by n-3 PUFA may modulate down-stream clonal expansion.

  5. Biglycan and decorin differentially regulate signaling in the fetal membranes (United States)

    Wu, Zhiping; Horgan, Casie E.; Carr, Olivia; Owens, Rick T.; Iozzo, Renato V.; Lechner, Beatrice E.


    Preterm birth is the leading cause of newborn mortality in the United States and about one third of cases are caused by preterm premature rupture of fetal membranes, a complication that is frequently observed in patients with Ehlers-Danlos Syndrome. Notably, a subtype of Ehlers-Danlos Syndrome is caused by expression of abnormal biglycan and decorin proteoglycans. As compound deficiency of these two small leucine-rich proteoglycans is a model of preterm birth, we investigated the fetal membranes of Bgn−/−;Dcn−/− double-null and single-null mice. Our results showed that biglycan signaling supported fetal membrane remodeling during early gestation in the absence of concomitant changes in TGFβ levels. In late gestation, biglycan signaling acted in a TGFβ–dependent manner to aid in membrane stabilization. In contrast, decorin signaling supported fetal membrane remodeling at early stages of gestation in a TGFβ–dependent manner, and fetal membrane stabilization at later stages of gestation without changes in TGFβ levels. Furthermore, exogenous soluble decorin was capable of rescuing the TGFβ signaling pathway in fetal membrane mesenchymal cells. Collectively, these findings provide novel targets for manipulation of fetal membrane extracellular matrix stability and could represent novel targets for research on preventive strategies for preterm premature rupture of fetal membranes. PMID:24373743

  6. Regulation of beta cell replication

    DEFF Research Database (Denmark)

    Lee, Ying C; Nielsen, Jens Høiriis


    Beta cell mass, at any given time, is governed by cell differentiation, neogenesis, increased or decreased cell size (cell hypertrophy or atrophy), cell death (apoptosis), and beta cell proliferation. Nutrients, hormones and growth factors coupled with their signalling intermediates have been...... suggested to play a role in beta cell mass regulation. In addition, genetic mouse model studies have indicated that cyclins and cyclin-dependent kinases that determine cell cycle progression are involved in beta cell replication, and more recently, menin in association with cyclin-dependent kinase...... inhibitors has been demonstrated to be important in beta cell growth. In this review, we consider and highlight some aspects of cell cycle regulation in relation to beta cell replication. The role of cell cycle regulation in beta cell replication is mostly from studies in rodent models, but whether...

  7. Membrane lipidome of an epithelial cell line

    DEFF Research Database (Denmark)

    Sampaio, Julio L; Gerl, Mathias J; Klose, Christian


    Tissue differentiation is an important process that involves major cellular membrane remodeling. We used Madin-Darby canine kidney cells as a model for epithelium formation and investigated the remodeling of the total cell membrane lipidome during the transition from a nonpolarized morphology...... to an epithelial morphology and vice versa. To achieve this, we developed a shotgun-based lipidomics workflow that enabled the absolute quantification of mammalian membrane lipidomes with minimal sample processing from low sample amounts. Epithelial morphogenesis was accompanied by a major shift from sphingomyelin...... to glycosphingolipid, together with an increase in plasmalogen, phosphatidylethanolamine, and cholesterol content, whereas the opposite changes took place during an epithelial-to-mesenchymal transition. Moreover, during polarization, the sphingolipids became longer, more saturated, and more hydroxylated as required...

  8. Lithium. Effects on excitable cell membranes

    NARCIS (Netherlands)

    Ploeger, Egbert Johan


    LITHIUM: Effects on excitable cell membranes. Lithium salts have been used in the treatment of manic-depressive psychosis for many years but their mechanism of action is not well understood. Many workers assume that the action of lithium on catecholamine metabolism and/or on electrolyte distribution

  9. Membrane depolarization regulates intracellular RANKL transport in non-excitable osteoblasts. (United States)

    Notomi, Takuya; Kuno, Miyuki; Hiyama, Akiko; Ezura, Yoichi; Honma, Masashi; Ishizuka, Toru; Ohura, Kiyoshi; Yawo, Hiromu; Noda, Masaki


    Parathyroid hormone (PTH) and 1α,25-dihydroxyvitamin D3 (VD3) are important factors in Ca(2+) homeostasis, and promote osteoclastogenesis by modulating receptor activator of nuclear factor kappa-B ligand (RANKL) mRNA expression. However, their contribution to RANKL intracellular transport (RANKLiT), including the trigger for RANKL lysosomal vesicle (RANKL-lv) fusion to the cell membrane, is unclear. In neurons, depolarization of membrane potential increases the intracellular Ca(2+) level ([Ca(2+)]i) and promotes neurotransmitter release via fusion of the synaptic vesicles to the cell membrane. To determine whether membrane depolarization also regulates cellular processes such as RANKLiT in MC3T3-E1 osteoblasts (OBs), we generated a light-sensitive OB cell line and developed a system for altering their membrane potential via delivery of a blue light stimulus. In the membrane fraction of RANKL-overexpressing OBs, PTH and VD3 increased the membrane-bound RANKL (mbRANKL) level at 10 min after application without affecting the mRNA expression level, and depolarized the cell membrane while transiently increasing [Ca(2+)]i. In our novel OB line stably expressing the channelrhodopsin-wide receiver, blue light-induced depolarization increased the mbRANKL level, which was reversed by treatment of blockers for L-type voltage-gated Ca(2+) channels and Ca(2+) release from the endoplasmic reticulum. In co-cultures of osteoclast precursor-like RAW264.7 cells and light-sensitive OBs overexpressing RANKL, light stimulation induced an increase in tartrate-resistant acid phosphatase activity and promoted osteoclast differentiation. These results indicate that depolarization of the cell membrane is a trigger for RANKL-lv fusion to the membrane and that membrane potential contributes to the function of OBs. In addition, the non-genomic action of VD3-induced RANKL-lv fusion included the membrane-bound VD3 receptor (1,25D3-MARRS receptor). Elucidating the mechanism of RANKLiT regulation

  10. Phospho-Caveolin-1 Mediates Integrin-Regulated Membrane Domain Internalisation (United States)

    del Pozo, Miguel A.; Alderson, Nazilla B.; Grande-García, Araceli; Balasubramanian, Nagaraj; Schwartz, Martin A.; Kiosses, William B.; Anderson, Richard G.W.


    Growth of normal cells is anchorage-dependent because signalling through multiple pathways including Erk, PI 3-kinase and Rac requires integrin-mediated cell adhesion 1. Components of these pathways localize to low density, cholesterol-rich domains in the plasma membrane named “lipid rafts” 2,3 or “cholesterol enriched membrane microdomains” (CEMM) 4. We previously reported that integrin-mediated adhesion regulates CEMM trafficking such that cell detachment from the extracellular matrix (ECM) triggers CEMM internalisation and clearance from the plasma membrane 5. We now report that this internalisation is mediated by dynamin-2 and caveolin-1. Internalisation requires phosphorylation of caveolin-1 on tyrosine 14. A shift in localisation of phospho-caveolin-1 from focal adhesions to caveolae induces CEMM internalisation upon cell detachment, which mediates inhibition of Erk, PI 3-kinase and Rac. These data define a novel molecular mechanism for growth and tumour suppression by caveolin-1. PMID:16113676

  11. Plasma Membrane Targeting of Protocadherin 15 Is Regulated by the Golgi-Associated Chaperone Protein PIST

    Directory of Open Access Journals (Sweden)

    Hongyun Nie


    Full Text Available Protocadherin 15 (PCDH15 is a core component of hair cell tip-links and crucial for proper function of inner ear hair cells. Mutations of PCDH15 gene cause syndromic and nonsyndromic hearing loss. At present, the regulatory mechanisms responsible for the intracellular transportation of PCDH15 largely remain unknown. Here we show that PIST, a Golgi-associated, PDZ domain-containing protein, interacts with PCDH15. The interaction is mediated by the PDZ domain of PIST and the C-terminal PDZ domain-binding interface (PBI of PCDH15. Through this interaction, PIST retains PCDH15 in the trans-Golgi network (TGN and reduces the membrane expression of PCDH15. We have previously showed that PIST regulates the membrane expression of another tip-link component, cadherin 23 (CDH23. Taken together, our finding suggests that PIST regulates the intracellular trafficking and membrane targeting of the tip-link proteins CDH23 and PCDH15.

  12. Selectivity of Direct Methanol Fuel Cell Membranes. (United States)

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


    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).

  13. Selectivity of Direct Methanol Fuel Cell Membranes (United States)

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


    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

  14. Selectivity of Direct Methanol Fuel Cell Membranes

    Directory of Open Access Journals (Sweden)

    Antonino S. Aricò


    Full Text Available 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.

  15. Proton exchange membrane fuel cells modeling

    CERN Document Server

    Gao, Fengge; Miraoui, Abdellatif


    The fuel cell is a potential candidate for energy storage and conversion in our future energy mix. It is able to directly convert the chemical energy stored in fuel (e.g. hydrogen) into electricity, without undergoing different intermediary conversion steps. In the field of mobile and stationary applications, it is considered to be one of the future energy solutions.Among the different fuel cell types, the proton exchange membrane (PEM) fuel cell has shown great potential in mobile applications, due to its low operating temperature, solid-state electrolyte and compactness.This book pre

  16. Interactions of Model Cell Membranes with Nanoparticles (United States)

    D'Angelo, S. M.; Camesano, T. A.; Nagarajan, R.


    The same properties that give nanoparticles their enhanced function, such as high surface area, small size, and better conductivity, can also alter the cytotoxicity of nanomaterials. Ultimately, many of these nanomaterials will be released into the environment, and can cause cytotoxic effects to environmental bacteria, aquatic organisms, and humans. Previous results from our laboratory suggest that nanoparticles can have a detrimental effect on cells, depending on nanoparticle size. It is our goal to characterize the properties of nanomaterials that can result in membrane destabilization. We tested the effects of nanoparticle size and chemical functionalization on nanoparticle-membrane interactions. Gold nanoparticles at 2, 5,10, and 80 nm were investigated, with a concentration of 1.1x1010 particles/mL. Model cell membranes were constructed of of L-α-phosphatidylcholine (egg PC), which has negatively charged lipid headgroups. A quartz crystal microbalance with dissipation (QCM-D) was used to measure frequency changes at different overtones, which were related to mass changes corresponding to nanoparticle interaction with the model membrane. In QCM-D, a lipid bilayer is constructed on a silicon dioxide crystal. The crystals, oscillate at different harmonic frequencies depending upon changes in mass or energy dissipation. When mass is added to the crystal surface, such as through addition of a lipid vesicle solution, the frequency change decreases. By monitoring the frequency and dissipation, we could verify that a supported lipid bilayer (SLB) formed on the silica surface. After formation of the SLB, the nanoparticles can be added to the system, and the changes in frequency and dissipation are monitored in order to build a mechanistic understanding of nanoparticle-cell membrane interactions. For all of the smaller nanoparticles (2, 5, and 10 nm), nanoparticle addition caused a loss of mass from the lipid bilayer, which appears to be due to the formation of holes

  17. A study for the research trends of membranes for proton exchange membrane fuel cells

    International Nuclear Information System (INIS)

    Sener, T.


    'Full text:' A single PEM fuel cell is comprised of a membrane electrode assembly, two bipolar plates and two fields. Membrane electrode assembly is the basic component of PEM fuel cell due to its cost and function, and it consists a membrane sandwiched between two electrocatalyst layers/electrodes and two gas diffusion layers. Increasing the PEM fuel cell operation temperature from 80 o C to 150-200 o C will prevent electrocatalysts CO poisoning and increase the fuel cell performance. Therefore, membranes must have chemical and mechanical resistance and must keep enough water at high temperatures. The aim of membrane studies through fuel cell commercialization is to produce a less expensive thin membrane with high operation temperature, chemical and mechanical resistance and water adsorption capacity. Within this frame, alternative membrane materials, membrane electrode assembly manufacture and evaluation methods are being studied. In this paper, recent studies are reviewed to give a conclusion for research trends. (author)

  18. Diphtheria toxin translocation across cellular membranes is regulated by sphingolipids

    International Nuclear Information System (INIS)

    Spilsberg, Bjorn; Hanada, Kentaro; Sandvig, Kirsten


    Diphtheria toxin is translocated across cellular membranes when receptor-bound toxin is exposed to low pH. To study the role of sphingolipids for toxin translocation, both a mutant cell line lacking the first enzyme in de novo sphingolipid synthesis, serine palmitoyltransferase, and a specific inhibitor of the same enzyme, myriocin, were used. The serine palmitoyltransferase-deficient cell line (LY-B) was found to be 10-15 times more sensitive to diphtheria toxin than the genetically complemented cell line (LY-B/cLCB1) and the wild-type cell line (CHO-K1), both when toxin translocation directly across the plasma membrane was induced by exposing cells with surface-bound toxin to low pH, and when the toxin followed its normal route via acidified endosomes into the cytosol. Toxin binding was similar in these three cell lines. Furthermore, inhibition of serine palmitoyltransferase activity by addition of myriocin sensitized the two control cell lines (LY-B/cLCB1 and CHO-K1) to diphtheria toxin, whereas, as expected, no effect was observed in cells lacking serine palmitoyltransferase (LY-B). In conclusion, diphtheria toxin translocation is facilitated by depletion of membrane sphingolipids

  19. Fuel cell membranes and crossover prevention (United States)

    Masel, Richard I [Champaign, IL; York, Cynthia A [Newington, CT; Waszczuk, Piotr [White Bear Lake, MN; Wieckowski, Andrzej [Champaign, IL


    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.

  20. Phosphatidylinositol 3-phosphates-at the interface between cell signalling and membrane traffic. (United States)

    Marat, Andrea L; Haucke, Volker


    Phosphoinositides (PIs) form a minor class of phospholipids with crucial functions in cell physiology, ranging from cell signalling and motility to a role as signposts of compartmental membrane identity. Phosphatidylinositol 3-phosphates are present at the plasma membrane and within the endolysosomal system, where they serve as key regulators of both cell signalling and of intracellular membrane traffic. Here, we provide an overview of the metabolic pathways that regulate cellular synthesis of PI 3-phosphates at distinct intracellular sites and discuss the mechanisms by which these lipids regulate cell signalling and membrane traffic. Finally, we provide a framework for how PI 3-phosphate metabolism is integrated into the cellular network. © 2016 The Authors.

  1. Cell-substrate interaction with cell-membrane-stress dependent adhesion. (United States)

    Jiang, H; Yang, B


    Cell-substrate interaction is examined in a two-dimensional mechanics model. The cell and substrate are treated as a shell and an elastic solid, respectively. Their interaction through adhesion is treated using nonlinear springs. Compared to previous cell mechanics models, the present model introduces a cohesive force law that is dependent not only on cell-substrate distance but also on internal cell-membrane stress. It is postulated that a living cell would establish focal adhesion sites with density dependent on the cell-membrane stress. The formulated mechanics problem is numerically solved using coupled finite elements and boundary elements for the cell and the substrate, respectively. The nodes in the adhesion zone from either side are linked by the cohesive springs. The specific cases of a cell adhering to a homogeneous substrate and a heterogeneous bimaterial substrate are examined. The analyses show that the substrate stiffness affects the adhesion behavior significantly and regulates the direction of cell adhesion, in good agreement with the experimental results in the literature. By introducing a reactive parameter (i.e., cell-membrane stress) linking biological responses of a living cell to a mechanical environment, the present model offers a unified mechanistic vehicle for characterization and prediction of living cell responses to various kinds of mechanical stimuli including local extracellular matrix and neighboring cells. Copyright © 2011 Elsevier Ltd. All rights reserved.

  2. Salmonella Rapidly Regulates Membrane Permeability To Survive Oxidative Stress. (United States)

    van der Heijden, Joris; Reynolds, Lisa A; Deng, Wanyin; Mills, Allan; Scholz, Roland; Imami, Koshi; Foster, Leonard J; Duong, Franck; Finlay, B Brett


    The outer membrane (OM) of Gram-negative bacteria provides protection against toxic molecules, including reactive oxygen species (ROS). Decreased OM permeability can promote bacterial survival under harsh circumstances and protects against antibiotics. To better understand the regulation of OM permeability, we studied the real-time influx of hydrogen peroxide in Salmonella bacteria and discovered two novel mechanisms by which they rapidly control OM permeability. We found that pores in two major OM proteins, OmpA and OmpC, could be rapidly opened or closed when oxidative stress is encountered and that the underlying mechanisms rely on the formation of disulfide bonds in the periplasmic domain of OmpA and TrxA, respectively. Additionally, we found that a Salmonella mutant showing increased OM permeability was killed more effectively by treatment with antibiotics. Together, these results demonstrate that Gram-negative bacteria regulate the influx of ROS for defense against oxidative stress and reveal novel targets that can be therapeutically targeted to increase bacterial killing by conventional antibiotics. Pathogenic bacteria have evolved ways to circumvent inflammatory immune responses. A decrease in bacterial outer membrane permeability during infection helps protect bacteria from toxic molecules produced by the host immune system and allows for effective colonization of the host. In this report, we reveal molecular mechanisms that rapidly alter outer membrane pores and their permeability in response to hydrogen peroxide and oxidative stress. These mechanisms are the first examples of pores that are rapidly opened or closed in response to reactive oxygen species. Moreover, one of these mechanisms can be targeted to artificially increase membrane permeability and thereby increase bacterial killing by the antibiotic cefotaxime during in vitro experiments and in a mouse model of infection. We envision that a better understanding of the regulation of membrane

  3. An Unusual Prohibitin Regulates Malaria Parasite Mitochondrial Membrane Potential

    Directory of Open Access Journals (Sweden)

    Joachim Michael Matz


    Full Text Available Summary: Proteins of the stomatin/prohibitin/flotillin/HfIK/C (SPFH family are membrane-anchored and perform diverse cellular functions in different organelles. Here, we investigate the SPFH proteins of the murine malaria model parasite Plasmodium berghei, the conserved prohibitin 1, prohibitin 2, and stomatin-like protein and an unusual prohibitin-like protein (PHBL. The SPFH proteins localize to the parasite mitochondrion. While the conserved family members could not be deleted from the Plasmodium genome, PHBL was successfully ablated, resulting in impaired parasite fitness and attenuated virulence in the mammalian host. Strikingly, PHBL-deficient parasites fail to colonize the Anopheles vector because of complete arrest during ookinete development in vivo. We show that this arrest correlates with depolarization of the mitochondrial membrane potential (ΔΨmt. Our results underline the importance of SPFH proteins in the regulation of core mitochondrial functions and suggest that fine-tuning of ΔΨmt in malarial parasites is critical for colonization of the definitive host. : Matz et al. present an experimental genetics study of an unusual prohibitin-like protein in the malaria parasite and find that it regulates mitochondrial membrane polarity. Ablation of this protein causes almost complete mitochondrial depolarization in the mosquito vector, which, in turn, leads to a block in malaria parasite transmission. Keywords: Plasmodium berghei, malaria, SPFH, prohibitin, stomatin-like protein, mitochondrion, membrane potential, ookinete, transmission

  4. Emergent properties arising from the assembly of amphiphiles. Artificial vesicle membranes as reaction promoters and regulators. (United States)

    Walde, Peter; Umakoshi, Hiroshi; Stano, Pasquale; Mavelli, Fabio


    This article deals with artificial vesicles and their membranes as reaction promoters and regulators. Among the various molecular assemblies which can form in an aqueous medium from amphiphilic molecules, vesicle systems are unique. Vesicles compartmentalize the aqueous solution in which they exist, independent on whether the vesicles are biological vesicles (existing in living systems) or whether they are artificial vesicles (formed in vitro from natural or synthetic amphiphiles). After the formation of artificial vesicles, their aqueous interior (the endovesicular volume) may become - or may be made - chemically different from the external medium (the exovesicular solution), depending on how the vesicles are prepared. The existence of differences between endo- and exovesicular composition is one of the features on the basis of which biological vesicles contribute to the complex functioning of living organisms. Furthermore, artificial vesicles can be formed from mixtures of amphiphiles in such a way that the vesicle membranes become molecularly, compositionally and organizationally highly complex, similarly to the lipidic matrix of biological membranes. All the various properties of artificial vesicles as membranous compartment systems emerge from molecular assembly as these properties are not present in the individual molecules the system is composed of. One particular emergent property of vesicle membranes is their possible functioning as promoters and regulators of chemical reactions caused by the localization of reaction components, and possibly catalysts, within or on the surface of the membranes. This specific feature is reviewed and highlighted with a few selected examples which range from the promotion of decarboxylation reactions, the selective binding of DNA or RNA to suitable vesicle membranes, and the reactivation of fragmented enzymes to the regulation of the enzymatic synthesis of polymers. Such type of emergent properties of vesicle membranes may

  5. Membrane tension: A challenging but universal physical parameter in cell biology. (United States)

    Pontes, Bruno; Monzo, Pascale; Gauthier, Nils C


    The plasma membrane separates the interior of cells from the outside environment. The membrane tension, defined as the force per unit length acting on a cross-section of membrane, regulates many vital biological processes. In this review, we summarize the first historical findings and the latest advances, showing membrane tension as an important physical parameter in cell biology. We also discuss how this parameter must be better integrated and we propose experimental approaches for key unanswered questions. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. [Parotid basal cell adenoma of membranous type]. (United States)

    Farah-Klibi, Faten; Ferchiou, Malek; Kourda, Jihène; El Amine, Olfa; Ferjaoui, Mohamed; Ben Jilani, Sarrah; Zermani, Rachida


    Basal cell adenoma (BCA) is a rare benign neoplasm characterized by the basaloid appearance of the tumour cells and the lack of myxo-chondroid stromal component present in pleomorphic adenoma. We report a case of basal cell adenoma of membranous type, highly suspected of malignancy because of the presence of mediastinal lymph nodes and pulmonary nodules which finally were related to an associated sarcoidosis. Our patient was an 80-year-old woman who presented a swelling of the right parotid two years ago. The clinical examination revealed a solid, indolent and mobile mass. A chest radiography noted mediastinal lymph nodes. The CT-scan confirmed the presence of mediastinal and tracheal lymph nodes with pulmonary nodules. So the diagnosis of metastatic malignant salivary gland tumor was suspected. Finally, the histological examination concluded to a basal cell adenoma of membranous type with sarcoidosis granulomas in the parotid and in the lymph nodes. The BCA is a benign tumor located generally in the parotid gland. When the malignancy is suspected, like in our case, this tumor must be differentiated from the basal cell adenocarcinoma using histological criteria.

  7. NCAM regulates cell motility

    DEFF Research Database (Denmark)

    Prag, Søren; Lepekhin, Eugene A; Kolkova, Kateryna


    Cell migration is required during development of the nervous system. The regulatory mechanisms for this process, however, are poorly elucidated. We show here that expression of or exposure to the neural cell adhesion molecule (NCAM) strongly affected the motile behaviour of glioma cells independe...

  8. Membrane Purification Cell for Aluminum Recycling

    Energy Technology Data Exchange (ETDEWEB)

    David DeYoung; James Wiswall; Cong Wang


    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

  9. Phospholipid synthesis participates in the regulation of diacylglycerol required for membrane trafficking at the Golgi complex. (United States)

    Sarri, Elisabet; Sicart, Adrià; Lázaro-Diéguez, Francisco; Egea, Gustavo


    The lipid metabolite diacylglycerol (DAG) is required for transport carrier biogenesis at the Golgi, although how cells regulate its levels is not well understood. Phospholipid synthesis involves highly regulated pathways that consume DAG and can contribute to its regulation. Here we altered phosphatidylcholine (PC) and phosphatidylinositol synthesis for a short period of time in CHO cells to evaluate the changes in DAG and its effects in membrane trafficking at the Golgi. We found that cellular DAG rapidly increased when PC synthesis was inhibited at the non-permissive temperature for the rate-limiting step of PC synthesis in CHO-MT58 cells. DAG also increased when choline and inositol were not supplied. The major phospholipid classes and triacylglycerol remained unaltered for both experimental approaches. The analysis of Golgi ultrastructure and membrane trafficking showed that 1) the accumulation of the budding vesicular profiles induced by propanolol was prevented by inhibition of PC synthesis, 2) the density of KDEL receptor-containing punctated structures at the endoplasmic reticulum-Golgi interface correlated with the amount of DAG, and 3) the post-Golgi transport of the yellow fluorescent temperature-sensitive G protein of stomatitis virus and the secretion of a secretory form of HRP were both reduced when DAG was lowered. We confirmed that DAG-consuming reactions of lipid synthesis were present in Golgi-enriched fractions. We conclude that phospholipid synthesis pathways play a significant role to regulate the DAG required in Golgi-dependent membrane trafficking.

  10. Developmentally regulated GTP-binding protein 2 is required for stabilization of Rac1-positive membrane tubules. (United States)

    Mani, Muralidharan; Lee, Unn Hwa; Yoon, Nal Ae; Yoon, Eun Hye; Lee, Byung Ju; Cho, Wha Ja; Park, Jeong Woo


    Previously we have reported that developmentally regulated GTP-binding protein 2 (DRG2) localizes on Rab5 endosomes and plays an important role in transferrin (Tfn) recycling. We here identified DRG2 as a key regulator of membrane tubule stability. At 30 min after Tfn treatment, DRG2 localized to membrane tubules which were enriched with phosphatidylinositol 4-monophosphate [PI(4)P] and did not contain Rab5. DRG2 interacted with Rac1 more strongly with GTP-bound Rac1 and tubular localization of DRG2 depended on Rac1 activity. DRG2 depletion led to destabilization of membrane tubules, while ectopic expression of DRG2 rescued the stability of the membrane tubules in DRG2-depleted cells. Our results reveal a novel mechanism for regulation of membrane tubule stability mediated by DRG2. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Annexin A4 and A6 induce membrane curvature and constriction during cell membrane repair

    DEFF Research Database (Denmark)

    Boye, Theresa Louise; Maeda, Kenji; Pezeshkian, Weria


    that annexin A4 binds to artificial membranes and generates curvature force initiated from free edges, whereas annexin A6 induces constriction force. In cells, plasma membrane injury and Ca2+ influx recruit annexin A4 to the vicinity of membrane wound edges where its homo-trimerization leads to membrane......Efficient cell membrane repair mechanisms are essential for maintaining membrane integrity and thus for cell life. Here we show that the Ca2+- and phospholipid-binding proteins annexin A4 and A6 are involved in plasma membrane repair and needed for rapid closure of micron-size holes. We demonstrate...... curvature near the edges. We propose that curvature force is utilized together with annexin A6-mediated constriction force to pull the wound edges together for eventual fusion. We show that annexin A4 can counteract various plasma membrane disruptions including holes of several micrometers indicating...

  12. Membrane fluidity adjustments in ethanol-stressed Oenococcus oeni cells

    NARCIS (Netherlands)

    Silveira, da M.G.; Golovina, E.A.; Hoekstra, F.A.; Rombouts, F.M.; Abee, T.


    The effect of ethanol on the cytoplasmic membrane of Oenococcus oeni cells and the role of membrane changes in the acquired tolerance to ethanol were investigated. Membrane tolerance to ethanol was defined as the resistance to ethanol-induced leakage of preloaded carboxyfluorescein (cF) from cells.

  13. Membrane rafts of the human red blood cell. (United States)

    Ciana, Annarita; Achilli, Cesare; Minetti, Giampaolo


    The cell type of election for the study of cell membranes, the mammalian non-nucleated erythrocyte, has been scarcely considered in the research of membrane rafts of the plasma membrane. However, detergent-resistant-membranes (DRM) were actually first described in human erythrocytes, as a fraction resisting solubilization by the nonionic detergent Triton X-100. These DRMs were insoluble entities of high density, easily pelleted by centrifugation, as opposed to the now accepted concept of lipid raft-like membrane fractions as material floating in low-density regions of sucrose gradients. The present article reviews the available literature on membrane rafts/DRMs in human erythrocytes from an historical point of view, describing the experiments that provided the solution to the above described discrepancy and suggesting possible avenue of research in the field of membrane rafts that, moving from the most studied model of living cell membrane, the erythrocyte's, could be relevant also for other cell types.

  14. Pharmacological targeting of membrane rigidity: implications on cancer cell migration and invasion

    International Nuclear Information System (INIS)

    Braig, Simone; Stoiber, Katharina; Zahler, Stefan; Vollmar, Angelika M


    The invasive potential of cancer cells strongly depends on cellular stiffness, a physical quantity that is not only regulated by the mechanical impact of the cytoskeleton but also influenced by the membrane rigidity. To analyze the specific role of membrane rigidity in cancer progression, we treated cancer cells with the Acetyl-CoA carboxylase inhibitor Soraphen A and revealed an alteration of the phospholipidome via mass spectrometry. Migration, invasion, and cell death assays were employed to relate this alteration to functional consequences, and a decrease of migration and invasion without significant impact on cell death has been recorded. Fourier fluctuation analysis of giant plasma membrane vesicles showed that Soraphen A increases membrane rigidity of carcinoma cell membranes. Mechanical measurements of the creep deformation response of whole intact cells were performed using the optical stretcher. The increase in membrane rigidity was observed in one cell line without changing the creep deformation response indicating no restructuring of the cytoskeleton. These data indicate that the increase of membrane rigidity alone is sufficient to inhibit invasiveness of cancer cells, thus disclosing the eminent role of membrane rigidity in migratory processes. (paper)

  15. Dividing Cells Regulate Their Lipid Composition and Localization (United States)

    Atilla-Gokcumen, G. Ekin; Muro, Eleonora; Relat-Goberna, Josep; Sasse, Sofia; Bedigian, Anne; Coughlin, Margaret L.; Garcia-Manyes, Sergi; Eggert, Ulrike S.


    Summary Although massive membrane rearrangements occur during cell division, little is known about specific roles that lipids might play in this process. We report that the lipidome changes with the cell cycle. LC-MS-based lipid profiling shows that 11 lipids with specific chemical structures accumulate in dividing cells. Using AFM, we demonstrate differences in the mechanical properties of live dividing cells and their isolated lipids relative to nondividing cells. In parallel, systematic RNAi knockdown of lipid biosynthetic enzymes identified enzymes required for division, which highly correlated with lipids accumulated in dividing cells. We show that cells specifically regulate the localization of lipids to midbodies, membrane-based structures where cleavage occurs. We conclude that cells actively regulate and modulate their lipid composition and localization during division, with both signaling and structural roles likely. This work has broader implications for the active and sustained participation of lipids in basic biology. PMID:24462247

  16. Indole prevents Escherichia coli cell division by modulating membrane potential


    Chimerel, Catalin; Field, Christopher M.; Pi?ero-Fernandez, Silvia; Keyser, Ulrich F.; Summers, David K.


    Indole is a bacterial signalling molecule that blocks E. coli cell division at concentrations of 3?5?mM. We have shown that indole is a proton ionophore and that this activity is key to the inhibition of division. By reducing the electrochemical potential across the cytoplasmic membrane of E. coli, indole deactivates MinCD oscillation and prevents formation of the FtsZ ring that is a prerequisite for division. This is the first example of a natural ionophore regulating a key biological proces...

  17. A Quaternary Polybenzimidazole Membrane for Intermediate Temperature Polymer Electrolyte Membrane Fuel Cells

    DEFF Research Database (Denmark)

    Xu, C.; Scott, K.; Li, Qingfeng


    A quaternary ammonium polybenzimidazole (QPBI) membrane was synthesized for applications in intermediate temperature (100–200 °C) hydrogen fuel cells. The QPBI membrane was imbibed with phosphoric acid to provide suitable proton conductivity. The proton conductivity of the membrane was 0.051 S cm–1...

  18. Plasma membranes from insect midgut cells

    Directory of Open Access Journals (Sweden)

    Walter R. Terra


    Full Text Available Plasma membranes from insect midgut cells are separated into apical and basolateral domains. The apical domain is usually modified into microvilli with a molecular structure similar to other animals. Nevertheless, the microvillar structure should differ in some insects to permit the traffic inside them of secretory vesicles that may budd laterally or pinch-off from the tips of microvilli. Other microvillar modifications are associated with proton-pumping or with the interplay with an ensheathing lipid membrane (the perimicrovilllar membrane observed in the midgut cells of hemipterans (aphids and bugs. The perimicrovillar membranes are thought to be involved in amino acid absorption from diluted diets. The microvillar and perimicrovillar membranes have densities (and protein content that depend on the insect taxon. The role played by the microvillar and perimicrovillar proteins in insect midgut physiology is reviewed here trying to provide a coherent picture of data and highlighting further research areas.As membranas plasmáticas das células intestinais dos insetos apresentam um domínio apical e outro basal. O domínio apical é geralmente modificado em microvilosidades com organização molecular similar a de outros animais, embora possam diferir naqueles insetos que apresentam vesículas secretoras em trânsito que brotam lateralmente ou destacam-se das extremidades das microvilosidades. Outras modificações microvilares estão associadas a bombeamento de prótons ou a interrelações com uma membrana lipídica (a membrana perimicrovilar que reveste as microvilosidades de células intestinais de hemípteros (pulgões e percevejos. Admite-se que as membranas perimicrovilares estejam envolvidas na absorção de aminoácidos a partir de dietas diluídas. As membranas microvilares e perimicrovilares tem densidades distintas (e conteúdo protéico que dependem do táxon do inseto. O papel desempenhado pelas proteínas microvilares e

  19. The exocyst at the interface between cytoskeleton and membranes in eukaryotic cells

    Directory of Open Access Journals (Sweden)

    Lukas eSynek


    Full Text Available Delivery and final fusion of the secretory vesicles with the relevant target membrane are hierarchically organized and reciprocally interconnected multi-step processes involving not only specific protein-protein interactions, but also specific protein-phospholipid interactions. The exocyst was discovered as a tethering complex mediating initial encounter of arriving exocytic vesicles with the plasma membrane. The exocyst complex is regulated by Rab and Rho small GTPases, resulting in docking of exocytic vesicles to the plasma membrane and finally their fusion mediated by specific SNARE complexes. In model Opisthokont cells, the exocyst was shown to directly interact with both microtubule and microfilament cytoskeleton and related motor proteins as well as with the plasma membrane via phosphatidylinositol 4,5-bisphosphate specific binding, which directly affects cortical cytoskeleton and plasma membrane dynamics. Here we summarize the current knowledge on exocyst-cytoskeleton-plasma membrane interactions in order to open a perspective for future research in this area in plant cells.

  20. Biophysics of Cell Membrane Lipids in Cancer Drug Resistance: Implications for Drug Transport and Drug Delivery with Nanoparticles (United States)

    Peetla, Chiranjeevi; Vijayaraghavalu, Sivakumar; Labhasetwar, Vinod


    In this review, we focus on the biophysics of cell membrane lipids, particularly when cancers develop acquired drug resistance, and how biophysical changes in resistant cell membrane influence drug transport and nanoparticle-mediated drug delivery. Recent advances in membrane lipid research show the varied roles of lipids in regulating membrane P-glycoprotein function, membrane trafficking, apoptotic pathways, drug transport, and endocytic functions, particularly endocytosis, the primary mechanism of cellular uptake of nanoparticle-based drug delivery systems. Since acquired drug resistance alters lipid biosynthesis, understanding the role of lipids in cell membrane biophysics and its effect on drug transport is critical for developing effective therapeutic and drug delivery approaches to overcoming drug resistance. Here we discuss novel strategies for (a) modulating the biophysical properties of membrane lipids of resistant cells to facilitate drug transport and regain endocytic function and (b) developing effective nanoparticles based on their biophysical interactions with membrane lipids to enhance drug delivery and overcome drug resistance. PMID:24055719

  1. Preparation and characterization of Nafion/titanium dioxide nanocomposite membranes for proton exchange membrane fuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Eroglu, Inci; Devrim, Yilser; Erkan, Serdar [Middle East Technical Univ., Ankara (Turkey). Dept. of Chemical Engineering; Bac, Nurcan [Yeditepe Univ., Istanbul (Turkey). Dept. of Chemical Engineering


    In the present study, Nafion/Titanium dioxide (TiO{sub 2}) nanocomposite membranes for use in proton exchange membrane fuel cells (PEMFC) were investigated. Nafion/TiO{sub 2} membranes were prepared using the recasting procedure. The composite membranes have been characterized by thermal analysis, XRD, SEM, proton conductivity measurements and single cell performance. Thermal analysis results showed that the composite membranes have good thermal properties. The introduction of the inorganic filler supplies the composite membrane with a good thermal resistance. The physico-chemical properties studied by means of scanning electron microscopy (SEM) and X-ray diffraction (XRD) techniques have proved the uniform and homogeneous distribution of TiO{sub 2} and the consequent enhancement of crystalline character of these membranes. The energy dispersive spectra (EDS) analysis indicated that the distribution of Ti element on the surface of the composite membrane was uniform. Performances of fabricated Membrane electrode assembly (MEA)'s measured via the PEMFC test station built at METU Fuel Cell Technology Laboratory. A single cell with a 5 cm{sup 2} active area was used in the experiments. These results should be conducive to the preparation of membranes suitable for PEMFC. We believe that Nafion/TiO{sub 2} nano composite membranes have good prospects for use in PEMFC. (orig.)

  2. Channels in cell membranes and synchrotron radiation

    International Nuclear Information System (INIS)

    Yan Xiaohui; Tian Liang; Zhang Xinyi


    For long time a lot of scientists have devoted to study how matter, such as water molecules and K + , Na + , Ca 2+ , Cl - ions, move through cell membranes and complete the matter exchange between the inside and outside of cells. Peter Agre discovered and characterized the first water channel protein in 1988 and Roderick MacKinnon elucidated the structural and mechanistic basis for ion channel function in 1998. These achievements have made it possible for us to 'see' these exquisitely designed molecular machines in action at the atomic level. The Nobel Prize in Chemistry for 2003 is shared between these two scientists. In determining the high resolution 3D structure of these channels, the synchrotron X-ray diffraction plays an important role

  3. Membrane plasmalogen composition and cellular cholesterol regulation: a structure activity study

    Directory of Open Access Journals (Sweden)

    Su-Myat Khine K


    Full Text Available Abstract Background Disrupted cholesterol regulation leading to increased circulating and membrane cholesterol levels is implicated in many age-related chronic diseases such as cardiovascular disease (CVD, Alzheimer's disease (AD, and cancer. In vitro and ex vivo cellular plasmalogen deficiency models have been shown to exhibit impaired intra- and extra-cellular processing of cholesterol. Furthermore, depleted brain plasmalogens have been implicated in AD and serum plasmalogen deficiencies have been linked to AD, CVD, and cancer. Results Using plasmalogen deficient (NRel-4 and plasmalogen sufficient (HEK293 cells we investigated the effect of species-dependent plasmalogen restoration/augmentation on membrane cholesterol processing. The results of these studies indicate that the esterification of cholesterol is dependent upon the amount of polyunsaturated fatty acid (PUFA-containing ethanolamine plasmalogen (PlsEtn present in the membrane. We further elucidate that the concentration-dependent increase in esterified cholesterol observed with PUFA-PlsEtn was due to a concentration-dependent increase in sterol-O-acyltransferase-1 (SOAT1 levels, an observation not reproduced by 3-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA reductase inhibition. Conclusion The present study describes a novel mechanism of cholesterol regulation that is consistent with clinical and epidemiological studies of cholesterol, aging and disease. Specifically, the present study describes how selective membrane PUFA-PlsEtn enhancement can be achieved using 1-alkyl-2-PUFA glycerols and through this action reduce levels of total and free cholesterol in cells.

  4. Sulfonated carbon black-based composite membranes for fuel cell ...

    Indian Academy of Sciences (India)

    Pristine and composite membranes prepared from SPEEK82 decomposed completely in <1 h, which is undesirable for fuel cell applications. SPEEK60 membrane having wt% of 0.25–0.5 with S–C particles led to higher proton conductivity than that of pristine membrane. No positive effect was observed on the properties of ...

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

    International Nuclear Information System (INIS)

    Haryadi,; Sugianto, D.; Ristopan, E.


    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

  6. Application of the nanocomposite membrane as electrolyte of proton exchange membrane fuel cell

    International Nuclear Information System (INIS)



    Hydrogen fuel cells proton exchange membrane fuel cell (PEMFC) is currently still in development and commercialization. Several barriers to the commercialization of these Nafion membrane as electrolyte is its very sensitive to humidity fluctuation. Nafion must be modified by making a composite Nafion-SiO 2 -HPA to increase electrolyte resistance against humidity fluctuations during the cell used. Research carried out by mixing Nafion solution with Tetra Ethoxy Ortho Silicate (TEOS) and conductive materials is phosphotungstic acid (PWA) by varying the ratio of Nafion, TEOS and PWA. The membrane is produced by heating a mixture of Nafion, TEOS and PWA by varying the evaporation temperature, time and annealing temperature to obtain the transparent membrane. The resulting membrane was analyzed its physical, chemical and electrochemical properties by applying the membrane as electrolyte of PEMFC at various humidity and temperature of operation. The results showed that at low temperatures (30-90 °C) and high humidity at 100 % RH, pure Nafion membrane is better than composite membrane (Nafion-SiO 2 -PWA), but at low humidity condition composite membrane is better than the pure Nafion membrane. It can be concluded that the composite membranes of (Nafion-SiO 2 -PWA) can be used as electrolyte of PEMFC operated at low humidity (40 % RH) and temperature between (30-90 °C). (author)

  7. Physiology and Regulation of Calcium Channels in Stomatal Guard Cells

    Energy Technology Data Exchange (ETDEWEB)

    Schroeder, Julian I.


    Stomatal pores in the epidermis of leaves regulate the diffusion of CO2 into leaves for photosynthetic carbon fixation and control water loss of plants during drought periods. Guard cells sense CO2, water status, light and other environmental conditions to regulate stomatal apertures for optimization of CO2 intake and plant growth under drought stress. The cytosolic second messenger calcium contributes to stomatal movements by transducing signals and regulating ion channels in guard cells. Studies suggest that both plasma membrane Ca2+ influx channels and vacuolar/organellar Ca2+ release channels contribute to ABA-induced Ca2+ elevations in guard cells. Recent research in the P.I.'s laboratory has led to identification of a novel major cation-selective Ca2+-permeable influx channel (Ica) in the plasma membrane of Arabidopsis guard cells. These advances will allow detailed characterization of Ica plasma membrane Ca2+ influx channels in guard cells. The long term goal of this research project is to gain a first detailed characterization of these novel plasma membrane Ca2+-permeable channel currents in Arabidopsis guard cells. The proposed research will investigate the hypothesis that Ica represents an important Ca2+ influx pathway for ABA and CO2 signal transduction in Arabidopsis guard cells. These studies will lead to elucidation of key signal transduction mechanisms by which plants balance CO2 influx into leaves and transpirational water loss and may contribute to future strategies for manipulating gas exchange for improved growth of crop plants and for biomass production.

  8. Regulators of Tfh cell differentiation

    Directory of Open Access Journals (Sweden)

    Gajendra Motiram Jogdand


    Full Text Available The follicular helper T (Tfh cells help is critical for activation of B cells, antibody class switching and germinal center formation. The Tfh cells are characterized by the expression of CXCR5, ICOS, PD-1, Bcl-6, and IL-21. They are involved in clearing infections and are adversely linked with autoimmune diseases and also have a role in viral replication as well as clearance. Tfh cells are generated from naïve CD4 T cells with sequential steps involving cytokine signaling (IL-21, IL-6, IL-12, activin A, migration and positioning in the germinal center by CXCR5, surface receptors (ICOS/ICOSL, SAP/SLAM as well as transcription factor (Bcl-6, c-Maf, STAT3 signaling and repressor miR155. On the other hand Tfh generation is negatively regulated at specific steps of Tfh generation by specific cytokine (IL-2, IL-7, surface receptor (PD-1, CTLA-4, transcription factors Blimp-1, STAT5, T-bet, KLF-2 signaling and repressor miR 146a. Interestingly, miR 17-92 and FOXO1 acts as a positive as well as a negative regulator of Tfh differentiation depending on the time of expression and disease specificity. Tfh cells are also generated from the conversion of other effector T cells as exemplified by Th1 cells converting into Tfh during viral infection. The mechanistic details of effector T cells conversion into Tfh are yet to be clear. To manipulate Tfh cells for therapeutic implication and or for effective vaccination strategies, it is important to know positive and negative regulators of Tfh generation. Hence, in this review we have highlighted and interlinked molecular signaling from cytokines, surface receptors, transcription factors, ubiquitin Ligase and miRNA as positive and negative regulators for Tfh differentiation.

  9. Protein kinase A regulates 3-phosphatidylinositide dynamics during platelet-derived growth factor-induced membrane ruffling and chemotaxis. (United States)

    Deming, Paula B; Campbell, Shirley L; Baldor, Linda C; Howe, Alan K


    Spatial regulation of the cAMP-dependent protein kinase (PKA) is required for chemotaxis in fibroblasts; however, the mechanism(s) by which PKA regulates the cell migration machinery remain largely unknown. Here we report that one function of PKA during platelet-derived growth factor (PDGF)-induced chemotaxis was to promote membrane ruffling by regulating phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) dynamics. Inhibition of PKA activity dramatically altered membrane dynamics and attenuated formation of peripheral membrane ruffles in response to PDGF. PKA inhibition also significantly decreased the number and size of PIP(3)-rich membrane ruffles in response to uniform stimulation and to gradients of PDGF. This ruffling defect was quantified using a newly developed method, based on computer vision edge-detection algorithms. PKA inhibition caused a marked attenuation in the bulk accumulation of PIP(3) following PDGF stimulation, without effects on PI3-kinase (PI3K) activity. The deficits in PIP(3) dynamics correlated with a significant inhibition of growth factor-induced membrane recruitment of endogenous Akt and Rac activation in PKA-inhibited cells. Simultaneous inhibition of PKA and Rac had an additive inhibitory effect on growth factor-induced ruffling dynamics. Conversely, the expression of a constitutively active Rac allele was able to rescue the defect in membrane ruffling and restore the localization of a fluorescent PIP(3) marker to membrane ruffles in PKA-inhibited cells, even in the absence of PI3K activity. These data demonstrate that, like Rac, PKA contributes to PIP(3) and membrane dynamics independently of direct regulation of PI3K activity and suggest that modulation of PIP(3)/3-phosphatidylinositol (3-PI) lipids represents a major target for PKA in the regulation of PDGF-induced chemotactic events.

  10. Independent regulation of reovirus membrane penetration and apoptosis by the mu1 phi domain.

    Directory of Open Access Journals (Sweden)

    Pranav Danthi


    Full Text Available Apoptosis plays an important role in the pathogenesis of reovirus encephalitis. Reovirus outer-capsid protein mu1, which functions to penetrate host cell membranes during viral entry, is the primary regulator of apoptosis following reovirus infection. Ectopic expression of full-length and truncated forms of mu1 indicates that the mu1 phi domain is sufficient to elicit a cell death response. To evaluate the contribution of the mu1 phi domain to the induction of apoptosis following reovirus infection, phi mutant viruses were generated by reverse genetics and analyzed for the capacity to penetrate cell membranes and elicit apoptosis. We found that mutations in phi diminish reovirus membrane penetration efficiency by preventing conformational changes that lead to generation of key reovirus entry intermediates. Independent of effects on membrane penetration, amino acid substitutions in phi affect the apoptotic potential of reovirus, suggesting that phi initiates apoptosis subsequent to cytosolic delivery. In comparison to wild-type virus, apoptosis-defective phi mutant viruses display diminished neurovirulence following intracranial inoculation of newborn mice. These results indicate that the phi domain of mu1 plays an important regulatory role in reovirus-induced apoptosis and disease.

  11. Integrated functions of membrane-type 1 matrix metalloproteinase in regulating cancer malignancy: Beyond a proteinase. (United States)

    Sakamoto, Takeharu; Seiki, Motoharu


    Membrane-type 1 matrix metalloproteinase (MT1-MMP) is expressed in different types of invasive and proliferative cells, including cancer cells and stromal cells. MT1-MMP cleaves extracellular matrix proteins, membrane proteins and other pericellular proteins, thereby changing the cellular microenvironment and regulating signal activation. Critical roles of protease activity in cancer cell proliferation, invasion and metastasis have been demonstrated by many groups. MT1-MMP also has a non-protease activity in that it inhibits the oxygen-dependent suppression of hypoxia-inducible factors (HIFs) via Munc18-1-interacting protein 3 (Mint3) and thereby enhances the expression of HIF target genes. Elevated HIF activity in MT1-MMP-expressing cancer cells is a fundamental mechanism underlying the Warburg effect, a well-known phenomenon where malignant cancer cells exhibit a higher rate of glucose metabolism. Because specific intervention of HIF activation by MT1-MMP suppresses tumor formation by cancer cells in mice, both the proteolytic and non-proteolytic activities of MT1-MMP are important for tumor malignancy and function in an integrated manner. In this review, we summarize recent findings relating to how MT1-MMP activates HIF and its effects on cancer cells and stromal cells. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  12. Nanoscale spin sensing in artificial cell membranes

    International Nuclear Information System (INIS)

    Simpson David


    The use of the nitrogen-vacancy (NV) centre in diamond as a single spin sensor or magnetometer has attracted considerable interest in recent years because of its unique combination of sensitivity, nanoscale resolution, and optical initialisation and readout at room temperature. Nanodiamonds in particular hold great promise as an optical magnetometer probe for bio applications. In this work we employ nanodiamonds containing single NV spins to detect freely diffusing Mn2+ ions by detecting changes in the transverse relaxation time (T2) of the single spin probe. We also report the detection of gadolinium spin labels present in an artificial cell membrane by measuring changes in the longitudinal relaxation time (T1) of the probe. (author)

  13. Plasma membrane microdomains regulate turnover of transport proteins in yeast

    Czech Academy of Sciences Publication Activity Database

    Grossmann, G.; Malínský, Jan; Stahlschmidt, W.; Loibl, M.; Weig-Meckl, I.; Frommer, W.B.; Opekarová, Miroslava; Tanner, W.


    Roč. 183, č. 6 (2008), s. 1075-1088 ISSN 0021-9525 R&D Projects: GA ČR GA204/06/0009; GA ČR GA204/07/0133; GA ČR GC204/08/J024 Institutional research plan: CEZ:AV0Z50390703; CEZ:AV0Z50200510 Keywords : Lithium acetate * Membrane compartment of Can1 * Monomeric red fluorescent protein Subject RIV: EA - Cell Biology Impact factor: 9.120, year: 2008

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

    Energy Technology Data Exchange (ETDEWEB)

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


    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.

  15. Proteomic analysis of plasma membranes isolated from undifferentiated and differentiated HepaRG cells

    Directory of Open Access Journals (Sweden)

    Sokolowska Izabela


    Full Text Available Abstract Liver infection with hepatitis B virus (HBV, a DNA virus of the Hepadnaviridae family, leads to severe disease, such as fibrosis, cirrhosis and hepatocellular carcinoma. The early steps of the viral life cycle are largely obscure and the host cell plasma membrane receptors are not known. HepaRG is the only proliferating cell line supporting HBV infection in vitro, following specific differentiation, allowing for investigation of new host host-cell factors involved in viral entry, within a more robust and reproducible environment. Viral infection generally begins with receptor recognition at the host cell surface, following highly specific cell-virus interactions. Most of these interactions are expected to take place at the plasma membrane of the HepaRG cells. In the present study, we used this cell line to explore changes between the plasma membrane of undifferentiated (− and differentiated (+ cells and to identify differentially-regulated proteins or signaling networks that might potentially be involved in HBV entry. Our initial study identified a series of proteins that are differentially expressed in the plasma membrane of (− and (+ cells and are good candidates for potential cell-virus interactions. To our knowledge, this is the first study using functional proteomics to study plasma membrane proteins from HepaRG cells, providing a platform for future experiments that will allow us to understand the cell-virus interaction and mechanism of HBV viral infection.

  16. Membrane phospholipids and radiation-induced death of mammalian cells

    International Nuclear Information System (INIS)

    Wolters, H.


    Radiation-induced cell killing is generally believed to be a consequence of residual DNA damage or damage that is mis-repaired. However, besides this DNA damage, damage to other molecules or structures of the cell may be involved in the killing. Especially membranes have been suggested as a determinant in cellular radiosensitivity. In this thesis experiments are described, dealing with the possible involvement of membranes in radiation-induced killing of mammalian cells. A general treatise of membrane structure is followed by information concerning deleterious effects of radiation on membranes. Consequences of damage to structure and function of membranes are reviewed. Thereafter evidence relating to the possible involvement of membranes in radiation-induced cell killing is presented. (Auth.)

  17. Regulated Mucin Secretion from Airway Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Kenneth Bruce Adler


    Full Text Available Secretory epithelial cells of the proximal airways synthesize and secrete gel-forming polymeric mucins. The secreted mucins adsorb water to form mucus that is propelled by neighboring ciliated cells, providing a mobile barrier which removes inhaled particles and pathogens from the lungs. Several features of the intracellular trafficking of mucins make the airway secretory cell an interesting comparator for the cell biology of regulated exocytosis. Polymeric mucins are exceedingly large molecules (up to 3x10^6 D per monomer whose folding and initial polymerization in the ER requires the protein disulfide isomerase Agr2. In the Golgi, mucins further polymerize to form chains and possibly branched networks comprising more than 20 monomers. The large size of mucin polymers imposes constraints on their packaging into transport vesicles along the secretory pathway. Sugar side chains account for >70% of the mass of mucins, and their attachment to the protein core by O-glycosylation occurs in the Golgi. Mature polymeric mucins are stored in large secretory granules ~1 um in diameter. These are translocated to the apical membrane to be positioned for exocytosis by cooperative interactions among MARCKS, cysteine string protein (CSP, HSP70 and the cytoskeleton. Mucin granules undergo exocytic fusion with the plasma membrane at a low basal rate and a high stimulated rate. Both rates are mediated by a regulated exocytic mechanism as indicated by phenotypes in both basal and stimulated secretion in mice lacking Munc13-2, a sensor of the second messengers calcium and diacylglycerol (DAG. Basal secretion is induced by low levels of activation of P2Y2 purinergic and A3 adenosine receptors by extracellular ATP released in paracrine fashion and its metabolite adenosine. Stimulated secretion is induced by high levels of the same ligands, and possibly by inflammatory mediators as well. Activated receptors are coupled to phospholipase C by Gq, resulting in the

  18. Polybenzimidazole and sulfonated polyhedral oligosilsesquioxane composite membranes for high temperature polymer electrolyte membrane fuel cells

    DEFF Research Database (Denmark)

    Aili, David; Allward, Todd; Alfaro, Silvia Martinez


    Composite membranes based on poly(2,2′(m-phenylene)-5,5́bibenzimidazole) (PBI) and sulfonated polyhedral oligosilsesquioxane (S-POSS) with S-POSS contents of 5 and 10wt.% were prepared by solution casting as base materials for high temperature polymer electrolyte membrane fuel cells. With membranes...... based on pure PBI as a reference point, the composite membranes were characterized with respect to spectroscopic and physicochemical properties. After doping with phosphoric acid, the composite membranes showed considerably improved ex situ proton conductivity under anhydrous as well as under fully...... humidified conditions in the 120-180°C temperature range. The conductivity improvements were also confirmed by in situ fuel cell tests at 160°C and further supported by the electrochemical impedance spectroscopy data based on the operating membrane electrode assemblies, demonstrating the technical...

  19. Large Deformation Mechanics of Plasma Membrane Chained Vesicles in Cells (United States)

    Kosawada, Tadashi; Sanada, Kouichi; Takano, Tetsuo

    The clathrin-coated pits, vesicles and chained vesicles on the inner surface of the plasma membrane facilitate the cell to transport specific extracellular macromolecules. This cellular process is strongly involved with large mechanical deformations of the plasma membrane accompanied by changes in membrane curvature. The assembly of the clathrin coat is thought to provide curvature into the membrane. Hence, effects of in-plane shear elasticity due to these coat structure may be significant on the vesicular mechanics. In this study, large deformation mechanics of plasma membrane chained vesicles in cells have been formulated based on minimization of bending and in-plane shear strain energy of the membrane. Effects of outer surrounding cytoplasmic flat membrane upon mechanically stable shapes of the vesicles were revealed, while effects of in-plane shear elasticity were partly discussed.

  20. Mechanosensitivity of cell membranes. Ion channels, lipid matrix and cytoskeleton. (United States)

    Petrov, A G; Usherwood, P N


    Physical and biophysical mechanisms of mechano-sensitivity of cell membranes are reviewed. The possible roles of the lipid matrix and of the cytoskeleton in membrane mechanoreception are discussed. Techniques for generation of static strains and dynamic curvatures of membrane patches are considered. A unified model for stress-activated and stress-inactivated ion channels under static strains is described. A review of work on stress-sensitive pores in lipid-peptide model membranes is presented. The possible role of flexoelectricity in mechano-electric transduction, e.g. in auditory receptors is discussed. Studies of flexoelectricity in model lipid membranes, lipid-peptide membranes and natural membranes containing ion channels are reviewed. Finally, possible applications in molecular electronics of mechanosensors employing some of the recognized principles of mechano-electric transduction in natural membranes are discussed.

  1. Review of cell performance in anion exchange membrane fuel cells (United States)

    Dekel, Dario R.


    Anion exchange membrane fuel cells (AEMFCs) have recently received increasing attention since in principle they allow for the use of non-precious metal catalysts, which dramatically reduces the cost per kilowatt of power in fuel cell devices. Until not long ago, the main barrier in the development of AEMFCs was the availability of highly conductive anion exchange membranes (AEMs); however, improvements on this front in the past decade show that newly developed AEMs have already reached high levels of conductivity, leading to satisfactory cell performance. In recent years, a growing number of research studies have reported AEMFC performance results. In the last three years, new records in performance were achieved. Most of the literature reporting cell performance is based on hydrogen-AEMFCs, although an increasing number of studies have also reported the use of fuels others than hydrogen - such as alcohols, non-alcohol C-based fuels, as well as N-based fuels. This article reviews the cell performance and performance stability achieved in AEMFCs through the years since the first reports in the early 2000s.

  2. Performance enhancement of membrane electrode assemblies with plasma etched polymer electrolyte membrane in PEM fuel cell

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Yong-Hun; Yoon, Won-Sub [School of Advanced Materials Engineering, Kookmin University, 861-1 Jeongneung-dong, Seongbuk-gu, Seoul 136-702 (Korea); Bae, Jin Woo; Cho, Yoon-Hwan; Lim, Ju Wan; Ahn, Minjeh; Jho, Jae Young; Sung, Yung-Eun [World Class University (WCU) program of Chemical Convergence for Energy and Environment (C2E2), School of Chemical and Biological Engineering, College of Engineering, Seoul National University (SNU), 599 Gwanak-Ro, Gwanak-gu, Seoul 151-744 (Korea); Kwon, Nak-Hyun [Fuel Cell Vehicle Team 3, Advanced Technology Center, Corporate Research and Development Division, Hyundai-Kia Motors, 104 Mabuk-dong, Giheung-gu, Yongin-si, Gyeonggi-do 446-912 (Korea)


    In this work, a surface modified Nafion 212 membrane was fabricated by plasma etching in order to enhance the performance of a membrane electrode assembly (MEA) in a polymer electrolyte membrane fuel cell. Single-cell performance of MEA at 0.7 V was increased by about 19% with membrane that was etched for 10 min compared to that with untreated Nafion 212 membrane. The MEA with membrane etched for 20 min exhibited a current density of 1700 mA cm{sup -2} at 0.35 V, which was 8% higher than that of MEA with untreated membrane (1580 mA cm{sup -2}). The performances of MEAs containing etched membranes were affected by complex factors such as the thickness and surface morphology of the membrane related to etching time. The structural changes and electrochemical properties of the MEAs with etched membranes were characterized by field emission scanning electron microscopy, Fourier transform-infrared spectrometry, electrochemical impedance spectroscopy, and cyclic voltammetry. (author)

  3. Membrane transport of anandamide through resealed human red blood cell membranes

    DEFF Research Database (Denmark)

    Bojesen, I.N.; Hansen, Harald S.


    The use of resealed red blood cell membranes (ghosts) allows the study of the transport of a compound in a nonmetabolizing system with a biological membrane. Transmembrane movements of anandamide (N-arachidonoylethanolamine, arachidonoylethanolamide) have been studied by exchange efflux experiments...... at 0°C and pH 7.3 with albumin-free and albumin-filled human red blood cell ghosts. The efflux kinetics is biexponential and is analyzed in terms of compartment models. The distribution of anandamide on the membrane inner to outer leaflet pools is determined to be 0.275 ± 0.023, and the rate constant...... of unidirectional flux from inside to outside is 0.361 ± 0.023 s. The rate constant of unidirectional flux from the membrane to BSA in the medium ([BSA]) increases with the square root of [BSA] in accordance with the theory of an unstirred layer around ghosts. Anandamide passed through the red blood cell membrane...

  4. Lactobacillus casei combats acid stress by maintaining cell membrane functionality. (United States)

    Wu, Chongde; Zhang, Juan; Wang, Miao; Du, Guocheng; Chen, Jian


    Lactobacillus casei strains have traditionally been recognized as probiotics and frequently used as adjunct culture in fermented dairy products where lactic acid stress is a frequently encountered environmental condition. We have investigated the effect of lactic acid stress on the cell membrane of L. casei Zhang [wild type (WT)] and its acid-resistant mutant Lbz-2. Both strains were grown under glucose-limiting conditions in chemostats; following challenge by low pH, the cell membrane stress responses were investigated. In response to acid stress, cell membrane fluidity decreased and its fatty acid composition changed to reduce the damage caused by lactic acid. Compared with the WT, the acid-resistant mutant exhibited numerous survival advantages, such as higher membrane fluidity, higher proportions of unsaturated fatty acids, and higher mean chain length. In addition, cell integrity analysis showed that the mutant maintained a more intact cellular structure and lower membrane permeability after environmental acidification. These results indicate that alteration in membrane fluidity, fatty acid distribution, and cell integrity are common mechanisms utilized by L. casei to withstand severe acidification and to reduce the deleterious effect of lactic acid on the cell membrane. This detailed comparison of cell membrane responses between the WT and mutant add to our knowledge of the acid stress adaptation and thus enable new strategies to be developed aimed at improving the industrial performance of this species under acid stress.

  5. Correlation between membrane fluidity cellular development and stem cell differentiation

    KAUST Repository

    Noutsi, Pakiza


    Cell membranes are made up of a complex structure of lipids and proteins that diffuse laterally giving rise to what we call membrane fluidity. During cellular development, such as neuronal differentiation, cell membranes undergo dramatic structural changes induced by proteins such as ARC and Cofilin among others in the case of synaptic modification. In this study we used the generalized polarization (GP) property of fluorescent probe Laurdan using two-photon microscopy to determine membrane fluidity as a function of time and for various cell lines. A low GP value corresponds to a higher fluidity and a higher GP value is associated with a more rigid membrane. Four different cell lines were monitored such as hN2, NIH3T3, HEK293 and L6 cells. As expected, NIH3T3 cells have more rigid membrane at earlier stages of their development. On the other hand neurons tend to have the highest membrane fluidity early in their development emphasizing its correlation with plasticity and the need for this malleability during differentiation. This study sheds light on the involvement of membrane fluidity during neuronal differentiation and development of other cell lines.

  6. Phosphoric acid doped imidazolium polysulfone membranes for high temperature proton exchange membrane fuel cells

    DEFF Research Database (Denmark)

    Yang, Jingshuai; Li, Qingfeng; Jensen, Jens Oluf


    A novel acid–base polymer membrane is prepared by doping of imidazolium polysulfone with phosphoric acid for high temperature proton exchange membrane fuel cells. Polysulfone is first chloromethylated, followed by functionalization of the chloromethylated polysulfone with alkyl imidazoles i.e. me...

  7. Plasma membrane--cortical cytoskeleton interactions: a cell biology approach with biophysical considerations. (United States)

    Kapus, András; Janmey, Paul


    From a biophysical standpoint, the interface between the cell membrane and the cytoskeleton is an intriguing site where a "two-dimensional fluid" interacts with an exceedingly complex three-dimensional protein meshwork. The membrane is a key regulator of the cytoskeleton, which not only provides docking sites for cytoskeletal elements through transmembrane proteins, lipid binding-based, and electrostatic interactions, but also serves as the source of the signaling events and molecules that control cytoskeletal organization and remolding. Conversely, the cytoskeleton is a key determinant of the biophysical and biochemical properties of the membrane, including its shape, tension, movement, composition, as well as the mobility, partitioning, and recycling of its constituents. From a cell biological standpoint, the membrane-cytoskeleton interplay underlies--as a central executor and/or regulator--a multitude of complex processes including chemical and mechanical signal transduction, motility/migration, endo-/exo-/phagocytosis, and other forms of membrane traffic, cell-cell, and cell-matrix adhesion. The aim of this article is to provide an overview of the tight structural and functional coupling between the membrane and the cytoskeleton. As biophysical approaches, both theoretical and experimental, proved to be instrumental for our understanding of the membrane/cytoskeleton interplay, this review will "oscillate" between the cell biological phenomena and the corresponding biophysical principles and considerations. After describing the types of connections between the membrane and the cytoskeleton, we will focus on a few key physical parameters and processes (force generation, curvature, tension, and surface charge) and will discuss how these contribute to a variety of fundamental cell biological functions. © 2013 American Physiological Society.

  8. Development and characterization of proton conductive membranes and membrane electrode assemblies for fuel cells (United States)

    Jiang, Ruichun

    Polymer electrolyte membrane fuel cells (PEMFCs), including hydrogen fuel cells (PEMFCs) and direct methanol fuel cells (DMFCs), are considered as attractive electrical power sources. However, there are some technical obstacles that impede the commercialization of PEMFCs. For instance, in H 2-PEMFCs, carbon monoxide (CO) poisoning of the anode catalyst causes serious performance loss; in DMFCs, methanol crossover through the membrane reduces the overall fuel cell efficiency. This work focused on: (1) developing high performance membrane electrode assemblies (MEAs) and investigating their behavior at higher temperature H2-PEMFC with H2+CO as the fuel; (2) improving DMFCs efficiency by preparing low methanol crossover/good proton conductivity membranes based on NafionRTM matrix; (3) synthesizing and modifying low cost sulfonated hydrocarbon (SPEEK) membranes for both H2-PEMFCs and DMFCs applications. High performance membrane electrode assemblies (MEAs) with composite NafionRTM-TeflonRTM-Zr(HPO 4)2 membranes were prepared, optimized and characterized at higher temperature (> 100°C)/lower relative humidity (oxidation mechanism of H2/CO in higher temperature PEMFC was investigated and simulated. Two type of membranes based on NafionRTM matrix were prepared: silica/NafionRTM membrane and palladium impregnated NafionRTM (Pd-NafionRTM) membrane. The composite silica/NafionRTM membrane was developed by in-situ sol-gel reaction followed by solution casting, while the Pd-NafionRTM was fabricated via a supercritical fluid CO2 (scCO 2) route. Reduced methanol crossover and enhanced efficiency was observed by applying each of the two membranes to DMFCs. In addition, the research demonstrated that scCO2 is a promising technique for modifying membranes or depositing nano-particle electrocatalysts onto electrolyte. Sulfonated poly(ether ether ketone) (SPEEK) was synthesized by a sulfonation reaction using poly(ether ether ketone) (PEEK). Multilayer structure SPEEK membranes with

  9. Cell swelling and volume regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay


    The extracellular space in the brain is typically 20% of the tissue volume and is reduced to at least half its size under conditions of neural insult. Whether there is a minimum size to the extracellular space was discussed. A general model for cell volume regulation was presented, followed...... by a discussion on how many of the generally involved mechanisms are identified in neural cells and (or) in astrocytes. There seems to be clear evidence suggesting that parallel K+ and Cl- channels mediate regulatory volume decrease in primary cultures of astrocytes, and a stretch-activated cation channel has...... been reported. The role of the different channels was discussed. A taurine leak pathway is clearly activated after cell swelling both in astrocytes and in neurones. The relations between the effect of glutamate and cell swelling were discussed. Discussion on the clearance of potassium from...

  10. Pyroelectricity as a possible mechanism for cell membrane permeabilization. (United States)

    García-Sánchez, Tomás; Muscat, Adeline; Leray, Isabelle; Mir, Lluis M


    The effects of pyroelectricity on cell membrane permeability had never been explored. Pyroelectricity consists in the generation of an electric field in the surface of some materials when a change in temperature is produced. In the present study, tourmaline microparticles, which are known to display pyroelectrical properties, were subjected to different changes in temperature upon exposure to cells in order to induce an electric field at their surface. Then, the changes in the permeability of the cell membrane to a cytotoxic agent (bleomycin) were assessed by a cloning efficacy test. An increase in the permeability of the cell membrane was only detected when tourmaline was subjected to a change in temperature. This suggests that the apparition of an induced pyroelectrical electric field on the material could actually be involved in the observed enhancement of the cell membrane permeability as a result of cell electropermeabilization. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Salmonella Rapidly Regulates Membrane Permeability To Survive Oxidative Stress

    Directory of Open Access Journals (Sweden)

    Joris van der Heijden


    Full Text Available The outer membrane (OM of Gram-negative bacteria provides protection against toxic molecules, including reactive oxygen species (ROS. Decreased OM permeability can promote bacterial survival under harsh circumstances and protects against antibiotics. To better understand the regulation of OM permeability, we studied the real-time influx of hydrogen peroxide in Salmonella bacteria and discovered two novel mechanisms by which they rapidly control OM permeability. We found that pores in two major OM proteins, OmpA and OmpC, could be rapidly opened or closed when oxidative stress is encountered and that the underlying mechanisms rely on the formation of disulfide bonds in the periplasmic domain of OmpA and TrxA, respectively. Additionally, we found that a Salmonella mutant showing increased OM permeability was killed more effectively by treatment with antibiotics. Together, these results demonstrate that Gram-negative bacteria regulate the influx of ROS for defense against oxidative stress and reveal novel targets that can be therapeutically targeted to increase bacterial killing by conventional antibiotics.

  12. How the antimicrobial peptides destroy bacteria cell membrane: Translocations vs. membrane buckling (United States)

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


    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. A cell culture technique for human epiretinal membranes to describe cell behavior and membrane contraction in vitro. (United States)

    Wertheimer, Christian; Eibl-Lindner, Kirsten H; Compera, Denise; Kueres, Alexander; Wolf, Armin; Docheva, Denitsa; Priglinger, Siegfried G; Priglinger, Claudia; Schumann, Ricarda G


    To introduce a human cell culture technique for investigating in-vitro behavior of primary epiretinal cells and membrane contraction of fibrocellular tissue surgically removed from eyes with idiopathic macular pucker. Human epiretinal membranes were harvested from ten eyes with idiopathic macular pucker during standard vitrectomy. Specimens were fixed on cell culture plastic using small entomological pins to apply horizontal stress to the tissue, and then transferred to standard cell culture conditions. Cell behavior of 400 epiretinal cells from 10 epiretinal membranes was observed in time-lapse microscopy and analyzed in terms of cell migration, cell velocity, and membrane contraction. Immunocytochemistry was performed for cell type-specific antigens. Cell specific differences in migration behavior were observed comprising two phenotypes: (PT1) epiretinal cells moving fast, less directly, with small round phenotype and (PT2) epiretinal cells moving slowly, directly, with elongated large phenotype. No mitosis, no outgrowth and no migration onto the plastic were seen. Horizontal contraction measurements showed variation between specimens. Masses of epiretinal cells with a myofibroblast-like phenotype expressed cytoplasmatic α-SMA stress fibers and correlated with cell behavior characteristics (PT2). Fast moving epiretinal cells (PT1) were identified as microglia by immunostaining. This in-vitro technique using traction application allows for culturing surgically removed epiretinal membranes from eyes with idiopathic macular pucker, demonstrating cell behavior and membrane contraction of primary human epiretinal cells. Our findings emphasize the abundance of myofibroblasts, the presence of microglia and specific differences of cell behavior in these membranes. This technique has the potential to improve the understanding of pathologies at the vitreomacular interface and might be helpful in establishing anti-fibrotic treatment strategies.

  14. Models of dynamic extraction of lipid tethers from cell membranes

    International Nuclear Information System (INIS)

    Nowak, Sarah A; Chou, Tom


    When a ligand that is bound to an integral membrane receptor is pulled, the membrane and the underlying cytoskeleton can deform before either the membrane delaminates from the cytoskeleton or the ligand detaches from the receptor. If the membrane delaminates from the cytoskeleton, it may be further extruded and form a membrane tether. We develop a phenomenological model for this process by assuming that deformations obey Hooke's law up to a critical force at which the cell membrane locally detaches from the cytoskeleton and a membrane tether forms. We compute the probability of tether formation and show that tethers can be extruded only within an intermediate range of force loading rates and pulling velocities. The mean tether length that arises at the moment of ligand detachment is computed as are the force loading rates and pulling velocities that yield the longest tethers

  15. Carbon nanotubes based nafion composite membranes for fuel cell applications

    CSIR Research Space (South Africa)

    Cele, NP


    Full Text Available composite membranes. Keywords: Carbon Nanotubes, Conductivity, Fuel Cell, Nafion, Nanocomposite Membranes, Thermal Properties, Water Uptake FUEL CELLS 00, 0000, No. 0, 1–8 ? 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim 1 ORIGINA L RESEAR CH PAPE... used strategies to overcome these drawbacks is the modification of Nafion by using polymer nanocomposite (PNC) technology. PNCs have recently shown a worldwide growth effort especially in the fabrication of high temperature PEM for fuel cells [18...

  16. Coating nanoparticles with cell membranes for targeted drug delivery. (United States)

    Gao, Weiwei; Zhang, Liangfang


    Targeted delivery allows drug molecules to preferentially accumulate at the sites of action and thus holds great promise to improve therapeutic index. Among various drug-targeting approaches, nanoparticle-based delivery systems offer some unique strengths and have achieved exciting preclinical and clinical results. Herein, we aim to provide a review on the recent development of cell membrane-coated nanoparticle system, a new class of biomimetic nanoparticles that combine both the functionalities of cellular membranes and the engineering flexibility of synthetic nanomaterials for effective drug delivery and novel therapeutics. This review is particularly focused on novel designs of cell membrane-coated nanoparticles as well as their underlying principles that facilitate the purpose of drug targeting. Three specific areas are highlighted, including: (i) cell membrane coating to prolong nanoparticle circulation, (ii) cell membrane coating to achieve cell-specific targeting and (iii) cell membrane coating for immune system targeting. Overall, cell membrane-coated nanoparticles have emerged as a novel class of targeted nanotherapeutics with strong potentials to improve on drug delivery and therapeutic efficacy for treatment of various diseases.

  17. Formation of Cell Membrane Component Domains in Artificial Lipid Bilayer. (United States)

    Tero, Ryugo; Fukumoto, Kohei; Motegi, Toshinori; Yoshida, Miyu; Niwano, Michio; Hirano-Iwata, Ayumi


    The lipid bilayer environment around membrane proteins strongly affects their structure and functions. Here, we aimed to study the fusion of proteoliposomes (PLs) derived from cultured cells with an artificial lipid bilayer membrane and the distribution of the PL components after the fusion. PLs, which were extracted as a crude membrane fraction from Chinese hamster ovary (CHO) cells, formed isolated domains in a supported lipid bilayer (SLB), comprising phosphatidylcholine (PC), phosphatidylethanolamine (PE), and cholesterol (Chol), after the fusion. Observation with a fluorescence microscope and an atomic force microscope showed that the membrane fusion occurred selectively at microdomains in the PC + PE + Chol-SLB, and that almost all the components of the PL were retained in the domain. PLs derived from human embryonic kidney 293 (HEK) cells also formed isolated domains in the PC + PE + Chol-SLB, but their fusion kinetics was different from that of the CHO-PLs. We attempted to explain the mechanism of the PL-SLB fusion and the difference between CHO- and HEK-PLs, based on a kinetic model. The domains that contained the whole cell membrane components provided environments similar to that of natural cell membranes, and were thus effective for studying membrane proteins using artificial lipid bilayer membranes.

  18. Improved Membrane Materials for PEM Fuel Cell Application

    Energy Technology Data Exchange (ETDEWEB)

    Kenneth A. Mauritz; Robert B. Moore


    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.

  19. Environmental and Genetic Factors Regulating Localization of the Plant Plasma Membrane H+-ATPase. (United States)

    Haruta, Miyoshi; Tan, Li Xuan; Bushey, Daniel B; Swanson, Sarah J; Sussman, Michael R


    A P-type H + -ATPase is the primary transporter that converts ATP to electrochemical energy at the plasma membrane of higher plants. Its product, the proton-motive force, is composed of an electrical potential and a pH gradient. Many studies have demonstrated that this proton-motive force not only drives the secondary transporters required for nutrient uptake, but also plays a direct role in regulating cell expansion. Here, we have generated a transgenic Arabidopsis ( Arabidopsis thaliana ) plant expressing H + -ATPase isoform 2 (AHA2) that is translationally fused with a fluorescent protein and examined its cellular localization by live-cell microscopy. Using a 3D imaging approach with seedlings grown for various times under a variety of light intensities, we demonstrate that AHA2 localization at the plasma membrane of root cells requires light. In dim light conditions, AHA2 is found in intracellular compartments, in addition to the plasma membrane. This localization profile was age-dependent and specific to cell types found in the transition zone located between the meristem and elongation zones. The accumulation of AHA2 in intracellular compartments is consistent with reduced H + secretion near the transition zone and the suppression of root growth. By examining AHA2 localization in a knockout mutant of a receptor protein kinase, FERONIA, we found that the intracellular accumulation of AHA2 in the transition zone is dependent on a functional FERONIA-dependent inhibitory response in root elongation. Overall, this study provides a molecular underpinning for understanding the genetic, environmental, and developmental factors influencing root growth via localization of the plasma membrane H + -ATPase. © 2018 American Society of Plant Biologists. All Rights Reserved.

  20. Durability Issues of High Temperature Proton Exchange Membrane Fuel Cells Based on Acid Doped Polybenzimidazole Membranes

    DEFF Research Database (Denmark)

    To achieve high temperature operation of proton exchange membrane fuel cells (PEMFC), preferably under ambient pressure, phosphoric acid doped polybenzimidazole (PBI) membrane represents an effective approach, which in recent years has motivated extensive research activities with great progress....... As a critical concern, issues of long term durability of PBI based fuel cells are addressed in this talk, including oxidative degradation of the polymer, mechanical failures of the membrane, acid leaching out, corrosion of carbon support and sintering of catalysts particles. Excellent polymer durability has...

  1. Layer-by-layer cell membrane assembly (United States)

    Matosevic, Sandro; Paegel, Brian M.


    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. Depletion of membrane cholesterol compromised caspase-8 imparts in autophagy induction and inhibition of cell migration in cancer cells. (United States)

    Kumar, Mukesh; Irungbam, Karuna; Kataria, Meena


    Cholesterol in lipid raft plays crucial role on cancer cell survival during metastasis of cancer cells. Cancer cells are reported to enrich cholesterol in lipid raft which make them more susceptible to cell death after cholesterol depletion than normal cells. Methyl-β-cyclodextrin (MβCD), an amphipathic polysaccharide known to deplete the membrane cholesterol, induces cell death selectively in cancer cells. Present work was designed to identify the major form of programmed cell death in membrane cholesterol depleted cancer cells (MDA-MB 231 and 4T1) and its impact on migration efficiency of cancer cells. Membrane cholesterol alteration and morphological changes in 4T1 and MDA-MB 231 cancer cells by MβCD were measured by fluorescent microscopy. Cell death and cell proliferation were observed by PI, AO/EB and MTT assay respectively. Programme cell death was confirmed by flow cytometer. Caspase activation was assessed by MTT and PI after treatments with Z-VAD [OME]-FMK, mitomycin c and cycloheximide. Necroptosis, autophagy, pyroptosis and paraptosis were examined by cell proliferation assay and flow cytometry. Relative quantitation of mRNA of caspase-8, necroptosis and autophagy genes were performed. Migration efficiency of cancer cells were determined by wound healing assay. We found caspase independent cell death in cholesterol depleted MDA-MB 231 cells which was reduced by (3-MA) an autophagy inhibitor. Membrane cholesterol depletion neither induces necroptosis, paraptosis nor pyroptosis in MDA-MB 231 cells. Subsequent activation of caspase-8 after co-incubation of mitomycin c and cycloheximide separately, restored the cell viability in cholesterol depleted MDA-MB 231 cells. Down regulation of caspase-8 mRNA in cholesterol depleted cancer cells ensures that caspase-8 indirectly promotes the induction of autophagy. In another experiment we have demonstrated that membrane cholesterol depletion reduces the migration efficiency in cancer cells. Together our

  3. Membrane orientation of droplets prepared from Chara corallina internodal cells. (United States)

    Berecki, G; Eijken, M; Van Iren, F; Van Duijn, B


    It is generally accepted that the membrane surrounding droplets from characean cells originates from the tonoplast, but there is some uncertainty regarding droplet membrane sidedness. This issue was addressed directly by combining two different droplet isolation methods and the patch clamp technique. Neutral red accumulation was used to demonstrate the presence of H(+)-transport over the membrane and to predict membrane orientation. Two types of droplet populations with differently oriented membranes could be formed in an iso-osmotic bath solution. Cytoplasmic droplets (cytosolic side of the tonoplast inside) contained cytoplasm, while the second type of droplet population contained vacuolar sap (vacuolar droplets, vacuolar side of the tonoplast inside). Smaller vesicels also appeared inside the droplets, with an apparently inversely oriented membrane. Confocal laser scanning microscopy indirectly demonstrated that, at least with one of the droplet isolation methods, the plasma membrane entirely remains in the internodal cell after intracellular perfusion. Both types of droplet populations allowed the formation of excised patches and single-channel measurements by the patch clamp technique. Properties of anion channels in the tonoplast could be used to prove the predicted membrane orientation, knowing that Ca2+ can only activate these channels from the cytosolic side. These results provide useful data for studies addressing ligand-binding, block and modulation, organization and interaction of proteins within the membrane or with other regulatory factors, where it is important to control membrane orientation.

  4. Glutamic Acid Residues in HIV-1 p6 Regulate Virus Budding and Membrane Association of Gag. (United States)

    Friedrich, Melanie; Setz, Christian; Hahn, Friedrich; Matthaei, Alina; Fraedrich, Kirsten; Rauch, Pia; Henklein, Petra; Traxdorf, Maximilian; Fossen, Torgils; Schubert, Ulrich


    The HIV-1 Gag p6 protein regulates the final abscission step of nascent virions from the cell membrane by the action of its two late (L-) domains, which recruit Tsg101 and ALIX, components of the ESCRT system. Even though p6 consists of only 52 amino acids, it is encoded by one of the most polymorphic regions of the HIV-1 gag gene and undergoes various posttranslational modifications including sumoylation, ubiquitination, and phosphorylation. In addition, it mediates the incorporation of the HIV-1 accessory protein Vpr into budding virions. Despite its small size, p6 exhibits an unusually high charge density. In this study, we show that mutation of the conserved glutamic acids within p6 increases the membrane association of Pr55 Gag followed by enhanced polyubiquitination and MHC-I antigen presentation of Gag-derived epitopes, possibly due to prolonged exposure to membrane bound E3 ligases. The replication capacity of the total glutamic acid mutant E0A was almost completely impaired, which was accompanied by defective virus release that could not be rescued by ALIX overexpression. Altogether, our data indicate that the glutamic acids within p6 contribute to the late steps of viral replication and may contribute to the interaction of Gag with the plasma membrane.

  5. Radiation Interaction with Therapeutic Drugs and Cell Membranes

    International Nuclear Information System (INIS)

    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.


    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

  6. Cubic membranes: a legend beyond the Flatland* of cell membrane organization. (United States)

    Almsherqi, Zakaria A; Kohlwein, Sepp D; Deng, Yuru


    Cubic membranes represent highly curved, three-dimensional nanoperiodic structures that correspond to mathematically well defined triply periodic minimal surfaces. Although they have been observed in numerous cell types and under different conditions, particularly in stressed, diseased, or virally infected cells, knowledge about the formation and function of nonlamellar, cubic structures in biological systems is scarce, and research so far is restricted to the descriptive level. We show that the "organized smooth endoplasmic reticulum" (OSER; Snapp, E.L., R.S. Hegde, M. Francolini, F. Lombardo, S. Colombo, E. Pedrazzini, N. Borgese, and J. Lippincott-Schwartz. 2003. J. Cell Biol. 163:257-269), which is formed in response to elevated levels of specific membrane-resident proteins, is actually the two-dimensional representation of two subtypes of cubic membrane morphology. Controlled OSER induction may thus provide, for the first time, a valuable tool to study cubic membrane formation and function at the molecular level.

  7. Ionic fluxes in erythrocyte membranes of sickle cell anaemia ...

    African Journals Online (AJOL)

    Ionic fluxes in erythrocyte membranes of sickle cell anaemia subjects at different tonicities. ... Journal of African Association of Physiological Sciences ... The aim of this study was to investigate ionic fluxes in membrane of erythrocytes at different tonicities with a view to highlighting any selective ionic-fluxing potential of ...

  8. Toughness of membranes applied in polymer electrolyte fuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Kiefer, J.; Brack, H.P.; Scherer, G.G. [Paul Scherrer Inst. (PSI), Villigen (Switzerland)


    Since several years we apply the radiation-grafting technique to prepare polymeric membranes for application in polymer electrolyte fuel cells (PEFCs). Our investigations presented here focus on changes in toughness of these materials after the various synthesis steps and the importance of membrane toughness for their application in PEFCs. (author) 2 figs., 4 refs.

  9. Perforate on CHO cell membranes induced by electromagnetic ...

    African Journals Online (AJOL)

    Atomic force microscopy (AFM) has been used to visualize the morphological change on the surface of Chinese hamster ovary (CHO) cell membranes before and after electromagnetic pulses (EMP) irradiation. The results show that there were different sizes and shapes of membrane perforate (width ranging from 0.39 - 0.66 ...

  10. Catalytic membranes for CO oxidation in fuel cells (United States)

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


    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.

  11. Cloning of B cell-specific membrane tetraspanning molecule BTS possessing B cell proliferation-inhibitory function. (United States)

    Suenaga, Tadahiro; Arase, Hisashi; Yamasaki, Sho; Kohno, Masayuki; Yokosuka, Tadashi; Takeuchi, Arata; Hattori, Takamichi; Saito, Takashi


    Lymphocyte proliferation is regulated by signals through antigen receptors, co-stimulatory receptors, and other positive and negative modulators. Several membrane tetraspanning molecules are also involved in the regulation of lymphocyte growth and death. We cloned a new B cell-specific tetraspanning (BTS) membrane molecule, which is similar to CD20 in terms of expression, structure and function. BTS is specifically expressed in the B cell line and its expression is increased after the pre-B cell stage. BTS is expressed in intracellular granules and on the cell surface. Overexpression of BTS in immature B cell lines induces growth retardation through inhibition of cell cycle progression and cell size increase without inducing apoptosis. This inhibitory function is mediated predominantly by the N terminus of BTS. The development of mature B cells is inhibited in transgenic mice expressing BTS, suggesting that BTS is involved in the in vivo regulation of B cells. These results indicate that BTS plays a role in the regulation of cell division and B cell growth.

  12. Investigating Sterol and Redox Regulation of the Ion Channel Activity of CLIC1 Using Tethered Bilayer Membranes

    Directory of Open Access Journals (Sweden)

    Heba Al Khamici


    Full Text Available The Chloride Intracellular Ion Channel (CLIC family consists of six conserved proteins in humans. These are a group of enigmatic proteins, which adopt both a soluble and membrane bound form. CLIC1 was found to be a metamorphic protein, where under specific environmental triggers it adopts more than one stable reversible soluble structural conformation. CLIC1 was found to spontaneously insert into cell membranes and form chloride ion channels. However, factors that control the structural transition of CLIC1 from being an aqueous soluble protein into a membrane bound protein have yet to be adequately described. Using tethered bilayer lipid membranes and electrical impedance spectroscopy system, herein we demonstrate that CLIC1 ion channel activity is dependent on the type and concentration of sterols in bilayer membranes. These findings suggest that membrane sterols play an essential role in CLIC1’s acrobatic switching from a globular soluble form to an integral membrane form, promoting greater ion channel conductance in membranes. What remains unclear is the precise nature of this regulation involving membrane sterols and ultimately determining CLIC1’s membrane structure and function as an ion channel. Furthermore, our impedance spectroscopy results obtained using CLIC1 mutants, suggest that the residue Cys24 is not essential for CLIC1’s ion channel function. However Cys24 does appear important for optimal ion channel activity. We also observe differences in conductance between CLIC1 reduced and oxidized forms when added to our tethered membranes. Therefore, we conclude that both membrane sterols and redox play a role in the ion channel activity of CLIC1.

  13. Plasma membrane organization and dynamics is probe and cell line dependent. (United States)

    Huang, Shuangru; Lim, Shi Ying; Gupta, Anjali; Bag, Nirmalya; Wohland, Thorsten


    The action and interaction of membrane receptor proteins take place within the plasma membrane. The plasma membrane, however, is not a passive matrix. It rather takes an active role and regulates receptor distribution and function by its composition and the interaction of its lipid components with embedded and surrounding proteins. Furthermore, it is not a homogenous fluid but contains lipid and protein domains of various sizes and characteristic lifetimes which are important in regulating receptor function and signaling. The precise lateral organization of the plasma membrane, the differences between the inner and outer leaflet, and the influence of the cytoskeleton are still debated. Furthermore, there is a lack of comparisons of the organization and dynamics of the plasma membrane of different cell types. Therefore, we used four different specific membrane markers to test the lateral organization, the differences between the inner and outer membrane leaflet, and the influence of the cytoskeleton of up to five different cell lines, including Chinese hamster ovary (CHO-K1), Human cervical carcinoma (HeLa), neuroblastoma (SH-SY5Y), fibroblast (WI-38) and rat basophilic leukemia (RBL-2H3) cells by Imaging Total Internal Reflection (ITIR)-Fluorescence Correlation Spectroscopy (FCS). We measure diffusion in the temperature range of 298-310K to measure the Arrhenius activation energy (E Arr ) of diffusion and apply the FCS diffusion law to obtain information on the spatial organization of the probe molecules on the various cell membranes. Our results show clear differences of the FCS diffusion law and E Arr for the different probes in dependence of their localization. These differences are similar in the outer and inner leaflet of the membrane. However, these values can differ significantly between different cell lines raising the question how molecular plasma membrane events measured in different cell lines can be compared. This article is part of a Special Issue

  14. Selectivity of biopolymer membranes using HepG2 cells. (United States)

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


    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.

  15. Low Crossover Polymer Electrolyte Membranes for Direct Methanol Fuel Cells (United States)

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


    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.

  16. Cell-free system for synthesizing membrane proteins cell free method for synthesizing membrane proteins (United States)

    Laible, Philip D; Hanson, Deborah K


    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.

  17. Two novel approaches targeting cancer cell membrane for tumor therapy. (United States)

    Feng, Yingzhu; Wang, Bochu; Cao, Yang; He, Rui


    Disruption of normal cell function by chemicals, UV radiation or viruses can cause various cancer. Drugs that have been developed for cancer therapy bind to various targets to correct disorder cell behavior, repair damaged DNA or promote cell apoptosis. However, there is rare study that focuses on cancer cell membrane as target. We propose two approaches for achieving our goal. One is to use phospholipase A2 (PLA2) to cleave phospholipid heads of the bilayer of cancer cells. Because PLA2 has unique Ca(2+) catalytic site and the pH of healthy tissue cells should be slightly alkaline at 7.2-7.5, it can be easily protected by CO3(2-) in the form of PLA2-CaCO3. While PLA2-CaCO3 accumulate in cancer cells in the acidic microenvironment of which the pH is below 7, it could be converted to active state (PLA2-Ca(2+)) which can intensively damage the cancer cell membrane. The other one is to use both monoclonal antibodies and dimethylsulfoxide (DMSO). The internalization of targeted cancer cell antibodies could change the curvature of cell membrane from order state to disorder state, therefore strong detergent DMSO can destroy cancer cells at extreme low concentration. These two approaches present no harm for normal cells, therefore, drugs targeted cancer cell membrane might become a new and high effective clinical cancer therapy. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Novel High Temperature Membrane for PEM Fuel Cells, Phase I (United States)

    National Aeronautics and Space Administration — The innovation proposed in this STTR program is a high temperature membrane to increase the efficiency and power density of PEM fuel cells. The NASA application is...

  19. Stimulated-healing of proton exchange membrane fuel cell catalyst

    NARCIS (Netherlands)

    Latsuzbaia, R.; Negro, E.; Koper, G.J.M.


    Platinum nanoparticles, which are used as catalysts in Proton Exchange Membrane Fuel Cells (PEMFC), tend to degrade after long-term operation. We discriminate the following mechanisms of the degradation: poisoning, migration and coalescence, dissolution, and electrochemical Ostwald ripening. There

  20. Novel Membrane for Highly Efficient Fuel Cells, Phase I (United States)

    National Aeronautics and Space Administration — Proton Exchange Membrane (PEM) fuel cells and electrolyzers are key technologies for NASA space systems utilizing hydrogen, oxygen, or water as reactants. In order...

  1. Nafion-based nanocomposite membranes for fuel cells

    CSIR Research Space (South Africa)

    Cele, NP


    Full Text Available , mechanical properties and electrical conductivity of nafion membrane for fuel cell applications. The results showed an improvement on the thermal behaviour of prepared nation nanocomposites compared to pure Nafion with an addition of only 1 wt% MWCNTs....

  2. Poly (ether ether ketone) membranes for fuel cells

    International Nuclear Information System (INIS)

    Marrero, Jacqueline C.; Gomes, Ailton de S.; Filho, Jose C.D.; Hui, Wang S.; Oliveira, Vivianna S. de


    Polymeric membranes were developed using a SPEEK polymer matrix (sulphonated poly (ether ether ketone)), containing hygroscopic particles of zirconia (Zr) (incorporated by sol-gel method), for use as electrolyte membranes in fuel cells. SPEEK with different sulfonation degrees were used: 63 and 86%. The thermal analysis (TGA and DSC) was carried out to characterize the membranes and electrochemical impedance spectroscopy (EIS) was carried out to evaluating the proton conductivity of the membranes. Additional analysis were underway in order to characterize these membranes, which include: X-ray diffraction (XRD), scanning electron microscopy (SEM) and transmission electron microscopy (TEM) in order to evaluate the influence of zirconia and sulfonation degree on the properties of the membranes. (author)

  3. Hybrid proton-conducting membranes for polymer electrolyte fuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Gomez-Romero, Pedro [Institut de Ciencia de Materials de Barcelona (CSIC), Campus UAB, E-08193 Bellaterra (Barcelona) (Spain)]. E-mail:; Asensio, Juan Antonio [Institut de Ciencia de Materials de Barcelona (CSIC), Campus UAB, E-08193 Bellaterra (Barcelona) (Spain); Institut Quimic de Sarria, Universitat Ramon Llull, Via Augusta 390, E-08017 Barcelona (Spain); Borros, Salvador [Institut Quimic de Sarria, Universitat Ramon Llull, Via Augusta 390, E-08017 Barcelona (Spain)


    The synthesis and characterization of a novel hybrid organic-inorganic material formed by phosphomolybdic acid H{sub 3}PMo{sub 12}O{sub 40} (PMo{sub 12}) and poly(2,5-benzimidazole) (ABPBI) is reported. This material, composed of two proton-conducting components, can be cast in the form of membranes from methanesulfonic acid (MSA) solutions. Upon impregnation with phosphoric acid, the hybrid membranes present higher conductivity than the best ABPBI polymer membranes impregnated in the same conditions. These electrolyte membranes are stable up to 200 deg. C, and have a proton conductivity of 3 x 10{sup -2} S cm{sup -1} at 185 deg. C without humidification. These properties make them very good candidates as membranes for polymer electrolyte membrane fuel cells (PEMFC) at temperatures of 100-200 deg. C.

  4. Membrane Protein Mobility and Orientation Preserved in Supported Bilayers Created Directly from Cell Plasma Membrane Blebs. (United States)

    Richards, Mark J; Hsia, Chih-Yun; Singh, Rohit R; Haider, Huma; Kumpf, Julia; Kawate, Toshimitsu; Daniel, Susan


    Membrane protein interactions with lipids are crucial for their native biological behavior, yet traditional characterization methods are often carried out on purified protein in the absence of lipids. We present a simple method to transfer membrane proteins expressed in mammalian cells to an assay-friendly, cushioned, supported lipid bilayer platform using cell blebs as an intermediate. Cell blebs, expressing either GPI-linked yellow fluorescent proteins or neon-green fused transmembrane P2X2 receptors, were induced to rupture on glass surfaces using PEGylated lipid vesicles, which resulted in planar supported membranes with over 50% mobility for multipass transmembrane proteins and over 90% for GPI-linked proteins. Fluorescent proteins were tracked, and their diffusion in supported bilayers characterized, using single molecule tracking and moment scaling spectrum (MSS) analysis. Diffusion was characterized for individual proteins as either free or confined, revealing details of the local lipid membrane heterogeneity surrounding the protein. A particularly useful result of our bilayer formation process is the protein orientation in the supported planar bilayer. For both the GPI-linked and transmembrane proteins used here, an enzymatic assay revealed that protein orientation in the planar bilayer results in the extracellular domains facing toward the bulk, and that the dominant mode of bleb rupture is via the "parachute" mechanism. Mobility, orientation, and preservation of the native lipid environment of the proteins using cell blebs offers advantages over proteoliposome reconstitution or disrupted cell membrane preparations, which necessarily result in significant scrambling of protein orientation and typically immobilized membrane proteins in SLBs. The bleb-based bilayer platform presented here is an important step toward integrating membrane proteomic studies on chip, especially for future studies aimed at understanding fundamental effects of lipid interactions

  5. Mechanical Stress Downregulates MHC Class I Expression on Human Cancer Cell Membrane

    DEFF Research Database (Denmark)

    La Rocca, Rosanna; Tallerico, Rossana; Hassan, Almosawy Talib


    In our body, cells are continuously exposed to physical forces that can regulate different cell functions such as cell proliferation, differentiation and death. In this work, we employed two different strategies to mechanically stress cancer cells. The cancer and healthy cell populations were...... treated either with mechanical stress delivered by a micropump (fabricated by deep X-ray nanolithography) or by ultrasound wave stimuli. A specific down-regulation of Major Histocompatibility Complex (MHC) class I molecules expression on cancer cell membrane compared to different kinds of healthy cells...... (fibroblasts, macrophages, dendritic and lymphocyte cells) was observed, stimulating the cells with forces in the range of nano-newton, and pressures between 1 and 10 bar (1 bar5100.000 Pascal), depending on the devices used. Moreover, Raman spectroscopy analysis, after mechanical treatment, in the range...

  6. PIST regulates the intracellular trafficking and plasma membrane expression of Cadherin 23

    Directory of Open Access Journals (Sweden)

    Oshima Kazuo


    Full Text Available Abstract Background The atypical cadherin protein cadherin 23 (CDH23 is crucial for proper function of retinal photoreceptors and inner ear hair cells. As we obtain more and more information about the specific roles of cadherin 23 in photoreceptors and hair cells, the regulatory mechanisms responsible for the transport of this protein to the plasma membrane are largely unknown. Results PIST, a Golgi-associated, PDZ domain-containing protein, interacted with cadherin 23 via the PDZ domain of PIST and the C-terminal PDZ domain-binding interface (PBI of cadherin 23. By binding to cadherin 23, PIST retained cadherin 23 in the trans-Golgi network of cultured cells. The retention was released when either of the two known cadherin 23-binding proteins MAGI-1 and harmonin was co-expressed. Similar to MAGI-1 and harmonin, PIST was detected in mouse inner ear sensory hair cells. Conclusions PIST binds cadherin 23 via its PDZ domain and retains cadherin 23 in trans-Golgi network. MAGI-1 and harmonin can compete with PIST for binding cadherin 23 and release cadherin 23 from PIST's retention. Our finding suggests that PIST, MAGI-1 and harmonin collaborate in intracellular trafficking of cadherin 23 and regulate the plasma membrane expression of cadherin 23.

  7. Single-molecule tracking of small GTPase Rac1 uncovers spatial regulation of membrane translocation and mechanism for polarized signaling (United States)

    Das, Sulagna; Yin, Taofei; Yang, Qingfen; Zhang, Jingqiao; Wu, Yi I.; Yu, Ji


    Polarized Rac1 signaling is a hallmark of many cellular functions, including cell adhesion, motility, and cell division. The two steps of Rac1 activation are its translocation to the plasma membrane and the exchange of nucleotide from GDP to GTP. It is, however, unclear whether these two processes are regulated independent of each other and what their respective roles are in polarization of Rac1 signaling. We designed a single-particle tracking (SPT) method to quantitatively analyze the kinetics of Rac1 membrane translocation in living cells. We found that the rate of Rac1 translocation was significantly elevated in protrusions during cell spreading on collagen. Furthermore, combining FRET sensor imaging with SPT measurements in the same cell, the recruitment of Rac1 was found to be polarized to an extent similar to that of the nucleotide exchange process. Statistical analysis of single-molecule trajectories and optogenetic manipulation of membrane lipids revealed that Rac1 membrane translocation precedes nucleotide exchange, and is governed primarily by interactions with phospholipids, particularly PI(3,4,5)P3, instead of protein factors. Overall, the study highlights the significance of membrane translocation in spatial Rac1 signaling, which is in addition to the traditional view focusing primarily on GEF distribution and exchange reaction. PMID:25561548

  8. Proton Exchange Membrane Fuel Cells Applied for Transport Sector

    DEFF Research Database (Denmark)

    Hosseinzadeh, Elham; Rokni, Masoud


    A thermodynamic analysis of a PEMFC (proton exchange membrane fuel cell) is investigated. PEMFC may be the most promising technology for fuel cell automotive systems, which is operating at quite low temperatures, (between 60 to 80℃). In this study the fuel cell motive power part of a lift truck has...

  9. Mouse endometrial stromal cells produce basement-membrane components

    DEFF Research Database (Denmark)

    Wewer, U M; Damjanov, A; Weiss, J


    . Mouse decidual cells isolated from 6- to 7-day pregnant uteri explanted in vitro continue to synthesize basement-membrane-like extracellular matrix. Using immunohistochemistry and metabolic labeling followed by immunoprecipitation, SDS-PAGE, and fluorography, it was shown that the decidual cells...... to undergo pseudodecidualization. We thus showed that stromal cells from pregnant and nonpregnant mouse uteri synthesize significant amounts of basement-membrane components in vitro, and hence could serve as a good model for the study of normal basement-membrane components.......During mouse pregnancy, uterine stromal cells transform into morphologically distinct decidual cells under the influence of the implanting embryo and a proper hormonal environment. Mechanical stimulation of hormonally primed uterine stromal cells leads to the same morphologic alterations...

  10. Catalyst layers for proton exchange membrane fuel cells prepared by electrospray deposition on Nafion membrane (United States)

    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.

  11. Activation of Endothelial Nitric Oxide (eNOS Occurs through Different Membrane Domains in Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Jason Tran

    Full Text Available Endothelial cells respond to a large range of stimuli including circulating lipoproteins, growth factors and changes in haemodynamic mechanical forces to regulate the activity of endothelial nitric oxide synthase (eNOS and maintain blood pressure. While many signalling pathways have been mapped, the identities of membrane domains through which these signals are transmitted are less well characterized. Here, we manipulated bovine aortic endothelial cells (BAEC with cholesterol and the oxysterol 7-ketocholesterol (7KC. Using a range of microscopy techniques including confocal, 2-photon, super-resolution and electron microscopy, we found that sterol enrichment had differential effects on eNOS and caveolin-1 (Cav1 colocalisation, membrane order of the plasma membrane, caveolae numbers and Cav1 clustering. We found a correlation between cholesterol-induced condensation of the plasma membrane and enhanced high density lipoprotein (HDL-induced eNOS activity and phosphorylation suggesting that cholesterol domains, but not individual caveolae, mediate HDL stimulation of eNOS. Vascular endothelial growth factor (VEGF-induced and shear stress-induced eNOS activity was relatively independent of membrane order and may be predominantly controlled by the number of caveolae on the cell surface. Taken together, our data suggest that signals that activate and phosphorylate eNOS are transmitted through distinct membrane domains in endothelial cells.

  12. Membrane-bound Dickkopf-1 in Foxp3+ regulatory T cells suppresses T-cell-mediated autoimmune colitis. (United States)

    Chae, Wook-Jin; Park, Jong-Hyun; Henegariu, Octavian; Yilmaz, Saliha; Hao, Liming; Bothwell, Alfred L M


    Induction of tolerance is a key mechanism to maintain or to restore immunological homeostasis. Here we show that Foxp3 + regulatory T (Treg) cells use Dickkopf-1 (DKK-1) to regulate T-cell-mediated tolerance in the T-cell-mediated autoimmune colitis model. Treg cells from DKK-1 hypomorphic doubleridge mice failed to control CD4 + T-cell proliferation, resulting in CD4 T-cell-mediated autoimmune colitis. Thymus-derived Treg cells showed a robust expression of DKK-1 but not in naive or effector CD4 T cells. DKK-1 expression in Foxp3 + Treg cells was further increased upon T-cell receptor stimulation in vitro and in vivo. Interestingly, Foxp3 + Treg cells expressed DKK-1 in the cell membrane and the functional inhibition of DKK-1 using DKK-1 monoclonal antibody abrogated the suppressor function of Foxp3 + Treg cells. DKK-1 expression was dependent on de novo protein synthesis and regulated by the mitogen-activated protein kinase pathway but not by the canonical Wnt pathway. Taken together, our results highlight membrane-bound DKK-1 as a novel Treg-derived mediator to maintain immunological tolerance in T-cell-mediated autoimmune colitis. © 2017 The Authors. Immunology Published by John Wiley & Sons Ltd.

  13. Physiology of cell volume regulation in vertebrates

    DEFF Research Database (Denmark)

    Hoffmann, Else K; Lambert, Ian H; Pedersen, Stine F


    organisms. Importantly, cell volume impacts on a wide array of physiological processes, including transepithelial transport; cell migration, proliferation, and death; and changes in cell volume function as specific signals regulating these processes. A discussion of this issue concludes the review....

  14. X-radiation effects on muscle cell membrane electrical parameters

    International Nuclear Information System (INIS)

    Portela, A.; Vaccari, J.G.; Llobera, O.; Campi, M.; Delbue, M.A.; Perez, J.C.; Stewart, P.A.; Gosztonyi, A.E.; Brown Univ., Providence, R.I.


    Early effects of 100 Kilorads of X-rays on muscle cell membrane properties have been measured in sartorius muscles from Leptodactylus ocellatus. Threshold strength for rectangular current pulses increased 10% after irradiation, and action potential propagation velocity decreased 10%. Passive membrane parameters were calculated from potential responses to sub-threshold current pulses, assuming conventional cable theory. Specific membrane conductance increased to 18% after irradiation, membrane capacitance increased 14%, and length constant decreased 10% but membrane time constant was unchanged. Cell diameter decreased 5%, and resting membrane potential decreased 8%. Membrane parameters during an action potential were also evaluated by the phase-plane and current-voltage plot techniques. Irradiation significantly decreased the action potential amplitude, the excitation potential, and the maximum rates of rise and fall of membrane potential. Increases were observed in dynamic sodium and potassium conductances, peak sodium current, and net charge accumulation per action potential. This X-ray dose also produced signficant changes in the timing of peak events during the action potential; in general the whole action potential process is slower after irradiation

  15. Cell surface area and membrane folding in glioblastoma cell lines differing in PTEN and p53 status.

    Directory of Open Access Journals (Sweden)

    Simon Memmel

    Full Text Available Glioblastoma multiforme (GBM is characterized by rapid growth, invasion and resistance to chemo-/radiotherapy. The complex cell surface morphology with abundant membrane folds, microvilli, filopodia and other membrane extensions is believed to contribute to the highly invasive behavior and therapy resistance of GBM cells. The present study addresses the mechanisms leading to the excessive cell membrane area in five GBM lines differing in mutational status for PTEN and p53. In addition to scanning electron microscopy (SEM, the membrane area and folding were quantified by dielectric measurements of membrane capacitance using the single-cell electrorotation (ROT technique. The osmotic stability and volume regulation of GBM cells were analyzed by video microscopy. The expression of PTEN, p53, mTOR and several other marker proteins involved in cell growth and membrane synthesis were examined by Western blotting. The combined SEM, ROT and osmotic data provided independent lines of evidence for a large variability in membrane area and folding among tested GBM lines. Thus, DK-MG cells (wild type p53 and wild type PTEN exhibited the lowest degree of membrane folding, probed by the area-specific capacitance C m = 1.9 µF/cm(2. In contrast, cell lines carrying mutations in both p53 and PTEN (U373-MG and SNB19 showed the highest C m values of 3.7-4.0 µF/cm(2, which corroborate well with their heavily villated cell surface revealed by SEM. Since PTEN and p53 are well-known inhibitors of mTOR, the increased membrane area/folding in mutant GBM lines may be related to the enhanced protein and lipid synthesis due to a deregulation of the mTOR-dependent downstream signaling pathway. Given that membrane folds and extensions are implicated in tumor cell motility and metastasis, the dielectric approach presented here provides a rapid and simple tool for screening the biophysical cell properties in studies on targeting chemo- or radiotherapeutically the

  16. Transmembrane protein OSTA-1 shapes sensory cilia morphology via regulation of intracellular membrane trafficking in C. elegans. (United States)

    Olivier-Mason, Anique; Wojtyniak, Martin; Bowie, Rachel V; Nechipurenko, Inna V; Blacque, Oliver E; Sengupta, Piali


    The structure and function of primary cilia are critically dependent on intracellular trafficking pathways that transport ciliary membrane and protein components. The mechanisms by which these trafficking pathways are regulated are not fully characterized. Here we identify the transmembrane protein OSTA-1 as a new regulator of the trafficking pathways that shape the morphology and protein composition of sensory cilia in C. elegans. osta-1 encodes an organic solute transporter alpha-like protein, mammalian homologs of which have been implicated in membrane trafficking and solute transport, although a role in regulating cilia structure has not previously been demonstrated. We show that mutations in osta-1 result in altered ciliary membrane volume, branch length and complexity, as well as defects in localization of a subset of ciliary transmembrane proteins in different sensory cilia types. OSTA-1 is associated with transport vesicles, localizes to a ciliary compartment shown to house trafficking proteins, and regulates both retrograde and anterograde flux of the endosome-associated RAB-5 small GTPase. Genetic epistasis experiments with sensory signaling, exocytic and endocytic proteins further implicate OSTA-1 as a crucial regulator of ciliary architecture via regulation of cilia-destined trafficking. Our findings suggest that regulation of transport pathways in a cell type-specific manner contributes to diversity in sensory cilia structure and might allow dynamic remodeling of ciliary architecture via multiple inputs.

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

    International Nuclear Information System (INIS)

    Harper, Jeffrey F.


    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

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

    Energy Technology Data Exchange (ETDEWEB)

    Jeffrey F. Harper, Ph.D.


    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.

  19. Regulation of the Plasma Membrane H+-ATPase

    DEFF Research Database (Denmark)

    Falhof, Janus

    The plasma membrane (PM) H+-ATPase is responsible for generating the electrochemical gradientthat drives the secondary transport of nutrients across the cellular membrane. It belongs to a familyof cation and lipid transporters that are vital to many organisms. PM H+-ATPases are Type P3AATPases...

  20. Dendronized Polymer Architectures for Fuel Cell Membranes

    DEFF Research Database (Denmark)

    Nielsen, Mads Møller; Dimitrov, Ivaylo; Takamuku, S.


    case the side chains are synthesized and introduced in their sulfonated form onto an azide‐functionalized PSU via click chemistry. Three degrees of substitution of each architecture were prepared in order to evaluate the dependence on number of sulfonated side chains. Solution cast membranes were...

  1. The lipid organisation of the cell membrane

    Directory of Open Access Journals (Sweden)

    Ladha, S.


    Full Text Available Lipids and proteins in biological membranes are arranged in a mosaic of domains in the membrane. These domains represent small-scale heterogeneities in composition, shape and fluidity within the plane of the membrane, over the range of hundreds of nanometers to a few micrometers. They arise from the complex interactions of the heterogeneous mixtures of phospholipids, sterols, and proteins that make up all biological membranes.Los lípidos y las proteínas en las membranas biológicas están dispuestos en un mosaico de campos en la membrana. Estos campos representan heterogeneidades a pequeña escala en la composición, forma y fluidez dentro del plano de la membrana, en un rango que va de los cientos de nanómetros a los pocos micrómetros. Estos campos se originan de las complejas interacciones de las mezclas heterogéneas de fosfolípidos, esteroles y proteínas de las que están hechas todas y cada una de las membranas biológicas.

  2. Polyunsaturation in cell membranes and lipid bilayers and its effects on membrane proteins. (United States)

    Slater, S J; Kelly, M B; Yeager, M D; Larkin, J; Ho, C; Stubbs, C D


    The effect of variation of the degree of cis-unsaturation on cell membrane protein functioning was investigated using a model lipid bilayer system and protein kinase C (PKC). This protein is a key element of signal transduction. Furthermore it is representative of a class of extrinsic membrane proteins that show lipid dependent interactions with cell membranes. To test for dependence of activity on the phospholipid unsaturation, experiments were devised using a vesicle assay system consisting of phosphatidylcholine (PC) and phosphatidylserine (PS) in which the unsaturation was systematically varied. Highly purified PKC alpha and epsilon were obtained using the baculovirus-insect cell expression system. It was shown that increased PC unsaturation elevated the activity of PKC alpha. By contrast, increasing the unsaturation of PS decreased the activity of PKC alpha, and to a lesser extent PKC epsilon. This result immediately rules out any single lipid bilayer physical parameter, such as lipid order, underlying the effect. It is proposed that while PC unsaturation effects are explainable on the basis of a contribution to membrane surface curvature stress, the effects of PS unsaturation may be due to specific protein-lipid interactions. Overall, the results indicate that altered phospholipid unsaturation in cell membranes that occurs in certain disease states such as chronic alcoholism, or by dietary manipulations, are likely to have profound effects on signal transduction pathways involving PKC and similar proteins.

  3. Regulation of cell polarity by cell adhesion receptors. (United States)

    Ebnet, Klaus; Kummer, Daniel; Steinbacher, Tim; Singh, Amrita; Nakayama, Masanori; Matis, Maja


    The ability of cells to polarize is an intrinsic property of almost all cells and is required for the devlopment of most multicellular organisms. To develop cell polarity, cells integrate various signals derived from intrinsic as well as extrinsic sources. In the recent years, cell-cell adhesion receptors have turned out as important regulators of cellular polarization. By interacting with conserved cell polarity proteins, they regulate the recruitment of polarity complexes to specific sites of cell-cell adhesion. By initiating intracellular signaling cascades at those sites, they trigger their specific subcellular activation. Not surprisingly, cell-cell adhesion receptors regulate diverse aspects of cell polarity, including apico-basal polarity in epithelial and endothelial cells, front-to-rear polarity in collectively migrating cells, and planar cell polarity during organ development. Here, we review the recent developments highlighting the central roles of cell-cell adhesion molecules in the development of cell polarity. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Tuning Collective Cell Migration by Cell-Cell Junction Regulation. (United States)

    Friedl, Peter; Mayor, Roberto


    Collective cell migration critically depends on cell-cell interactions coupled to a dynamic actin cytoskeleton. Important cell-cell adhesion receptor systems implicated in controlling collective movements include cadherins, immunoglobulin superfamily members (L1CAM, NCAM, ALCAM), Ephrin/Eph receptors, Slit/Robo, connexins and integrins, and an adaptive array of intracellular adapter and signaling proteins. Depending on molecular composition and signaling context, cell-cell junctions adapt their shape and stability, and this gradual junction plasticity enables different types of collective cell movements such as epithelial sheet and cluster migration, branching morphogenesis and sprouting, collective network migration, as well as coordinated individual-cell migration and streaming. Thereby, plasticity of cell-cell junction composition and turnover defines the type of collective movements in epithelial, mesenchymal, neuronal, and immune cells, and defines migration coordination, anchorage, and cell dissociation. We here review cell-cell adhesion systems and their functions in different types of collective cell migration as key regulators of collective plasticity. Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.

  5. Polybenzimidazole/Mxene composite membranes for intermediate temperature polymer electrolyte membrane fuel cells (United States)

    Fei, Mingming; Lin, Ruizhi; Deng, Yuming; Xian, Hongxi; Bian, Renji; Zhang, Xiaole; Cheng, Jigui; Xu, Chenxi; Cai, Dongyu


    This report demonstrated the first study on the use of a new 2D nanomaterial (Mxene) for enhancing membrane performance of intermediate temperature (>100 °C) polymer electrolyte membrane fuel cells (ITPEMFCs). In this study, a typical Ti3C2T x -MXene was synthesized and incorporated into polybenzimidazole (PBI)-based membranes by using a solution blending method. The composite membrane with 3 wt% Ti3C2T x -MXene showed the proton conductivity more than 2 times higher than that of pristine PBI membrane at the temperature range of 100 °C-170 °C, and led to substantial increase in maximum power density of fuel cells by ˜30% tested at 150 °C. The addition of Ti3C2T x -MXene also improved the mechanical properties and thermal stability of PBI membranes. At 3 wt% Ti3C2T x -MXene, the elongation at break of phosphoric acid doped PBI remained unaffected at 150 °C, and the tensile strength and Young’s modulus was increased by ˜150% and ˜160%, respectively. This study pointed out promising application of MXene in ITPEMFCs.

  6. Microbubble-based enhancement of radiation effect: Role of cell membrane ceramide metabolism.

    Directory of Open Access Journals (Sweden)

    Azza Al-Mahrouki

    Full Text Available Ultrasound (US stimulated microbubbles (MB is a new treatment approach that sensitizes cancer cells to radiation (XRT. The molecular pathways in this response remain unelucidated, however, previous data has supported a role for cell membrane-metabolism related pathways including an up regulation of UDP glycosyltransferase 8 (UGT8, which catalyzes the transfer of galactose to ceramide, a lipid that is associated with the induction of apoptotic signalling. In this study, the role of UGT8 in responses of prostate tumours to ultrasound-stimulated microbubble radiation enhancement therapy is investigated. Experiments were carried out with cells in vitro and tumours in vivo in which UGT8 levels had been up regulated or down regulated. Genetically modified PC3 cells were treated with XRT, US+MB, or a combination of XRT+US+MB. An increase in the immunolabelling of ceramide was observed in cells where UGT8 was down-regulated as opposed to cells where UGT8 was either not regulated or was up-regulated. Clonogenic assays have revealed a decreased level of cellular survival with the down-regulation of UGT8. Xenograft tumours generated from stably transfected PC3 cells were also treated with US+MB, XRT or US+MB+XRT. Histology demonstrated more cellular damage in tumours with down-regulated UGT8 in comparison with control tumours. In contrast, tumours with up-regulated UGT8 had less damage than control tumours. Power Doppler imaging indicated a reduction in the vascular index with UGT8 down-regulation and photoacoustic imaging revealed a reduction in oxygen saturation. This was contrary to when UGT8 was up regulated. The down regulation of UGT8 led to the accumulation of ceramide resulting in more cell death signalling and therefore, a greater enhancement of radiation effect when vascular disruption takes place through the use of ultrasound-stimulated microbubbles.

  7. Cell Membrane-Cloaked Nanoparticles for Targeted Therapeutics (United States)

    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

  8. Nonlinear electro-mechanobiological behavior of cell membrane during electroporation

    KAUST Repository

    Deng, Peigang


    A nonlinear electroporation (EP) model is proposed to study the electro-mechanobiological behavior of cell membrane during EP, by taking the nonlinear large deformation of the membrane into account. The proposed model predicts the critical transmembrane potential and the activation energy for EP, the equilibrium pore size, and the resealing process of the pore. Single-cell EP experiments using a micro EP chip were conducted on chicken red blood cells at different temperatures to determine the activation energy and the critical transmembrane potential for EP. The experimental results are in good agreement with the theoretical predictions. © 2012 American Institute of Physics.

  9. Cell-penetrating peptides for drug delivery across membrane barriers

    DEFF Research Database (Denmark)

    Foged, Camilla; Nielsen, Hanne Moerck


    During the last decade, cell-penetrating peptides have been investigated for their ability to overcome the plasma membrane barrier of mammalian cells for the intracellular or transcellular delivery of cargoes as diverse as low molecular weight drugs, imaging agents, oligonucleotides, peptides......-penetrating peptides as transmembrane drug delivery agents, according to the recent literature, and discusses critical issues and future challenges in relation to fully understanding the fundamental principles of the cell-penetrating peptide-mediated membrane translocation of cargoes and the exploitation......, proteins and colloidal carriers such as liposomes and polymeric nanoparticles. Their ability to cross biological membranes in a non-disruptive way without apparent toxicity is highly desired for increasing drug bioavailability. This review provides an overview of the application of cell...

  10. The formation of endosymbiotic membrane compartments: membrane identity markers and the regulation of vesicle trafficking

    NARCIS (Netherlands)

    Ivanov, S.


    In symbiosis of plants and arbuscular mycorrhizal fungi as well as in rhizobium-legume symbiosis the microbes are hosted intracellularly, inside specialized membrane compartments of the host. These membrane compartments are morphologically different but similar in function, since they control the

  11. The formation of endosymbiotic membrane compartments: membrane identity markers and the regulation of vesicle trafficking

    NARCIS (Netherlands)

    Ivanov, S.


    In symbiosis of plants and arbuscular mycorrhizal fungi as well as in rhizobium-legume symbiosis the microbes are hosted intracellularly, inside specialized membrane compartments of the host. These membrane compartments are morphologically different but similar in function, since they control

  12. Isotropic Versus Bipolar Functionalized Biomimetic Artificial Basement Membranes and Their Evaluation in Long-Term Human Cell Co-Culture. (United States)

    Rossi, Angela; Wistlich, Laura; Heffels, Karl-Heinz; Walles, Heike; Groll, Jürgen


    In addition to dividing tissues into compartments, basement membranes are crucial as cell substrates and to regulate cellular behavior. The development of artificial basement membranes is indispensable for the ultimate formation of functional engineered tissues; however, pose a challenge due to their complex structure. Herein, biodegradable electrospun polyester meshes are presented, exhibiting isotropic or bipolar bioactivation as a biomimetic and biofunctional model of the natural basement membrane. In a one-step preparation process, reactive star-shaped prepolymer additives, which generate a hydrophilic fiber surface, are electrospun with cell-adhesion-mediating peptides, derived from major components of the basement membrane. Human skin cells adhere to the functionalized meshes, and long-term co-culture experiments confirm that the artificial basement membranes recapitulate and preserve tissue specific functions. Several layers of immortalized human keratinocytes grow on the membranes, differentiating toward the surface and expressing typical epithelial markers. Fibroblasts migrate into the reticular lamina mimicking part of the mesh. Both cells types begin to produce extracellular matrix proteins and to remodel the initial membrane. It is shown at the example of skin that the artificial basement membrane design provokes biomimetic responses of different cell types and can thus be used as basis for the future development of basement membrane containing tissues. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Polyarylenethioethersulfone Membranes for Fuel Cells (Postprint) (United States)


    with deionized water in an attempt to completely remove the salts and then soxhlet -extracted in methanol for 72 h. Finally, it was vacuum dried at...were evaluated us- ing methods based on ASTM-D882 and ASTM-1004. SPTES-50 and Nafion samples both dry and wet were characterized. Samples of the...four-point-probe method provides membrane resistance without the effect of charge-transfer resistance and other nonohmic resis- tances. Proton

  14. Protein kinase Gin4 negatively regulates flippase function and controls plasma membrane asymmetry (United States)

    Roelants, Françoise M.; Su, Brooke M.; von Wulffen, Joachim; Ramachandran, Subramaniam; Sartorel, Elodie; Trott, Amy E.


    Plasma membrane function requires distinct leaflet lipid compositions. Two of the P-type ATPases (flippases) in yeast, Dnf1 and Dnf2, translocate aminoglycerophospholipids from the outer to the inner leaflet, stimulated via phosphorylation by cortically localized protein kinase Fpk1. By monitoring Fpk1 activity in vivo, we found that Fpk1 was hyperactive in cells lacking Gin4, a protein kinase previously implicated in septin collar assembly. Gin4 colocalized with Fpk1 at the cortical site of future bud emergence and phosphorylated Fpk1 at multiple sites, which we mapped. As judged by biochemical and phenotypic criteria, a mutant (Fpk111A), in which 11 sites were mutated to Ala, was hyperactive, causing increased inward transport of phosphatidylethanolamine. Thus, Gin4 is a negative regulator of Fpk1 and therefore an indirect negative regulator of flippase function. Moreover, we found that decreasing flippase function rescued the growth deficiency of four different cytokinesis mutants, which suggests that the primary function of Gin4 is highly localized control of membrane lipid asymmetry and is necessary for optimal cytokinesis. PMID:25646086

  15. Mouse endometrial stromal cells produce basement-membrane components

    DEFF Research Database (Denmark)

    Wewer, U M; Damjanov, A; Weiss, J


    . Mouse decidual cells isolated from 6- to 7-day pregnant uteri explanted in vitro continue to synthesize basement-membrane-like extracellular matrix. Using immunohistochemistry and metabolic labeling followed by immunoprecipitation, SDS-PAGE, and fluorography, it was shown that the decidual cells...

  16. Development of new membrane materials for direct methanol fuel cells

    NARCIS (Netherlands)

    Yildirim, M.H.


    Development of new membrane materials for direct methanol fuel cells Direct methanol fuel cells (DMFCs) can convert the chemical energy of a fuel directly into electrical energy with high efficiency and low emission of pollutants. DMFCs can be used as the power sources to portable electronic devices

  17. Polymer electrolyte membrane fuel cell control with feed-forward ...

    African Journals Online (AJOL)

    Feed-forward and feedback control is developed in this work for Polymer electrolyte membrane (PEM) fuel cell stacks. The feed-forward control is achieved using different methods, including look-up table, fuzzy logic and neural network, to improve the fuel cell stack breathing control and prevent the problem of oxygen ...

  18. The influence of platelet membranes on tumour cell behaviour. (United States)

    Coupland, L A; Hindmarsh, E J; Gardiner, E E; Parish, C R


    The significant role of platelets in the protection of tumour cells from immune attack and shear forces and the promotion of tumour cell extravasation from the bloodstream in the process of haematogenous metastasis have been extensively studied. The role of platelets, and in particular platelet membranes, in the promotion of a more metastatic phenotype in tumour cells is a more recent and, therefore, less well-recognised area of research. This review article summarises studies that have focused on the impact of tumour cell interactions with platelets and platelet membranes on tumour cell behaviour in vitro and in vivo. Furthermore, the gene expression changes that occur within tumour cells following contact with platelet membranes are also extensively reviewed. Overall, the interaction of platelet membranes with tumour cells results in a more invasive phenotype and the promotion of epithelial to mesenchymal transition with our own genetic studies revealing that matrix metalloproteinase-1, plasminogen activator inhibitor-1 and interleukin-8 are globally upregulated in a range of tumour cell lines.

  19. Advanced nanocomposite membranes for fuel cell applications: a comprehensive review

    Directory of Open Access Journals (Sweden)

    Kolsoum Pourzare


    Full Text Available Combination of inorganic fillers into organic polymer membranes (organic–inorganic hybrid membranes has drawn a significant deal of attention over the last few decades. This is because of the incorporated influence of the organic and inorganic phases towards proton conductivity and membrane stability, in addition to cost decline, improved water retention property, and also suppressing fuel crossover by increasing the transport pathway tortuousness. The preparation methods of the composite membranes and the intrinsic characteristics of the used particles as filler, such as size, type, surface acidity, shape, and their interactions with the polymer matrix can significantly affect the properties of the resultant matrix. The membranes currently used in proton exchange membrane fuel cells (PEMFCs are perfluorinated polymers containing sulfonic acid, such as Nafion®. Although these membranes possess superior properties, such as high proton conductivity and acceptable chemical, mechanical, and thermal stability, they suffer from several disadvantages such as water management, CO poisoning, and fuel crossover. Organic-inorganic nanocomposite PEMs offer excellent potentials for overcoming these shortcomings in order to achieve improved FC performance. Various inorganic fillers for the fabrication of composite membranes have been comprehensively reviewed in the present article. Moreover, the properties of polymer composites containing different nanoparticles have been thoroughly discussed.

  20. Towards Extrusion of Ionomers to Process Fuel Cell Membranes

    Directory of Open Access Journals (Sweden)

    Jean-Yves Sanchez


    Full Text Available While Proton Exchange Membrane Fuel Cell (PEMFC membranes are currently prepared by film casting, this paper demonstrates the feasibility of extrusion, a solvent-free alternative process. Thanks to water-soluble process-aid plasticizers, duly selected, it was possible to extrude acidic and alkaline polysulfone ionomers. Additionally, the feasibility to extrude composites was demonstrated. The impact of the plasticizers on the melt viscosity was investigated. Following the extrusion, the plasticizers were fully removed in water. The extrusion was found to impact neither on the ionomer chains, nor on the performances of the membrane. This environmentally friendly process was successfully validated for a variety of high performance ionomers.

  1. Regulated membrane remodeling by Mic60 controls formation of mitochondrial crista junctions. (United States)

    Hessenberger, Manuel; Zerbes, Ralf M; Rampelt, Heike; Kunz, Séverine; Xavier, Audrey H; Purfürst, Bettina; Lilie, Hauke; Pfanner, Nikolaus; van der Laan, Martin; Daumke, Oliver


    The mitochondrial contact site and cristae organizing system (MICOS) is crucial for the formation of crista junctions and mitochondrial inner membrane architecture. MICOS contains two core components. Mic10 shows membrane-bending activity, whereas Mic60 (mitofilin) forms contact sites between inner and outer membranes. Here we report that Mic60 deforms liposomes into thin membrane tubules and thus displays membrane-shaping activity. We identify a membrane-binding site in the soluble intermembrane space-exposed part of Mic60. This membrane-binding site is formed by a predicted amphipathic helix between the conserved coiled-coil and mitofilin domains. The mitofilin domain negatively regulates the membrane-shaping activity of Mic60. Binding of Mic19 to the mitofilin domain modulates this activity. Membrane binding and shaping by the conserved Mic60-Mic19 complex is crucial for crista junction formation, mitochondrial membrane architecture and efficient respiratory activity. Mic60 thus plays a dual role by shaping inner membrane crista junctions and forming contact sites with the outer membrane.

  2. Numerical modeling transport phenomena in proton exchange membrane fuel cells (United States)

    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

  3. Ion Channels Involved in Cell Volume Regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay


    This mini review outlines studies of cell volume regulation in two closely related mammalian cell lines: nonadherent Ehrlich ascites tumour cells (EATC) and adherent Ehrlich Lettre ascites (ELA) cells. Focus is on the regulatory volume decrease (RVD) that occurs after cell swelling, the volume...

  4. Cell membrane damage by iron nanoparticles: an invitro study

    Directory of Open Access Journals (Sweden)

    Gelare Hajsalimi


    Full Text Available Application of nanotechnology in medicinal and biological fields has attracted a great interest in the recent yeras. In this paper the cell membrane leakage induced by iron nanoparticles (Fe-NP against PC12 cell line which is known as a model of nervous system cell line was investigated by the lactate dehydrogenase (LDH test. Therefore, PC12 cells were incubated with different concentration of Fe-NP and test was performed after 48h of incubation of the cells with Fe-NP. The resulting data showed that the Fe-NP induced the damage of PC12 cell membrane in a concentration dependent manner. Hence, it may be concluded that the different cytotoxicty effect of NPs may be referred to the concentration of NPs, type of the NPs and the cells. Indeed, the kind of cytotoxic impacts of NPs on the cells can be reduced by the considering of above-mentioned parameters. The resulting data showed that the Fe-NP induced the damage of PC12 cell membrane in a concentration dependent manner. Hence, it may be concluded that the different cytotoxicty effect of NPs may be referred to the concentration of NPs, type of the NPs and the cells. Indeed, the kind of cytotoxic impacts of NPs on the cells can be reduced by the considering of above-mentioned parameters.

  5. Catalyst Degradation in High Temperature Proton Exchange Membrane Fuel Cells Based on Acid Doped Polybenzimidazole Membranes

    DEFF Research Database (Denmark)

    Cleemann, Lars Nilausen; Buazar, F.; Li, Qingfeng


    confirmed by the post TEM and XRD analysis. A strong dependence of the fuel cell performance degradation on the catalyst supports was observed. Graphitization of the carbon blacks improved the stability and catalyst durability though at the expense of a significant decrease in the specific surface area......Degradation of carbon supported platinum catalysts is a major failure mode for the long term durability of high temperature proton exchange membrane fuel cells based on phosphoric acid doped polybenzimidazole membranes. With Vulcan carbon black as a reference, thermally treated carbon black...... and multi‐walled carbon nanotubes were used as supports for electrode catalysts and evaluated in accelerated durability tests under potential cycling at 150 °C. Measurements of open circuit voltage, area specific resistance and hydrogen permeation through the membrane were carried out, indicating little...

  6. Electrically Conductive, Hydrophilic Porous Membrane for Fuel Cell Applications, Phase I (United States)

    National Aeronautics and Space Administration — This Phase I effort seeks to produce a conductive polyethersulfone (PES) microporous membrane for fuel cell water management applications. This membrane will...

  7. Regulated portals of entry into the cell (United States)

    Conner, Sean D.; Schmid, Sandra L.


    The plasma membrane is the interface between cells and their harsh environment. Uptake of nutrients and all communication among cells and between cells and their environment occurs through this interface. `Endocytosis' encompasses several diverse mechanisms by which cells internalize macromolecules and particles into transport vesicles derived from the plasma membrane. It controls entry into the cell and has a crucial role in development, the immune response, neurotransmission, intercellular communication, signal transduction, and cellular and organismal homeostasis. As the complexity of molecular interactions governing endocytosis are revealed, it has become increasingly clear that it is tightly coordinated and coupled with overall cell physiology and thus, must be viewed in a broader context than simple vesicular trafficking.

  8. Importance of balancing membrane and electrode water in anion exchange membrane fuel cells (United States)

    Omasta, T. J.; Wang, L.; Peng, X.; Lewis, C. A.; Varcoe, J. R.; Mustain, W. E.


    Anion exchange membrane fuel cells (AEMFCs) offer several potential advantages over proton exchange membrane fuel cells (PEMFCs), most notably to overcome the cost barrier that has slowed the growth and large scale implementation of fuel cells for transportation. However, limitations in performance have held back AEMFCs, specifically in the areas of stability, carbonation, and maximum achievable current and power densities. In order for AEMFCs to contend with PEMFCs for market viability, it is necessary to realize a competitive cell performance. This work demonstrates a new benchmark for a H2/O2 AEMFC with a peak power density of 1.4 W cm-2 at 60 °C. This was accomplished by taking a more precise look at balancing necessary membrane hydration while preventing electrode flooding, which somewhat surprisingly can occur both at the anode and the cathode. Specifically, radiation-grafted ETFE-based anion exchange membranes and anion exchange ionomer powder, functionalized with benchmark benzyltrimethylammonium groups, were utilized to examine the effects of the following parameters on AEMFC performance: feed gas flow rate, the use of hydrophobic vs. hydrophilic gas diffusion layers, and gas feed dew points.

  9. Estimation of membrane hydration status for standby proton exchange membrane fuel cell systems by impedance measurement

    DEFF Research Database (Denmark)

    Bidoggia, Benoit; Rugholt, Mark; Nielsen, Morten Busk


    Fuel cells are getting growing interest in both backup systems and electric vehicles. Although these systems are characterized by long periods of inactivity, they must be able to start at any instant in the shortest time. However, the membrane of which PEMFCs are made tends to dry out when not in...

  10. In Plant and Animal Cells, Detergent-Resistant Membranes Do Not Define Functional Membrane Rafts

    Czech Academy of Sciences Publication Activity Database

    Tanner, W.; Malínský, Jan; Opekarová, Miroslava


    Roč. 23, č. 4 (2011), s. 1191-1193 ISSN 1040-4651 Institutional research plan: CEZ:AV0Z50390512; CEZ:AV0Z50200510 Keywords : plasma-membrane * lipod rafts * proteins Subject RIV: EA - Cell Biology Impact factor: 8.987, year: 2011

  11. Fusion Pore Diameter Regulation by Cations Modulating Local Membrane Anisotropy

    Directory of Open Access Journals (Sweden)

    Doron Kabaso


    Full Text Available The fusion pore is an aqueous channel that is formed upon the fusion of the vesicle membrane with the plasma membrane. Once the pore is open, it may close again (transient fusion or widen completely (full fusion to permit vesicle cargo discharge. While repetitive transient fusion pore openings of the vesicle with the plasma membrane have been observed in the absence of stimulation, their frequency can be further increased using a cAMP-increasing agent that drives the opening of nonspecific cation channels. Our model hypothesis is that the openings and closings of the fusion pore are driven by changes in the local concentration of cations in the connected vesicle. The proposed mechanism of fusion pore dynamics is considered as follows: when the fusion pore is closed or is extremely narrow, the accumulation of cations in the vesicle (increased cation concentration likely leads to lipid demixing at the fusion pore. This process may affect local membrane anisotropy, which reduces the spontaneous curvature and thus leads to the opening of the fusion pore. Based on the theory of membrane elasticity, we used a continuum model to explain the rhythmic opening and closing of the fusion pore.

  12. Comprehensive quantitative comparison of the membrane proteome, phosphoproteome, and sialiome of human embryonic and neural stem cells

    DEFF Research Database (Denmark)

    Melo-Braga, Marcella Nunes; Schulz, Melanie; Liu, Qiuyue


    Human embryonic stem cells (hESCs) can differentiate into neural stem cells (NSCs), which can further be differentiated into neurons and glia cells. Therefore, these cells have huge potential as source for treatment of neurological diseases. Membrane-associated proteins are very important......ESCs and NSCs as well as to investigate potential new markers for these two cell stages, we performed large-scale quantitative membrane-proteomic of hESCs and NSCs. This approach employed membrane purification followed by peptide dimethyl labeling and peptide enrichment to study the membrane subproteome as well...... in which 78% of phosphopeptides were identified with ≥99% confidence in site assignment and 1810 unique formerly sialylated N-linked glycopeptides. Several proteins were identified as significantly regulated in hESCs and NSC, including proteins involved in the early embryonic and neural development...

  13. Nanofiber Composite Membranes for Alkaline Fuel Cells: Generation of Compositional, Morphological, and Functional Property Relationships (United States)


    properties of nanofiber composite anion-exchange membranes for alkaline fuel cells. A new membrane fabrication strategy, utilizing polymer fiber...Approved for Public Release; Distribution Unlimited Final Report: Nanofiber Composite Membranes for Alkaline Fuel Cells: Generation of Compositional...Park, NC 27709-2211 nanofibers, electrospinning, composite membranes, alkaline fuel cells REPORT DOCUMENTATION PAGE 11. SPONSOR/MONITOR’S REPORT NUMBER

  14. Cell Receptor-Basement Membrane Interactions in Health and Disease: a Kidney-Centric View (United States)

    Borza, Corina M.; Chen, Xiwu; Zent, Roy; Pozzi, Ambra


    Cell-extracellular matrix (ECM) interactions are essential for tissue development, homeostasis, and response to injury. Basement membranes (BMs) are specialized ECMs that separate epithelial or endothelial cells from stromal components and interact with cells via cellular receptors, including integrins and discoidin domain receptors. Disruption of cell-BM interactions due to either injury or genetic defects in either the ECM components or cellular receptors often lead to irreversible tissue injury and loss of organ function. Animal models that lack specific BM components or receptors either globally or in selective tissues have been used to help with our understanding of the molecular mechanisms whereby cell-BM interactions regulate organ function in physiological and pathological conditions. We review recently published work on animal models that explore how cell-BM interactions regulate kidney homeostasis in both health and disease. PMID:26610916

  15. DNA repair genes RAD52 and SRS2, a cell wall synthesis regulator gene SMI1, and the membrane sterol synthesis scaffold gene ERG28 are important in efficient Agrobacterium-mediated yeast transformation with chromosomal T-DNA. (United States)

    Ohmine, Yuta; Satoh, Yukari; Kiyokawa, Kazuya; Yamamoto, Shinji; Moriguchi, Kazuki; Suzuki, Katsunori


    Plant pathogenic Agrobacterium strains can transfer T-DNA regions of their Ti plasmids to a broad range of eukaryotic hosts, including fungi, in vitro. In the recent decade, the yeast Saccharomyces cerevisiae is used as a model host to reveal important host proteins for the Agrobacterium-mediated transformation (AMT). Further investigation is required to understand the fundamental mechanism of AMT, including interaction at the cell surface, to expand the host range, and to develop new tools. In this study, we screened a yeast mutant library for low AMT mutant strains by advantage of a chromosome type T-DNA, which transfer is efficient and independent on integration into host chromosome. By the mutant screening, we identified four mutant strains (srs2Δ, rad52Δ, smi1Δ and erg28Δ), which showed considerably low AMT efficiency. Structural analysis of T-DNA product replicons in AMT colonies of mutants lacking each of the two DNA repair genes, SRS2 and RAD52, suggested that the genes act soon after T-DNA entry for modification of the chromosomal T-DNA to stably maintain them as linear replicons and to circularize certain T-DNA simultaneously. The cell wall synthesis regulator SMI1 might have a role in the cell surface interaction between the donor and recipient cells, but the smi1Δ mutant exhibited pleiotropic effect, i.e. low effector protein transport as well as low AMT for the chromosomal T-DNA, but relatively high AMT for integrative T-DNAs. The ergosterol synthesis regulator/enzyme-scaffold gene ERG28 probably contributes by sensing a congested environment, because growth of erg28Δ strain was unaffected by the presence of donor bacterial cells, while the growth of the wild-type and other mutant yeast strains was suppressed by their presence. RAD52 and the DNA helicase/anti-recombinase gene SRS2 are necessary to form and maintain artificial chromosomes through the AMT of chromosomal T-DNA. A sterol synthesis scaffold gene ERG28 is important in the high

  16. Regulation of the V-ATPase along the endocytic pathway occurs through reversible subunit association and membrane localization.

    Directory of Open Access Journals (Sweden)

    Céline Lafourcade


    Full Text Available The lumen of endosomal organelles becomes increasingly acidic when going from the cell surface to lysosomes. Luminal pH thereby regulates important processes such as the release of internalized ligands from their receptor or the activation of lysosomal enzymes. The main player in endosomal acidification is the vacuolar ATPase (V-ATPase, a multi-subunit transmembrane complex that pumps protons from the cytoplasm to the lumen of organelles, or to the outside of the cell. The active V-ATPase is composed of two multi-subunit domains, the transmembrane V(0 and the cytoplasmic V(1. Here we found that the ratio of membrane associated V(1/Vo varies along the endocytic pathway, the relative abundance of V(1 being higher on late endosomes than on early endosomes, providing an explanation for the higher acidity of late endosomes. We also found that all membrane-bound V-ATPase subunits were associated with detergent resistant membranes (DRM isolated from late endosomes, raising the possibility that association with lipid-raft like domains also plays a role in regulating the activity of the proton pump. In support of this, we found that treatment of cells with U18666A, a drug that leads to the accumulation of cholesterol in late endosomes, affected acidification of late endosome. Altogether our findings indicate that the activity of the vATPase in the endocytic pathway is regulated both by reversible association/dissociation and the interaction with specific lipid environments.

  17. Nafion®/ODF-silica composite membranes for medium temperature proton exchange membrane fuel cells

    KAUST Repository

    Treekamol, Yaowapa


    A series of composite membranes were prepared by dispersing fluorinated polyoxadiazole oligomer (ODF)-functionalized silica nanoparticles in a Nafion matrix. Both melt-extrusion and solvent casting processes were explored. Ion exchange capacity, conductivity, water uptake and dimensional stability, thermal stability and morphology were characterized. The inclusion of functionalized nanoparticles proved advantageous, mainly due to a physical crosslinking effect and better water retention, with functionalized nanoparticles performing better than the pristine silica particles. For the same filler loading, better nanoparticle dispersion was achieved for solvent-cast membranes, resulting in higher proton conductivity. Filler agglomeration, however,was more severe for solvent-castmembranes at loadings beyond 5wt.%. The composite membranes showed excellent thermal stability, allowing for operation in medium temperature PEM fuel cells. Fuel cell performance of the compositemembranesdecreaseswithdecreasing relativehumidity, but goodperformance values are still obtained at 34% RHand 90 °C,with the best results obtained for solvent castmembranes loaded with 10 wt.% ODF-functionalized silica. Hydrogen crossover of the composite membranes is higher than that forpureNafion membranes,possiblydue toporosityresulting fromsuboptimalparticle- matrixcompatibility. © 2013 Crown Copyright and Elsevier BV. All rights reserved.

  18. Determination of apical membrane polarity in mammary epithelial cell cultures: The role of cell-cell, cell-substratum, and membrane-cytoskeleton interactions

    Energy Technology Data Exchange (ETDEWEB)

    Parry, G.; Beck, J.C.; Moss, L.; Bartley, J. (Lawrence Berkeley Lab., CA (United States)); Ojakian, G.K. (State Univ. of New York, Brooklyn (United States))


    The membrane glycoprotein, PAS-O, is a major differentiation antigen on mammary epithelial cells and is located exclusively in the apical domain of the plasma membrane. The authors have used 734B cultured human mammary carcinoma cells as a model system to study the role of tight junctions, cell-substratum contacts, and submembranous cytoskeletal elements in restricting PAS-O to the apical membrane. Immunofluorescence and immunoelectronmicroscopy experiments demonstrated that while tight junctions demarcate PAS-O distribution in confluent cultures, apical polarity could be established at low culture densities when cells could not form tight junctions with neighboring cells. They suggest, then, that interactions between vitronectin and its receptor, are responsible for establishment of membrane domains in the absence of tight junctions. The role of cytoskeletal elements in restricting PAS-O distribution was examined by treating cultures with cytochalasin D, colchicine, or acrylamide. Cytochalasin D led to a redistribution of PAS0O while colchicine and acrylamide did not. They hypothesize that PAS-O is restricted to the apical membrane by interactions with a microfilament network and that the cytoskeletal organization is dependent upon cell-cell and cell-substratum interactions.

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

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


    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.

  20. Red Blood Cell Membrane-Cloaked Nanoparticles For Drug Delivery (United States)

    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.

  1. Microstructured Electrolyte Membranes to Improve Fuel Cell Performance (United States)

    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

  2. Poly (Ethylene-Alit-Tetrafluoroethylene) Based Membranes For Fuel Cells: Synthesis And Fuel Cell Performance

    Energy Technology Data Exchange (ETDEWEB)

    Alkan Guersel, S.; Gubler, L.; Scherer, G.G.


    Proton exchange membranes have been synthesized by pre-irradiation grafting of styrene onto poly (ethylene-alt-tetrafluoroethylene) (ETFE) in the presence of divinyl benzene (DVB) as the cross linker and characterized ex-situ for their fuel cell relevant properties. The optimum graft level was determined as between 20 and 30 %. ETFE based membranes exhibited encouraging fuel cell performance yet, there is room for improvement through optimization of the membrane-electrode interface. (author)

  3. Role of membranes and membrane reactors in the hydrogen supply of fuel cells for transports

    Energy Technology Data Exchange (ETDEWEB)

    Julbe, A.; Guizard, Ch. [Institut Europeen des Membranes, UMII, Lab. des Materiaux et des Procedes Membranaires, CNRS UMR 5635, 34 - Montpellier (France)


    Production, storage and supply of high-purity hydrogen as a clean and efficient fuel is central to fuel cells technology, in particular in vehicle traction. Actually, technologies for handling liquefied or gaseous hydrogen in transports are not available so that a number of alternative fuels are considered with the aim of in-situ generation of hydrogen through catalytic processes. The integrated concept of membrane reactors (MRs) can greatly benefit to these technologies. Particular emphasis is put on inorganic membranes and their role in MRs performance for H{sub 2} production.

  4. Radiation effects on membranes - 1. Cellular permeability and cell survival

    International Nuclear Information System (INIS)

    Khare, S.; Jayakumar, A.; Trivedi, A.; Kesavan, P.C.; Prasad, R.


    The effect of various doses of γ 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 γ 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 γ radiation

  5. Translocation across biological membranes: activity, structure and regulation of transporters

    NARCIS (Netherlands)

    Ruiz, Stephanie


    Alle cellen zijn omsloten door biologische membranen met selectief doorlatende membrane, die het mogelijk maken om omstandigheden "binnen" en "buiten" de cel te handhaven. De gereguleerde beweging van moleculen door deze membranen is nodig voor talrijke essentiële processen, waaronder het verkrijgen

  6. Lipids that determine detergent resistance of MDCK cell membrane fractions. (United States)

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


    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. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  7. Membrane and MEA Development in Polymer Electrolyte Fuel Cells (United States)

    Trogadas, Panagiotis; Ramani, Vijay

    The polymer electrolyte fuel cell (PEFC) is based on Nafion polymer membranes operating at a temperature of 80°C. The main characteristics (structure and properties) and problems of Nafion-based PEFC technology are discussed. The primary drawbacks of Nafion membranes are poor conductivity at low relative humidities (and consequently at temperatures >100°C and ambient pressure) and large crossover of methanol in direct methanol fuel cell (DMFC) applications. These drawbacks have prompted an extensive effort to improve the properties of Nafion and identify alternate materials to replace Nafion. Polymer electrolyte membranes (PEMs) are classified in modified Nafion, membranes based on functionalized non-fluorinated backbones and acid-base polymer systems. Perhaps the most widely employed approach is the addition of inorganic additives to Nafion membranes to yield organic/inorganic composite membranes. Four major types of inorganic additives that have been studied (zirconium phosphates, heteropolyacids, metal hydrogen sulfates, and metal oxides) are reviewed in the following. DMFC and H2/O2 (air) cells based on modified Nafion membranes have been successfully operated at temperatures up to 120°C under ambient pressure and up to 150°C under 3-5 atm. Membranes based on functionalized non-fluorinated backbones are potentially promising for high-temperature operation. High conductivities have been obtained at temperatures up to 180°C. The final category of polymeric PEMs comprises non-functionalized polymers with basic character doped with proton-conducting acids such as phosphoric acid. The advanced features include high CO tolerance and thermal management. The advances made in the fabrication of electrodes for PEM fuel cells from the PTFE-bound catalyst layers of almost 20 years ago to the present technology are briefly discussed. There are two widely employed electrode designs: (1) PTFE-bound, and (2) thin-film electrodes. Emerging methods include those featuring

  8. Optical Trapping Techniques Applied to the Study of Cell Membranes (United States)

    Morss, Andrew J.

    Optical tweezers allow for manipulating micron-sized objects using pN level optical forces. In this work, we use an optical trapping setup to aid in three separate experiments, all related to the physics of the cellular membrane. In the first experiment, in conjunction with Brian Henslee, we use optical tweezers to allow for precise positioning and control of cells in suspension to evaluate the cell size dependence of electroporation. Theory predicts that all cells porate at a transmembrane potential VTMof roughly 1 V. The Schwann equation predicts that the transmembrane potential depends linearly on the cell radius r, thus predicting that cells should porate at threshold electric fields that go as 1/r. The threshold field required to induce poration is determined by applying a low voltage pulse to the cell and then applying additional pulses of greater and greater magnitude, checking for poration at each step using propidium iodide dye. We find that, contrary to expectations, cells do not porate at a constant value of the transmembrane potential but at a constant value of the electric field which we find to be 692 V/cm for K562 cells. Delivering precise dosages of nanoparticles into cells is of importance for assessing toxicity of nanoparticles or for genetic research. In the second experiment, we conduct nano-electroporation—a novel method of applying precise doses of transfection agents to cells—by using optical tweezers in conjunction with a confocal microscope to manipulate cells into contact with 100 nm wide nanochannels. This work was done in collaboration with Pouyan Boukany of Dr. Lee's group. The small cross sectional area of these nano channels means that the electric field within them is extremely large, 60 MV/m, which allows them to electrophoretically drive transfection agents into the cell. We find that nano electroporation results in excellent dose control (to within 10% in our experiments) compared to bulk electroporation. We also find that

  9. Lysophosphatidylcholine Induces Taurine Release from HeLa Cells

    DEFF Research Database (Denmark)

    Lambert, Ian H.; Falktoft, Birgitte


    Cell volume regulation, Membrane permeabilization, Vitamin E, Tyrosine phosphorylation, Lysophospholipids......Cell volume regulation, Membrane permeabilization, Vitamin E, Tyrosine phosphorylation, Lysophospholipids...

  10. Membrane with internal passages to permit fluid flow and an electrochemical cell containing the same (United States)

    Cisar, Alan J. (Inventor); Gonzalez-Martin, Anuncia (Inventor); Hitchens, G. Duncan (Inventor); Murphy, Oliver J. (Inventor)


    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.

  11. Live cell plasma membranes do not exhibit a miscibility phase transition over a wide range of temperatures. (United States)

    Lee, Il-Hyung; Saha, Suvrajit; Polley, Anirban; Huang, Hector; Mayor, Satyajit; Rao, Madan; Groves, Jay T


    Lipid/cholesterol mixtures derived from cell membranes as well as their synthetic reconstitutions exhibit well-defined miscibility phase transitions and critical phenomena near physiological temperatures. This suggests that lipid/cholesterol-mediated phase separation plays a role in the organization of live cell membranes. However, macroscopic lipid-phase separation is not generally observed in cell membranes, and the degree to which properties of isolated lipid mixtures are preserved in the cell membrane remain unknown. A fundamental property of phase transitions is that the variation of tagged particle diffusion with temperature exhibits an abrupt change as the system passes through the transition, even when the two phases are distributed in a nanometer-scale emulsion. We support this using a variety of Monte Carlo and atomistic simulations on model lipid membrane systems. However, temperature-dependent fluorescence correlation spectroscopy of labeled lipids and membrane-anchored proteins in live cell membranes shows a consistently smooth increase in the diffusion coefficient as a function of temperature. We find no evidence of a discrete miscibility phase transition throughout a wide range of temperatures: 14-37 °C. This contrasts the behavior of giant plasma membrane vesicles (GPMVs) blebbed from the same cells, which do exhibit phase transitions and macroscopic phase separation. Fluorescence lifetime analysis of a DiI probe in both cases reveals a significant environmental difference between the live cell and the GPMV. Taken together, these data suggest the live cell membrane may avoid the miscibility phase transition inherent to its lipid constituents by actively regulating physical parameters, such as tension, in the membrane.

  12. FlnA binding to PACSIN2 F-BAR domain regulates membrane tubulation in megakaryocytes and platelets. (United States)

    Begonja, Antonija Jurak; Pluthero, Fred G; Suphamungmee, Worawit; Giannini, Silvia; Christensen, Hilary; Leung, Richard; Lo, Richard W; Nakamura, Fumihiko; Lehman, William; Plomann, Markus; Hoffmeister, Karin M; Kahr, Walter H A; Hartwig, John H; Falet, Hervé


    Bin-Amphiphysin-Rvs (BAR) and Fes-CIP4 homology BAR (F-BAR) proteins generate tubular membrane invaginations reminiscent of the megakaryocyte (MK) demarcation membrane system (DMS), which provides membranes necessary for future platelets. The F-BAR protein PACSIN2 is one of the most abundant BAR/F-BAR proteins in platelets and the only one reported to interact with the cytoskeletal and scaffold protein filamin A (FlnA), an essential regulator of platelet formation and function. The FlnA-PACSIN2 interaction was therefore investigated in MKs and platelets. PACSIN2 associated with FlnA in human platelets. The interaction required FlnA immunoglobulin-like repeat 20 and the tip of PACSIN2 F-BAR domain and enhanced PACSIN2 F-BAR domain membrane tubulation in vitro. Most human and wild-type mouse platelets had 1 to 2 distinct PACSIN2 foci associated with cell membrane GPIbα, whereas Flna-null platelets had 0 to 4 or more foci. Endogenous PACSIN2 and transfected enhanced green fluorescent protein-PACSIN2 were concentrated in midstage wild-type mouse MKs in a well-defined invagination of the plasma membrane reminiscent of the initiating DMS and dispersed in the absence of FlnA binding. The DMS appeared less well defined, and platelet territories were not readily visualized in Flna-null MKs. We conclude that the FlnA-PACSIN2 interaction regulates membrane tubulation in MKs and platelets and likely contributes to DMS formation. © 2015 by The American Society of Hematology.

  13. Cysteines and N-Glycosylation Sites Conserved among All Alphaherpesviruses Regulate Membrane Fusion in Herpes Simplex Virus 1 Infection. (United States)

    Rider, Paul J F; Naderi, Misagh; Bergeron, Scott; Chouljenko, Vladimir N; Brylinski, Michal; Kousoulas, Konstantin G


    Neurotropism is a defining characteristic of alphaherpesvirus pathogenicity. Glycoprotein K (gK) is a conserved virion glycoprotein of all alphaherpesviruses that is not found in other herpesvirus subfamilies. The extracellular amino terminus of gK has been shown to be important to the ability of the prototypic alphaherpesvirus herpes simplex virus 1 (HSV-1) to enter neurons via axonal termini. Here, we determined the role of the two conserved N-linked glycosylation (N48 and N58) sites of gK in virus-induced cell fusion and replication. We found that N-linked glycosylation is important to the regulation of HSV-1-induced membrane fusion since mutating N58 to alanine (N58A) caused extensive virus-induced cell fusion. Due to the known contributions of N-linked glycosylation to protein processing and correct disulfide bond formation, we investigated whether the conserved extracellular cysteine residues within the amino terminus of gK contributed to the regulation of HSV-1-induced membrane fusion. We found that mutation of C37 and C114 residues led to a gK-null phenotype characterized by very small plaque formation and drastic reduction in infectious virus production, while mutation of C82 and C243 caused extensive virus-induced cell fusion. Comparison of N-linked glycosylation and cysteine mutant replication kinetics identified disparate effects on infectious virion egress from infected cells. Specifically, cysteine mutations caused defects in the accumulation of infectious virus in both the cellular and supernatant fractions, while glycosylation site mutants did not adversely affect virion egress from infected cells. These results demonstrate a critical role for the N glycosylation sites and cysteines for the structure and function of the amino terminus of gK. IMPORTANCE We have previously identified important entry and neurotropic determinants in the amino terminus of HSV-1 glycoprotein K (gK). Alphaherpesvirus-mediated membrane fusion is a complex and highly

  14. Durable, Low-cost, Improved Fuel Cell Membranes

    Energy Technology Data Exchange (ETDEWEB)

    Chris Roger; David Mountz; Wensheng He; Tao Zhang


    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

  15. Vimentin is involved in regulation of mitochondrial motility and membrane potential by Rac1

    Directory of Open Access Journals (Sweden)

    Elena A. Matveeva


    Full Text Available In this study we show that binding of mitochondria to vimentin intermediate filaments (VIF is regulated by GTPase Rac1. The activation of Rac1 leads to a redoubling of mitochondrial motility in murine fibroblasts. Using double-mutants Rac1(G12V, F37L and Rac1(G12V, Y40H that are capable to activate different effectors of Rac1, we show that mitochondrial movements are regulated through PAK1 kinase. The involvement of PAK1 kinase is also confirmed by the fact that expression of its auto inhibitory domain (PID blocks the effect of activated Rac1 on mitochondrial motility. The observed effect of Rac1 and PAK1 kinase on mitochondria depends on phosphorylation of the Ser-55 of vimentin. Besides the effect on motility Rac1 activation also decreases the mitochondrial membrane potential (MMP which is detected by ∼20% drop of the fluorescence intensity of mitochondria stained with the potential sensitive dye TMRM. One of important consequences of the discovered regulation of MMP by Rac1 and PAK1 is a spatial differentiation of mitochondria in polarized fibroblasts: at the front of the cell they are less energized (by ∼25% than at the rear part.

  16. Minireview: Prolactin Regulation of Adult Stem Cells (United States)

    Sackmann-Sala, Lucila; Guidotti, Jacques-Emmanuel


    Adult stem/progenitor cells are found in many tissues, where their primary role is to maintain homeostasis. Recent studies have evaluated the regulation of adult stem/progenitor cells by prolactin in various target tissues or cell types, including the mammary gland, the prostate, the brain, the bone marrow, the hair follicle, and colon cancer cells. Depending on the tissue, prolactin can either maintain stem cell quiescence or, in contrast, promote stem/progenitor cell expansion and push their progeny towards differentiation. In many instances, whether these effects are direct or involve paracrine regulators remains debated. This minireview aims to overview the current knowledge in the field. PMID:25793405

  17. Electrospun fiber membranes enable proliferation of genetically modified cells (United States)

    Borjigin, Mandula; Eskridge, Chris; Niamat, Rohina; Strouse, Bryan; Bialk, Pawel; Kmiec, Eric B


    Polycaprolactone (PCL) and its blended composites (chitosan, gelatin, and lecithin) are well-established biomaterials that can enrich cell growth and enable tissue engineering. However, their application in the recovery and proliferation of genetically modified cells has not been studied. In the study reported here, we fabricated PCL-biomaterial blended fiber membranes, characterized them using physicochemical techniques, and used them as templates for the growth of genetically modified HCT116-19 colon cancer cells. Our data show that the blended polymers are highly miscible and form homogenous electrospun fiber membranes of uniform texture. The aligned PCL nanofibers support robust cell growth, yielding a 2.5-fold higher proliferation rate than cells plated on standard plastic plate surfaces. PCL-lecithin fiber membranes yielded a 2.7-fold higher rate of proliferation, while PCL-chitosan supported a more modest growth rate (1.5-fold higher). Surprisingly, PCL-gelatin did not enhance cell proliferation when compared to the rate of cell growth on plastic surfaces. PMID:23467983

  18. Materials as stem cell regulators (United States)

    Murphy, William L.; McDevitt, Todd C.; Engler, Adam J.


    The stem cell/material interface is a complex, dynamic microenvironment in which the cell and the material cooperatively dictate one another's fate: the cell by remodelling its surroundings, and the material through its inherent properties (such as adhesivity, stiffness, nanostructure or degradability). Stem cells in contact with materials are able to sense their properties, integrate cues via signal propagation and ultimately translate parallel signalling information into cell fate decisions. However, discovering the mechanisms by which stem cells respond to inherent material characteristics is challenging because of the highly complex, multicomponent signalling milieu present in the stem cell environment. In this Review, we discuss recent evidence that shows that inherent material properties may be engineered to dictate stem cell fate decisions, and overview a subset of the operative signal transduction mechanisms that have begun to emerge. Further developments in stem cell engineering and mechanotransduction are poised to have substantial implications for stem cell biology and regenerative medicine. PMID:24845994

  19. Mechanisms and regulation of water transport across the peritoneal membrane


    Morelle, Johann


    Peritoneal dialysis represents the major home-based dialysis modality for patients with end-stage renal disease. The capacity of the peritoneal membrane to transport water in response to an osmotic gradient contributes to the efficiency of the technique. Previous studies have demonstrated that aquaporin water channels are expressed in the endothelium lining peritoneal microvessels and mediate ultrafiltration during peritoneal dialysis. The investigations presented in this work provide novel i...

  20. Cell biology symposium: Membrane trafficking and signal transduction (United States)

    In general, membrane trafficking is a broad group of processes where proteins and other large molecules are distributed throughout the cell as well as adjacent extracellular spaces. Whereas signal transduction is a process where signals are transmitted through a series of chemical or molecular event...

  1. Polymers application in proton exchange membranes for fuel cells (PEMFCs) (United States)

    Walkowiak-Kulikowska, Justyna; Wolska, Joanna; Koroniak, Henryk


    This review presents the most important research on alternative polymer membranes with ionic groups attached, provides examples of materials with a well-defined chemical structure that are described in the literature. Furthermore, it elaborates on the synthetic methods used for preparing PEMs, the current status of fuel cell technology and its application. It also briefly discusses the development of the PEMFC market.

  2. Application of Proton Exchange Membrane Fuel Cell for Lift Trucks

    DEFF Research Database (Denmark)

    Hosseinzadeh, Elham; Rokni, Masoud


    in order to account for water back diffusion. Further Membrane water content is assumed to be a linear function of thickness. PEM fuel cell is working at rather low operating conditions which makes it suitable for the automotive systems. In this paper motive power part of a lift truck has been investigated...

  3. Perforate on CHO cell membranes induced by electromagnetic ...

    African Journals Online (AJOL)



    Jun 17, 2009 ... A number of studies have suggested that cell membranes may be a target of EMP irradiation (Lai et al.,. 1998). Over the past 2 decades, advances in specify instru- mentation have greatly contributed to cellular and mole- cular biology tools available to investigate nanoscaled bio- molecular structures.

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

    NARCIS (Netherlands)

    Coll-Almela, L.; Saura-Lopez, D.; Laencina-Sanchez, J.; Schols, H.A.; Voragen, A.G.J.; Ros-García, J.M.


    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

  5. Salinity induced changes in cell membrane stability, protein and ...

    African Journals Online (AJOL)

    control), 4.7, 9.4 and 14.1 dS m-1 to determine the effect of salt on vegetative growth, relative water content, cell membrane stability, protein and RNA contents in sand culture experiment. Fresh and dry weights of plants, shoots and roots decreased ...

  6. hydrogel membrane as electrolyte for direct borohydride fuel cells

    Indian Academy of Sciences (India)


    Abstract. A direct borohydride fuel cell (DBFC) employing a poly (vinyl alcohol) hydrogel membrane electrolyte (PHME) is reported. The DBFC employs an AB5 Misch metal alloy as anode and a gold- plated stainless steel mesh as cathode in conjunction with aqueous alkaline solution of sodium boro- hydride as fuel and ...

  7. hydrogel membrane as electrolyte for direct borohydride fuel cells

    Indian Academy of Sciences (India)

    A direct borohydride fuel cell (DBFC) employing a poly (vinyl alcohol) hydrogel membrane electrolyte (PHME) is reported. The DBFC employs an AB5 Misch metal alloy as anode and a goldplated stainless steel mesh as cathode in conjunction with aqueous alkaline solution of sodium borohydride as fuel and aqueous ...

  8. Denaturation of membrane proteins and hyperthermic cell killing

    NARCIS (Netherlands)

    Burgman, Paulus Wilhelmus Johannes Jozef


    Summarizing: heat induced denaturation of membrane proteins is probably related to hyperthermic cell killing. Induced resistance of heat sensitive proteins seems to be involved in the development of thermotolerance. Although many questions remain still to be answered, it appears that HSP72, when

  9. The roles of membrane microdomains (rafts) in T cell activation

    Czech Academy of Sciences Publication Activity Database

    Hořejší, Václav


    Roč. 191, - (2003), s. 148-164 ISSN 0105-2896 R&D Projects: GA MŠk LN00A026 Grant - others:Wellcome Trust(GB) J1116W24Z Institutional research plan: CEZ:AV0Z5052915 Keywords : membrane microdomain * raft * T cell Subject RIV: EC - Immunology Impact factor: 7.052, year: 2003

  10. Sulfonated carbon black-based composite membranes for fuel cell ...

    Indian Academy of Sciences (India)

    Sci., Vol. 36, No. 4, August 2013, pp. 563–573. c Indian Academy of Sciences. Sulfonated carbon black-based composite membranes for fuel cell applications .... All data were collected from a second heating cycle and glass tran- sition temperatures (Tg) were calculated as a midpoint of thermogram. 2.5d FTIR studies: FTIR ...

  11. The F-BAR domains from srGAP1, srGAP2 and srGAP3 regulate membrane deformation differently (United States)

    Coutinho-Budd, Jaeda; Ghukasyan, Vladimir; Zylka, Mark J.; Polleux, Franck


    Summary Coordination of membrane deformation and cytoskeletal dynamics lies at the heart of many biological processes critical for cell polarity, motility and morphogenesis. We have recently shown that Slit-Robo GTPase-activating protein 2 (srGAP2) regulates neuronal morphogenesis through the ability of its F-BAR domain to regulate membrane deformation and induce filopodia formation. Here, we demonstrate that the F-BAR domains of two closely related family members, srGAP1 and srGAP3 [designated F-BAR(1) and F-BAR(3), respectively] display significantly different membrane deformation properties in non-neuronal COS7 cells and in cortical neurons. F-BAR(3) induces filopodia in both cell types, though less potently than F-BAR(2), whereas F-BAR(1) prevents filopodia formation in cortical neurons and reduces plasma membrane dynamics. These three F-BAR domains can heterodimerize, and they act synergistically towards filopodia induction in COS7 cells. As measured by fluorescence recovery after photobleaching, F-BAR(2) displays faster molecular dynamics than F-BAR(3) and F-BAR(1) at the plasma membrane, which correlates well with its increased potency to induce filopodia. We also show that the molecular dynamic properties of F-BAR(2) at the membrane are partially dependent on F-Actin. Interestingly, acute phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] depletion in cells does not interfere with plasma membrane localization of F-BAR(2), which is compatible with our result showing that F-BAR(2) binds to a broad range of negatively-charged phospholipids present at the plasma membrane, including phosphatidylserine (PtdSer). Overall, our results provide novel insights into the functional diversity of the membrane deformation properties of this subclass of F-BAR-domains required for cell morphogenesis. PMID:22467852

  12. An adhesion-based method for plasma membrane isolation: evaluating cholesterol extraction from cells and their membranes. (United States)

    Bezrukov, Ludmila; Blank, Paul S; Polozov, Ivan V; Zimmerberg, Joshua


    A method to isolate large quantities of directly accessible plasma membrane from attached cells is presented. The method is based on the adhesion of cells to an adsorbed layer of polylysine on glass plates, followed by hypotonic lysis with ice-cold distilled water and subsequent washing steps. Optimal conditions for coating glass plates and time for cell attachment were established. No additional chemical or mechanical treatments were used. Contamination of the isolated plasma membrane by cell organelles was less than 5%. The method uses inexpensive, commercially available polylysine and reusable glass plates. Plasma membrane preparations can be made in 15 min. Using this method, we determined that methyl-beta-cyclodextrin differentially extracts cholesterol from fibroblast cells and their plasma membranes and that these differences are temperature dependent. Determination of the cholesterol/phospholipid ratio from intact cells does not reflect methyl-beta-cyclodextrin plasma membrane extraction properties.

  13. NCAM180 regulates Ric8A membrane localization and potentiates β-adrenergic response.

    Directory of Open Access Journals (Sweden)

    Marie-Claude Amoureux

    Full Text Available Cooperation between receptors allows integrated intracellular signaling leading to appropriate physiological responses. The Neural Cell Adhesion Molecule (NCAM has three main isoforms of 120, 140 and 180 kDa, with adhesive and signaling properties, but their respective functions remains to be fully identified. Here we show that the human NCAM180 intracellular domain is a novel interactor of the human guanosine exchange factor (GEF Ric8A using the yeast two hybrid system and immunoprecipitation. Furthermore, NCAM, Ric8A and G(αs form a tripartite complex. Colocalization experiments by confocal microscopy revealed that human NCAM180 specifically induces the recruitment of Ric8A to the membrane. In addition, using an in vitro recombinant system, and in vivo by comparing NCAM knock-out mouse brain to NCAM heterozygous and wild type brains, we show that NCAM expression dose dependently regulates Ric8A redistribution in detergent resistent membrane microdomains (DRM. Previous studies have demonstrated essential roles for Ric8 in G(α protein activity at G protein coupled receptors (GPCR, during neurotransmitter release and for asymmetric cell division. We observed that inhibition of Ric8A by siRNA or its overexpression, decreases or increases respectively, cAMP production following β-adrenergic receptor stimulation. Furthermore, in human HEK293T recombinant cells, NCAM180 potentiates the G(αs coupled β-adrenergic receptor response, in a Ric8A dependent manner, whereas NCAM120 or NCAM140 do not. Finally, in mouse hippocampal neurons expressing endogenously NCAM, NCAM is required for the agonist isoproterenol to induce cAMP production, and this requirement depends on Ric8A. These data illustrate a functional crosstalk between a GPCR and an IgCAM in the nervous system.

  14. Improving dynamic performance of proton-exchange membrane fuel cell system using time delay control (United States)

    Kim, Young-Bae

    Transient behaviour is a key parameter for the vehicular application of proton-exchange membrane (PEM) fuel cell. The goal of this presentation is to construct better control technology to increase the dynamic performance of a PEM fuel cell. The PEM fuel cell model comprises a compressor, an injection pump, a humidifier, a cooler, inlet and outlet manifolds, and a membrane-electrode assembly. The model includes the dynamic states of current, voltage, relative humidity, stoichiometry of air and hydrogen, cathode and anode pressures, cathode and anode mass flow rates, and power. Anode recirculation is also included with the injection pump, as well as anode purging, for preventing anode flooding. A steady-state, isothermal analytical fuel cell model is constructed to analyze the mass transfer and water transportation in the membrane. In order to prevent the starvation of air and flooding in a PEM fuel cell, time delay control is suggested to regulate the optimum stoichiometry of oxygen and hydrogen, even when there are dynamical fluctuations of the required PEM fuel cell power. To prove the dynamical performance improvement of the present method, feed-forward control and Linear Quadratic Gaussian (LQG) control with a state estimator are compared. Matlab/Simulink simulation is performed to validate the proposed methodology to increase the dynamic performance of a PEM fuel cell system.

  15. Plasma membrane calcium channels in cancer: Alterations and consequences for cell proliferation and migration. (United States)

    Déliot, Nadine; Constantin, Bruno


    The study of calcium channels in molecular mechanisms of cancer transformation is still a novel area of research. Several studies, mostly conducted on cancer cell lines, however support the idea that a diversity of plasma membrane channels participates in the remodeling of Ca2+ homeostasis, which regulates various cancer hallmarks such as uncontrolled multiplication and increase in migration and invasion abilities. However few is still understood concerning the intracellular signaling cascades mobilized by calcium influx participating to cancer cell behavior. This review intends to gather some of these pathways dependent on plasma membrane calcium channels and described in prostate, breast and lung cancer cell lines. In these cancer cell types, the calcium channels involved in calcium signaling pathways promoting cancer behaviors are mostly non-voltage activated calcium channels and belong to the TRP superfamily (TRPC, TPRPV and TRPM families) and the Orai family. TRP and Orai channels are part of many signaling cascades involving the activation of transmembrane receptors by extracellular ligand from the tumor environment. TRPV can sense changes in the physical and chemical environment of cancer cells and TRPM7 are stretch activated and sensitive to cholesterol. Changes in activation and or expression of plasma-membrane calcium channels affect calcium-dependent signaling processes relevant to tumorigenesis. The studies cited in this review suggest that an increase in plasma membrane calcium channel expression and/or activity sustain an elevated calcium entry (constitutive or under the control of extracellular signals) promoting higher cell proliferation and migration in most cases. A variety of non-voltage-operated calcium channels display change expression and/or activity in a same cancer type and cooperate to the same process relevant to cancer cell behavior, or can be involved in a different sequence of events during the tumorigenesis. This article is part of a

  16. Innovative membrane development for fuel cells

    CSIR Research Space (South Africa)

    Vaivars, G


    Full Text Available will take time, and the first alternative commercial car will be hybrid. The critical issue is the power source for an electrical engine. The fuel cell (FC)-battery hybrid is a promising solution to replace the combustion engine. Liquid fuel (e.g. methanol...

  17. VMP1 related autophagy and apoptosis in colorectal cancer cells: VMP1 regulates cell death

    Energy Technology Data Exchange (ETDEWEB)

    Qian, Qinyi [Department of Ultrasonograph, Changshu No. 2 People’s Hospital, Changshu (China); Zhou, Hao; Chen, Yan [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China); Shen, Chenglong [Department of General Surgery, Changshu No. 2 People’s Hospital, Changshu (China); He, Songbing; Zhao, Hua; Wang, Liang [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China); Wan, Daiwei, E-mail: [Department of Hepatobiliary Surgery, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou (China); Gu, Wen, E-mail: [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China)


    Highlights: •This research confirmed VMP1 as a regulator of autophagy in colorectal cancer cell lines. •We proved the pro-survival role of VMP1-mediated autophagy in colorectal cancer cell lines. •We found the interaction between VMP1 and BECLIN1 also existing in colorectal cancer cell lines. -- Abstract: Vacuole membrane protein 1 (VMP1) is an autophagy-related protein and identified as a key regulator of autophagy in recent years. In pancreatic cell lines, VMP1-dependent autophagy has been linked to positive regulation of apoptosis. However, there are no published reports on the role of VMP1 in autophagy and apoptosis in colorectal cancers. Therefore, to address this gap of knowledge, we decided to interrogate regulation of autophagy and apoptosis by VMP1. We have studied the induction of autophagy by starvation and rapamycin treatment in colorectal cell lines using electron microscopy, immunofluorescence, and immunoblotting. We found that starvation-induced autophagy correlated with an increase in VMP1 expression, that VMP1 interacted with BECLIN1, and that siRNA mediated down-regulation of VMP1-reduced autophagy. Next, we examined the relationship between VMP1-dependent autophagy and apoptosis and found that VMP1 down-regulation sensitizes cells to apoptosis and that agents that induce apoptosis down-regulate VMP1. In conclusion, similar to its reported role in other cell types, VMP1 is an important regulator of autophagy in colorectal cell lines. However, in contrast to its role in pancreatic cell lines, in colorectal cancer cells, VMP1-dependent autophagy appears to be pro-survival rather than pro-cell death.

  18. Chemical Imaging of the Cell Membrane by NanoSIMS

    Energy Technology Data Exchange (ETDEWEB)

    Weber, P K; Kraft, M L; Frisz, J F; Carpenter, K J; Hutcheon, I D


    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

  19. Durability aspects of polymer electrolyte membrane fuel cells (United States)

    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

  20. Identification of glycan structure alterations on cell membrane proteins in desoxyepothilone B resistant leukemia cells. (United States)

    Nakano, Miyako; Saldanha, Rohit; Göbel, Anja; Kavallaris, Maria; Packer, Nicolle H


    Resistance to tubulin-binding agents used in cancer is often multifactorial and can include changes in drug accumulation and modified expression of tubulin isotypes. Glycans on cell membrane proteins play important roles in many cellular processes such as recognition and apoptosis, and this study investigated whether changes to the glycan structures on cell membrane proteins occur when cells become resistant to drugs. Specifically, we investigated the alteration of glycan structures on the cell membrane proteins of human T-cell acute lymphoblastic leukemia (CEM) cells that were selected for resistance to desoxyepothilone B (CEM/dEpoB). The glycan profile of the cell membrane glycoproteins was obtained by sequential release of N- and O-glycans from cell membrane fraction dotted onto polyvinylidene difluoride membrane with PNGase F and β-elimination respectively. The released glycan alditols were analyzed by liquid chromatography (graphitized carbon)-electrospray ionization tandem MS. The major N-glycan on CEM cell was the core fucosylated α2-6 monosialo-biantennary structure. Resistant CEM/dEpoB cells had a significant decrease of α2-6 linked sialic acid on N-glycans. The lower α2-6 sialylation was caused by a decrease in activity of β-galactoside α2-6 sialyltransferase (ST6Gal), and decreased expression of the mRNA. It is clear that the membrane glycosylation of leukemia cells changes during acquired resistance to dEpoB drugs and that this change occurs globally on all cell membrane glycoproteins. This is the first identification of a specific glycan modification on the surface of drug resistant cells and the mechanism of this downstream effect on microtubule targeting drugs may offer a route to new interventions to overcome drug resistance.

  1. Anion exchange membrane based on alkali doped poly(2,5-benzimidazole) for alkaline membrane fuel cell

    CSIR Research Space (South Africa)

    Luo, H


    Full Text Available Alkaline membrane fuel cell (AMFC) has been received increasing attention among the different types of fuel cells. Ammonium quaternized polymers such as poly (arylene ether sulfones) are being developed and studied as candidates of ionomeric...

  2. Synthesis and characterization of Nafion/TiO2 nanocomposite membrane for proton exchange membrane fuel cell. (United States)

    Kim, Tae Young; Cho, Sung Yong


    In this study, the syntheses and characterizations of Nafion/TiO2 membranes for a proton exchange membrane fuel cell (PEMFC) were investigated. Porous TiO2 powders were synthesized using the sol-gel method; with Nafion/TiO2 nanocomposite membranes prepared using the casting method. An X-ray diffraction analysis demonstrated that the synthesized TiO2 had an anatase structure. The specific surface areas of the TiO2 and Nafion/TiO2 nanocomposite membrane were found to be 115.97 and 33.91 m2/g using a nitrogen adsorption analyzer. The energy dispersive spectra analysis indicated that the TiO2 particles were uniformly distributed in the nanocomposite membrane. The membrane electrode assembly prepared from the Nafion/TiO2 nanocomposite membrane gave the best PEMFC performance compared to the Nafion/P-25 and Nafion membranes.

  3. Polybenzimidazole Membranes Containing Benzimidazole Side Groups for High Temprature Polymer Electrolyte Membrane Fuel Cells

    DEFF Research Database (Denmark)

    Yang, Jingshuai; Li, Xueyuan; Xu, Yizin


    temperatures without humidification. At an acid doping level of 13.1 mol H3PO4 per average molar repeat unit, the PBI membranes with a benzimidazole grafting degree of 10.6% demonstrated a conductivity of 0.15 S cm-1 and a H2-air fuel cell peak power density of 378 mW cm-2 at 180 oC at ambient pressure without...

  4. A Novel Unitized Regenerative Proton Exchange Membrane Fuel Cell (United States)

    Murphy, O. J.; Cisar, A. J.; Gonzalez-Martin, A.; Salinas, C. E.; Simpson, S. F.


    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.

  5. The Chemical Potential of Plasma Membrane Cholesterol: Implications for Cell Biology. (United States)

    Ayuyan, Artem G; Cohen, Fredric S


    Cholesterol is abundant in plasma membranes and exhibits a variety of interactions throughout the membrane. Chemical potential accounts for thermodynamic consequences of molecular interactions, and quantifies the effective concentration (i.e., activity) of any substance participating in a process. We have developed, to our knowledge, the first method to measure cholesterol chemical potential in plasma membranes. This was accomplished by complexing methyl-β-cyclodextrin with cholesterol in an aqueous solution and equilibrating it with an organic solvent containing dissolved cholesterol. The chemical potential of cholesterol was thereby equalized in the two phases. Because cholesterol is dilute in the organic phase, here activity and concentration were equivalent. This equivalence allowed the amount of cholesterol bound to methyl-β-cyclodextrin to be converted to cholesterol chemical potential. Our method was used to determine the chemical potential of cholesterol in erythrocytes and in plasma membranes of nucleated cells in culture. For erythrocytes, the chemical potential did not vary when the concentration was below a critical value. Above this value, the chemical potential progressively increased with concentration. We used standard cancer lines to characterize cholesterol chemical potential in plasma membranes of nucleated cells. This chemical potential was significantly greater for highly metastatic breast cancer cells than for nonmetastatic breast cancer cells. Chemical potential depended on density of the cancer cells. A method to alter and fix the cholesterol chemical potential to any value (i.e., a cholesterol chemical potential clamp) was also developed. Cholesterol content did not change when cells were clamped for 24-48 h. It was found that the level of activation of the transcription factor STAT3 increased with increasing cholesterol chemical potential. The cholesterol chemical potential may regulate signaling pathways. Copyright © 2018. Published by

  6. Analysis of membrane proteome and secretome in cells over-expressing ADAM17 using quantitative proteomics

    Energy Technology Data Exchange (ETDEWEB)

    Kawahara, R.; Simabuco, F.M. [Laboratorio Nacional de Biociencias - LNBIO, Campinas, SP (Brazil); Yokoo, S.; Paes Leme, A.F. [Laboratorio Nacional de Luz Sincrotron (LNLS), Campinas, SP (Brazil); Sherman, N. [University of Virginia, Charlottesville, VA (United States)


    Full text: A disintegrin and metalloproteinase (ADAM) protease is involved in proteolytic ectodomain shedding of several membrane-associated proteins and modulation of key cell signaling pathways in the tumor microenvironment. In this study, we examined the effect of over-expressing the full length human ADAM17 in membrane and secreted proteins. To this end, we constructed a stable Flp-In T-RExHEK293 cells expressing ADAM17 by tetracycline induction. These cells were grown in Dulbeccos modified Eagles medium containing light lysine, arginine or heavy, L-Arg-13C615N4 and L-Lys -13C615N2 (SILAC: stable isotope labeling with amino acid in cell culture) media and they were treated with an ADAM17 activator, phorbolester (PMA). Controls such as Flp-In T-RExHEK293 cell without PMA treatment and without ADAM17 cloned were cultivated in light medium. The ADAM17 overexpression was induced with tetracycline 500 ng/ml for 24 hours. Cells in a heavy condition were treated with PMA 50 ng/ml for 1 hour and vehicle DMSO was used as control in a light cell condition. The extracellular media were collected, concentrated and used to evaluate the secretome and a cell surface biotinylation-based approach was used to capture cell surface-associated proteins. The biotinylated proteins were eluted with dithiothreitol, alkylated with iodoacetamide and then digested with trypsin. The resulting peptides were subjected to LC-MS/MS analysis on an ETD enabled Orbitrap Velos instrument. The results showed different proteins up or down regulated in membrane and secretome analysis which might represent potential molecules involved in signaling or ADAM17 regulation events. (author)


    Directory of Open Access Journals (Sweden)

    Ya. A. Udalova


    Full Text Available Affine chromatography was used to isolate Lec antibodies from the sera of a healthy female donor with the high titers of these anti- bodies, which were labeled with biotin. The study enrolled 51 patients with primary breast cancer (BC. Antigen expression was found by immunohistochemistry and flow cytometry. With these two techniques being used, the detection rate of Lec expression in BC cells was 65% (33/51; the antigen was most frequently found by flow cytometry as compared with immunohistochemistry: 72 and 58% of cases, respectively.

  8. Mass Spectrometry of Polymer Electrolyte Membrane Fuel Cells

    Directory of Open Access Journals (Sweden)

    Viktor Johánek


    Full Text Available 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.

  9. Mass Spectrometry of Polymer Electrolyte Membrane Fuel Cells. (United States)

    Johánek, Viktor; Ostroverkh, Anna; Fiala, Roman; Rednyk, Andrii; Matolín, Vladimír


    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.

  10. Isolation of a Wheat Cell Line with Altered Membrane Properties (United States)

    Erdei, László; Vigh, László; Dudits, Dénes


    A spontaneous dimethylsulfoxide (DMSO)-tolerant cell line was isolated from a cell culture of wheat (Triticum monococcum L.). The tolerant cells were able to grow in the presence of 4% DMSO. Cells formed from protoplasts of the tolerant line required DMSO for division in culture medium of high osmotic value. Fatty acid composition and the molar ratio of phospholipids/sterols suggest a more ordered membrane structure in the tolerant line. Accordingly, a lower K+ influx rate was detected in the tolerant cells in comparison with the original line. These characteristics were maintained after 6 months' cultivation of the cells in DMSO-free growth medium. This suggested that genetic changes could be responsible for differences between the two cell lines. PMID:16662251

  11. Cell membrane wrapping of a spherical thin elastic shell. (United States)

    Yi, Xin; Gao, Huajian


    Nanocapsules that can be tailored intelligently and specifically have drawn considerable attention in the fields of drug delivery and bioimaging. Here we conduct a theoretical study on cell uptake of a spherical nanocapsule which is modeled as a linear elastic solid thin shell in three dimensions. It is found that there exist five wrapping phases based on the stability of three wrapping states: no wrapping, partial wrapping and full wrapping. The wrapping phase diagrams are strongly dependent on the capsule size, adhesion energy, cell membrane tension, and bending rigidity ratio between the capsule and membrane. Discussion is made on similarities and differences between the cell uptake of solid nanocapsules and fluid vesicles. The reported results may have important implications for biomedical applications of nanotechnology.

  12. Phytochemicals prevent mitochondrial membrane permeabilization and protect SH-SY5Y cells against apoptosis induced by PK11195, a ligand for outer membrane translocator protein. (United States)

    Wu, Yuqiu; Shamoto-Nagai, Masayo; Maruyama, Wakako; Osawa, Toshihiko; Naoi, Makoto


    Epidemiological studies present the beneficial effects of dietary habits on prevention of aging-associated decline of brain function. Phytochemicals, the second metabolites of food, protect neuronal cells from cell death in cellular models of neurodegenerative disorders, and the neuroprotective activity has been ascribed to the anti-oxidant and anti-inflammatory functions. In this paper, the cellular mechanism of neuroprotection by phytochemicals was investigated, using the cellular model of mitochondrial apoptosis induced by PK11195, a ligand of outer membrane translocator protein, in SH-SY5Y cells. PK11195 induced mitochondrial membrane permeabilization with rapid transit production of superoxide (superoxide flashes) and calcium release from mitochondria, and activated apoptosis signal pathway. Study on the structure-activity relationship of astaxanthin, ferulic acid derivatives, and sesame lignans revealed that these phytochemicals inhibited mitochondrial membrane permeabilization and protected cells from apoptosis. Ferulic acid derivatives and sesame lignans inhibited or enhanced the mitochondrial pore formation and cell death by PK11195 according to their amphiphilic properties, not directly depending on the antioxidant activity. Regulation of pore formation at mitochondrial membrane is discussed as a novel mechanism behind neuroprotective activity of phytochemicals in aging and age-associated neurodegenerative disorders, and also behind dual functions of phytochemicals in neuronal and cancer cells.

  13. Mechanical stress downregulates MHC class I expression on human cancer cell membrane.

    Directory of Open Access Journals (Sweden)

    Rosanna La Rocca

    Full Text Available In our body, cells are continuously exposed to physical forces that can regulate different cell functions such as cell proliferation, differentiation and death. In this work, we employed two different strategies to mechanically stress cancer cells. The cancer and healthy cell populations were treated either with mechanical stress delivered by a micropump (fabricated by deep X-ray nanolithography or by ultrasound wave stimuli. A specific down-regulation of Major Histocompatibility Complex (MHC class I molecules expression on cancer cell membrane compared to different kinds of healthy cells (fibroblasts, macrophages, dendritic and lymphocyte cells was observed, stimulating the cells with forces in the range of nano-newton, and pressures between 1 and 10 bar (1 bar = 100.000 Pascal, depending on the devices used. Moreover, Raman spectroscopy analysis, after mechanical treatment, in the range between 700-1800 cm(-1, indicated a relative concentration variation of MHC class I. PCA analysis was also performed to distinguish control and stressed cells within different cell lines. These mechanical induced phenotypic changes increase the tumor immunogenicity, as revealed by the related increased susceptibility to Natural Killer (NK cells cytotoxic recognition.

  14. Mechanical Stress Downregulates MHC Class I Expression on Human Cancer Cell Membrane

    KAUST Repository

    La Rocca, Rosanna


    In our body, cells are continuously exposed to physical forces that can regulate different cell functions such as cell proliferation, differentiation and death. In this work, we employed two different strategies to mechanically stress cancer cells. The cancer and healthy cell populations were treated either with mechanical stress delivered by a micropump (fabricated by deep X-ray nanolithography) or by ultrasound wave stimuli. A specific down-regulation of Major Histocompatibility Complex (MHC) class I molecules expression on cancer cell membrane compared to different kinds of healthy cells (fibroblasts, macrophages, dendritic and lymphocyte cells) was observed, stimulating the cells with forces in the range of nano-newton, and pressures between 1 and 10 bar (1 bar = 100.000 Pascal), depending on the devices used. Moreover, Raman spectroscopy analysis, after mechanical treatment, in the range between 700–1800 cm−1, indicated a relative concentration variation of MHC class I. PCA analysis was also performed to distinguish control and stressed cells within different cell lines. These mechanical induced phenotypic changes increase the tumor immunogenicity, as revealed by the related increased susceptibility to Natural Killer (NK) cells cytotoxic recognition.

  15. Nafion/silane nanocomposite membranes for high temperature polymer electrolyte membrane fuel cell. (United States)

    Ghi, Lee Jin; Park, Na Ri; Kim, Moon Sung; Rhee, Hee Woo


    The polymer electrolyte membrane fuel cell (PEMFC) has been studied actively for both potable and stationary applications because it can offer high power density and be used only hydrogen and oxygen as environment-friendly fuels. Nafion which is widely used has mechanical and chemical stabilities as well as high conductivity. However, there is a drawback that it can be useless at high temperatures (> or = 90 degrees C) because proton conducting mechanism cannot work above 100 degrees C due to dehydration of membrane. Therefore, PEMFC should be operated for long-term at high temperatures continuously. In this study, we developed nanocomposite membrane using stable properties of Nafion and phosphonic acid groups which made proton conducting mechanism without water. 3-Aminopropyl triethoxysilane (APTES) was used to replace sulfonic acid groups of Nafion and then its aminopropyl group was chemically modified to phosphonic acid groups. The nanocomposite membrane showed very high conductivity (approximately 0.02 S/cm at 110 degrees C, <30% RH).

  16. Phosphoric acid distribution in the membrane electrode assembly of high temperature proton exchange membrane fuel cells

    International Nuclear Information System (INIS)

    Kwon, Kyungjung; Park, Jung Ock; Yoo, Duck Young; Yi, Jung S.


    The ionomer content in electrode is one of the most important parameters for the high performance of fuel cells. The high temperature PEMFC based on phosphoric acid (PA)-doped polymer membrane with unhumidified reactant gases has a difficulty in controlling the liquid state PA ionomer content in electrode. To evaluate the PA content in electrode, the three techniques of cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and acid-base titration (ABT) are carried out in situ or ex situ. The properties of membrane electrode assembly (MEA) such as electrochemical surface area (ESA), ohmic resistance, charge transfer resistance, double layer capacitance and the amount of PA in MEA components (anode, cathode and membrane) are extracted by each technique. Ex situ CV with the usage of dry gases has a limitation in assessing the reliable ESA of unhumidified PEMFC. While in situ EIS presents some informative values of resistance and capacitance for understanding the PA distribution in MEA, its sensitivity to the PA content in MEA components needs to be higher for detecting a subtle change in PA distribution. Ex situ ABT supplies a clear PA distribution in MEA at room temperature but does not seem to reflect the operating state well at high temperatures. However, it can be used as a detection tool for the loss of the initial acid content in membrane during a long-term MEA durability study.

  17. Model of fission yeast cell shape driven by membrane-bound growth factors and the cytoskeleton.

    Directory of Open Access Journals (Sweden)

    Tyler Drake

    Full Text Available Fission yeast serves as a model for how cellular polarization machinery consisting of signaling molecules and the actin and microtubule cytoskeleton regulates cell shape. In this work, we develop mathematical models to investigate how these cells maintain a tubular shape of approximately constant diameter. Many studies identify active Cdc42, found in a cap at the inner membrane of growing cell tips, as an important regulator of local cell wall remodeling, likely through control of exocyst tethering and the targeting of other polarity-enhancing structures. First, we show that a computational model with Cdc42-dependent local cell wall remodeling under turgor pressure predicts a relationship between spatial extent of growth signal and cell diameter that is in agreement with prior experiments. Second, we model the consequences of feedback between cell shape and distribution of Cdc42 growth signal at cell tips. We show that stability of cell diameter over successive cell divisions places restrictions on their mutual dependence. We argue that simple models where the spatial extent of the tip growth signal relies solely on geometrical alignment of confined microtubules might lead to unstable width regulation. Third, we study a computational model that combines a growth signal distributed over a characteristic length scale (as, for example, by a reaction-diffusion mechanism with an axis-sensing microtubules system that places landmarks at positions where microtubule tips touch the cortex. A two-dimensional implementation of this model leads to stable cell diameter for a wide range of parameters. Changes to the parameters of this model reproduce straight, bent, and bulged cell shapes, and we discuss how this model is consistent with other observed cell shapes in mutants. Our work provides an initial quantitative framework for understanding the regulation of cell shape in fission yeast, and a scaffold for understanding this process on a more molecular

  18. Interaction of membrane/lipid rafts with the cytoskeleton: impact on signaling and function: membrane/lipid rafts, mediators of cytoskeletal arrangement and cell signaling. (United States)

    Head, Brian P; Patel, Hemal H; Insel, Paul A


    The plasma membrane in eukaryotic cells contains microdomains that are enriched in certain glycosphingolipids, gangliosides, and sterols (such as cholesterol) to form membrane/lipid rafts (MLR). These regions exist as caveolae, morphologically observable flask-like invaginations, or as a less easily detectable planar form. MLR are scaffolds for many molecular entities, including signaling receptors and ion channels that communicate extracellular stimuli to the intracellular milieu. Much evidence indicates that this organization and/or the clustering of MLR into more active signaling platforms depends upon interactions with and dynamic rearrangement of the cytoskeleton. Several cytoskeletal components and binding partners, as well as enzymes that regulate the cytoskeleton, localize to MLR and help regulate lateral diffusion of membrane proteins and lipids in response to extracellular events (e.g., receptor activation, shear stress, electrical conductance, and nutrient demand). MLR regulate cellular polarity, adherence to the extracellular matrix, signaling events (including ones that affect growth and migration), and are sites of cellular entry of certain pathogens, toxins and nanoparticles. The dynamic interaction between MLR and the underlying cytoskeleton thus regulates many facets of the function of eukaryotic cells and their adaptation to changing environments. Here, we review general features of MLR and caveolae and their role in several aspects of cellular function, including polarity of endothelial and epithelial cells, cell migration, mechanotransduction, lymphocyte activation, neuronal growth and signaling, and a variety of disease settings. This article is part of a Special Issue entitled: Reciprocal influences between cell cytoskeleton and membrane channels, receptors and transporters. Guest Editor: Jean Claude Hervé. Published by Elsevier B.V.

  19. β(IV)-Spectrin regulates TREK-1 membrane targeting in the heart. (United States)

    Hund, Thomas J; Snyder, Jedidiah S; Wu, Xiangqiong; Glynn, Patric; Koval, Olha M; Onal, Birce; Leymaster, Nicholas D; Unudurthi, Sathya D; Curran, Jerry; Camardo, Celia; Wright, Patrick J; Binkley, Philip F; Anderson, Mark E; Mohler, Peter J


    Cardiac function depends on the highly regulated and co-ordinate activity of a large ensemble of potassium channels that control myocyte repolarization. While voltage-gated K(+) channels have been well characterized in the heart, much less is known about regulation and/or targeting of two-pore K(+) channel (K(2P)) family members, despite their potential importance in modulation of heart function. Here, we report a novel molecular pathway for membrane targeting of TREK-1, a mechano-sensitive K(2P) channel regulated by environmental and physical factors including membrane stretch, pH, and polyunsaturated fatty acids (e.g. arachidonic acid). We demonstrate that β(IV)-spectrin, an actin-associated protein, is co-localized with TREK-1 at the myocyte intercalated disc, associates with TREK-1 in the heart, and is required for TREK-1 membrane targeting. Mice expressing β(IV)-spectrin lacking TREK-1 binding (qv(4J)) display aberrant TREK-1 membrane localization, decreased TREK-1 activity, delayed action potential repolarization, and arrhythmia without apparent defects in localization/function of other cardiac potassium channel subunits. Finally, we report abnormal β(IV)-spectrin levels in human heart failure. These data provide new insight into membrane targeting of TREK-1 in the heart and establish a broader role for β(IV)-spectrin in organizing functional membrane domains critical for normal heart function.

  20. βIV-Spectrin regulates TREK-1 membrane targeting in the heart (United States)

    Hund, Thomas J.; Snyder, Jedidiah S.; Wu, Xiangqiong; Glynn, Patric; Koval, Olha M.; Onal, Birce; Leymaster, Nicholas D.; Unudurthi, Sathya D.; Curran, Jerry; Camardo, Celia; Wright, Patrick J.; Binkley, Philip F.; Anderson, Mark E.; Mohler, Peter J.


    Aims Cardiac function depends on the highly regulated and co-ordinate activity of a large ensemble of potassium channels that control myocyte repolarization. While voltage-gated K+ channels have been well characterized in the heart, much less is known about regulation and/or targeting of two-pore K+ channel (K2P) family members, despite their potential importance in modulation of heart function. Methods and results Here, we report a novel molecular pathway for membrane targeting of TREK-1, a mechano-sensitive K2P channel regulated by environmental and physical factors including membrane stretch, pH, and polyunsaturated fatty acids (e.g. arachidonic acid). We demonstrate that βIV-spectrin, an actin-associated protein, is co-localized with TREK-1 at the myocyte intercalated disc, associates with TREK-1 in the heart, and is required for TREK-1 membrane targeting. Mice expressing βIV-spectrin lacking TREK-1 binding (qv4J) display aberrant TREK-1 membrane localization, decreased TREK-1 activity, delayed action potential repolarization, and arrhythmia without apparent defects in localization/function of other cardiac potassium channel subunits. Finally, we report abnormal βIV-spectrin levels in human heart failure. Conclusions These data provide new insight into membrane targeting of TREK-1 in the heart and establish a broader role for βIV-spectrin in organizing functional membrane domains critical for normal heart function. PMID:24445605

  1. Alteration of membrane complement regulators is associated with transporter status in patients on peritoneal dialysis (United States)

    Biegger, Dagmar; Segerer, Stephan; Braun, Niko; Alscher, M. Dominik; Latus, Joerg


    Introduction A growing body of evidence from animal models and cell culture studies indicate an important role of a local regulatory complement system (CS) in peritoneal injury during peritoneal dialysis (PD). We investigated the expression of the local regulatory CS (reflected by CD46,CD55,CD59) in the peritoneal tissue of patients with different membrane function characteristics. Patients and methods Biopsies from the parietal peritoneum were taken from 24 patients on PD, 22 uremic patients prior to PD. PD patients were grouped according to the dialysate-to-plasma ratio of creatinine (D/P Cre) and ratio of dialysate glucose at 4 hours versus dialysate glucose at time zero (D/D0 glucose) into low or low-average peritoneal transport status (L/LA) and high-average or high-transport status (HA/H) groups. CD46, CD55, and CD59 RNA expression were analyzed by real-time polymerase chain reaction (RT-PCR). Further localization of membrane complement regulators (CRegs) and semiquantitatively analysis was done by immunohistochemistry (IHC). Results CD46 and CD59 expression were similar in all groups. CD55 expression was significantly decreased in the HA/H group compared to the L/LA group and to uremic controls (p peritonitis. There was no statistically significant correlation between PD duration and the expressions of CD46, CD55, and CD59. IHC revealed strong CD46, CD55, and CD59 expression in mesothelial cells. CD55 and CD59 were additionally detected in the vasculature. Using IHC, CD46 was lower in PD patients compared to uremic controls (p>0.05), but there was no difference between the L/LA compared to the H/HA group. Moreover IHC confirmed decreased expression of CD55 in the HA/H group compared to the L/LA group and uremic controls (pperitonitis and PD duration do not appear to alter CReg expression. PMID:28542228

  2. Interplay between membrane elasticity and active cytoskeleton forces regulates the aggregation dynamics of the immunological synapse (United States)

    Dharan, Nadiv; Farago, Oded

    Adhesion between a T cell and an antigen presenting cell is achieved by TCR-pMHC and LFA1-ICAM1 protein complexes. These segregate to form a special pattern, known as the immunological synapse (IS), consisting of a central quasi-circular domain of TCR-pMHC bonds surrounded by a peripheral domain of LFA1-ICAM1 complexes. Insights gained from imaging studies had led to the conclusion that the formation of the central adhesion domain in the IS is driven by active (ATP-driven) mechanisms. Recent studies, however, suggested that passive (thermodynamic) mechanisms may also play an important role in this process. Here, we present a simple physical model, taking into account the membrane-mediated thermodynamic attraction between the TCR-pMHC bonds and the effective forces that they experience due to ATP-driven actin retrograde flow and transport by dynein motor proteins. Monte Carlo simulations of the model exhibit a good spatio-temporal agreement with the experimentally observed pattern evolution of the TCR-pMHC microclusters. The agreement is lost when one of the aggregation mechanisms is "muted", which helps to identify the respective roles in the process. We conclude that actin retrograde flow drives the centripetal motion of TCR-pMHC bonds, while the membrane-mediated interactions facilitate microcluster formation and growth. In the absence of dynein motors, the system evolves into a ring-shaped pattern, which highlights the role of dynein motors in the formation of the final concentric pattern. The interplay between the passive and active mechanisms regulates the rate of the accumulation process, which in the absence of one them proceeds either too quickly or slowly.

  3. Modeling and Simulation for Fuel Cell Polymer Electrolyte Membrane

    Directory of Open Access Journals (Sweden)

    Takahiro Hayashi


    Full Text Available We have established methods to evaluate key properties that are needed to commercialize polyelectrolyte membranes for fuel cell electric vehicles such as water diffusion, gas permeability, and mechanical strength. These methods are based on coarse-graining models. For calculating water diffusion and gas permeability through the membranes, the dissipative particle dynamics–Monte Carlo approach was applied, while mechanical strength of the hydrated membrane was simulated by coarse-grained molecular dynamics. As a result of our systematic search and analysis, we can now grasp the direction necessary to improve water diffusion, gas permeability, and mechanical strength. For water diffusion, a map that reveals the relationship between many kinds of molecular structures and diffusion constants was obtained, in which the direction to enhance the diffusivity by improving membrane structure can be clearly seen. In order to achieve high mechanical strength, the molecular structure should be such that the hydrated membrane contains narrow water channels, but these might decrease the proton conductivity. Therefore, an optimal design of the polymer structure is needed, and the developed models reviewed here make it possible to optimize these molecular structures.

  4. Membrane properties of smooth muscle cells in pulmonary hypertensive rats. (United States)

    Suzuki, H; Twarog, B M


    The membrane properties of smooth muscle cells in rat main pulmonary artery (MPA) and small pulmonary artery (SPA) were investigated during chronic normobaric hypoxia and after monocrotaline injection. As chronic pulmonary hypertension developed, pronounced differences between MPA and SPA were observed. These findings may shed light on mechanisms of smooth muscle hypertrophy. 1) The resting membrane potential of smooth muscle in MPA became less negative than the normal (depolarized), whereas the resting membrane potential of smooth muscle in SPA became more negative (hyperpolarized). 2) In MPA, both the length and time constants diminished. 3) In MPA, the maximum membrane depolarization produced by a 10-fold increase in extracellular [K+] decreased. 4) In SPA, the depolarization observed in K+-free solution was more rapid and greater in amplitude, and the transient hyperpolarization following restoration of K+-containing solution increased. 5) In SPA, initial and sustained depolarization evoked by Na+-deficient solutions were increased. 6) Depolarization in MPA was due to increased membrane permeability, perhaps to Cl-, whereas hyperpolarization in SPA could be attributed to increased activity of an electrogenic Na+-K+ pump.

  5. Modified SPEEK membranes for direct ethanol fuel cell

    KAUST Repository

    Maab, Husnul


    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 x 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 x 10(-10) kg m s(-1) m(-2) respectively, which is 2- to 50-fold lower than that for plain SPEEK (19 x 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 degrees C and 90 degrees C. (C) 2010 Elsevier B.V. All rights reserved.

  6. Bidirectional regulation between B cells and T cells

    NARCIS (Netherlands)

    Margry, B.


    B cells were often thought of as simple precursors of end-stage effector cells that are merely in charge of antibody production. Research in the last decades has shown that B cells possess important other roles as well, including their involvement in the regulation and functioning of T cell-mediated

  7. Membrane-Associated Transporter Protein (MATP Regulates Melanosomal pH and Influences Tyrosinase Activity.

    Directory of Open Access Journals (Sweden)

    Bum-Ho Bin

    Full Text Available The SLC45A2 gene encodes a Membrane-Associated Transporter Protein (MATP. Mutations of this gene cause oculocutaneous albinism type 4 (OCA4. However, the molecular mechanism of its action in melanogenesis has not been elucidated. Here, we discuss the role of MATP in melanin production. The SLC45A2 gene is highly enriched in human melanocytes and melanoma cell lines, and its protein, MATP, is located in melanosomes. The knockdown of MATP using siRNAs reduced melanin content and tyrosinase activity without any morphological change in melanosomes or the expression of melanogenesis-related proteins. Interestingly, the knockdown of MATP significantly lowered the melanosomal pH, as verified through DAMP analysis, suggesting that MATP regulates melanosomal pH and therefore affects tyrosinase activity. Finally, we found that the reduction of tyrosinase activity associated with the knockdown of MATP was readily recovered by copper treatment in the in vitro L-DOPA oxidase activity assay of tyrosinase. Considering that copper is an important element for tyrosinase activity and that its binding to tyrosinase depends on melanosomal pH, MATP may play an important role in regulating tyrosinase activity via controlling melanosomal pH.

  8. Cell swelling and volume regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay


    been reported. The role of the different channels was discussed. A taurine leak pathway is clearly activated after cell swelling both in astrocytes and in neurones. The relations between the effect of glutamate and cell swelling were discussed. Discussion on the clearance of potassium from...

  9. Immune regulation by mast cells

    NARCIS (Netherlands)

    Suurmond, Jolien


    The objective of this PhD thesis is to understand mast cell (and basophil) functions and their role in autoimmune disease by focusing on three main aims: 1. To characterize the interaction between innate and Fc receptor triggers on mast cell and basophil function 2. To analyze the interaction

  10. Ubiquitination of the bacterial inositol phosphatase, SopB, regulates its biological activity at the plasma membrane.

    LENUS (Irish Health Repository)

    Knodler, Leigh A


    The Salmonella type III effector, SopB, is an inositol polyphosphate phosphatase that modulates host cell phospholipids at the plasma membrane and the nascent Salmonella-containing vacuole (SCV). Translocated SopB persists for many hours after infection and is ubiquitinated but the significance of this covalent modification has not been investigated. Here we identify by mass spectrometry six lysine residues of SopB that are mono-ubiquitinated. Substitution of these six lysine residues with arginine, SopB-K(6)R, almost completely eliminated SopB ubiquitination. We found that ubiquitination does not affect SopB stability or membrane association, or SopB-dependent events in SCV biogenesis. However, two spatially and temporally distinct events are dependent on ubiquitination, downregulation of SopB activity at the plasma membrane and prolonged retention of SopB on the SCV. Activation of the mammalian pro-survival kinase Akt\\/PKB, a downstream target of SopB, was intensified and prolonged after infection with the SopB-K(6)R mutant. At later times, fewer SCV were decorated with SopB-K(6)R compared with SopB. Instead SopB-K(6)R was present as discrete vesicles spread diffusely throughout the cell. Altogether, our data show that ubiquitination of SopB is not related to its intracellular stability but rather regulates its enzymatic activity at the plasma membrane and intracellular localization.

  11. Crystalline polymorphism induced by charge regulation in ionic membranes. (United States)

    Leung, Cheuk-Yui; Palmer, Liam C; Kewalramani, Sumit; Qiao, Baofu; Stupp, Samuel I; Olvera de la Cruz, Monica; Bedzyk, Michael J


    The crystallization of molecules with polar and hydrophobic groups, such as ionic amphiphiles and proteins, is of paramount importance in biology and biotechnology. By coassembling dilysine (+2) and carboxylate (-1) amphiphiles of various tail lengths into bilayer membranes at different pH values, we show that the 2D crystallization process in amphiphile membranes can be controlled by modifying the competition of long-range and short-range interactions among the polar and the hydrophobic groups. The pH and the hydrophobic tail length modify the intermolecular packing and the symmetry of their crystalline phase. For hydrophobic tail lengths of 14 carbons (C14), we observe the coassembly into crystalline bilayers with hexagonal molecular ordering via in situ small- and wide-angle X-ray scattering. As the tail length increases, the hexagonal lattice spacing decreases due to an increase in van der Waals interactions, as demonstrated by atomistic molecular dynamics simulations. For C16 and C18 we observe a reentrant crystalline phase transition sequence, hexagonal-rectangular-C-rectangular-P-rectangular-C-hexagonal, as the solution pH is increased from 3 to 10.5. The stability of the rectangular phases, which maximize tail packing, increases with increasing tail length. As a result, for very long tails (C22), the possibility of observing packing symmetries other than rectangular-C phases diminishes. Our work demonstrates that it is possible to systematically exchange chemical and mechanical energy by changing the solution pH value within a range of physiological conditions at room temperature in bilayers of molecules with ionizable groups.

  12. Leptin Regulation of Mammary Cell Growth

    National Research Council Canada - National Science Library

    Pighetti, Gina


    .... The studies of this proposal were designed to test the hypothesis that the interaction of leptin with its receptor regulates normal and pathologic mammary epithelial cell proliferation and/or differentiation...

  13. Stimulating Effect of Terfenadine on Erythrocyte Cell Membrane Scrambling

    Directory of Open Access Journals (Sweden)

    Elena Signoretto


    Full Text Available Background/Aims: The antihistaminic drug Terfenadine may trigger apoptosis of tumor cells, an effect unrelated to its effect on histamine receptors. Similar to apoptosis of nucleated cells, erythrocytes may enter eryptosis, the suicidal death of erythrocytes characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Signaling triggering eryptosis include increase of cytosolic Ca2+ activity ([Ca2+]i, oxidative stress, and ceramide. The present study explored, whether Terfenadine is capable to trigger eryptosis. Methods: Flow cytometry was employed to estimate phosphatidylserine abundance at the erythrocyte surface from annexin-V-binding, cell volume from forward scatter, [Ca2+]i from Fluo3-fluorescence, abundance of reactive oxygen species (ROS from 2′,7′-dichlorodihydrofluorescein (DCF diacetate dependent fluorescence, and ceramide abundance at the human erythrocyte surface utilizing specific antibodies. Hemolysis was quantified from haemoglobin concentration in the supernatant. Results: A 48 hours exposure of human erythrocytes to Terfenadine (≥ 5 µM significantly increased the percentage of annexin-V-binding cells and triggered hemolysis without significantly modifying the average forward scatter. Terfenadine (7.5 µM significantly increased Fluo3-fluorescence, but did not significantly modify DCF fluorescence or ceramide abundance. The effect of Terfenadine on annexin-V-binding was significantly blunted but not abolished by removal of extracellular Ca2+. Exposure of human erythrocytes to Ca2+ ionophore ionomycin (1 µM, 15 min triggered annexin-V-binding, an effect augmented by Terfenadine pretreatment (10 µM, 48 hours. Conclusions: Terfenadine triggers phospholipid scrambling of the human erythrocyte cell membrane, an effect in part due to entry of extracellular Ca2+ and in part due to sensitizing human erythrocyte cell membrane scrambling to Ca2+.

  14. Sensitivity of mitochondrial DNA depleted ρ0 cells to H2O2 depends on the plasma membrane status. (United States)

    Tomita, Kazuo; Kuwahara, Yoshikazu; Takashi, Yuko; Tsukahara, Takao; Kurimasa, Akihiro; Fukumoto, Manabu; Nishitani, Yoshihiro; Sato, Tomoaki


    To clarify the relationship between mitochondrial DNA (mtDNA)-depleted ρ0 cells and the cellular sensitivity to hydrogen peroxide (H 2 O 2 ), we established HeLa and SAS ρ0 cell lines and investigated their survival rate in H 2 O 2 , radical scavenging enzymes, plasma membrane potential status, and chronological change in intracellular H 2 O 2 amount under the existence of extracellular hydrogen peroxide compared with the parental cells. The results revealed that ρ0 cells had higher sensitivity to H 2 O 2 than their parental cells, even though the catalase activity of ρ0 cells was up-regulated, and the membrane potential of the ρ0 cells was lower than their parental cells. Furthermore, the internal H 2 O 2 amount significantly increased only in ρ0 cells after 50 μM H 2 O 2 treatment for 1 h. These results suggest that plasma membrane status of ρ0 cells may cause degradation, and the change could lead to enhanced membrane permeability to H 2 O 2 . As a consequence, ρ0 cells have a higher H 2 O 2 sensitivity than the parental cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Na+/K+-pump modulates intercellular communication viainteraction with other membrane transporters in vascularsmooth muscle cells

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Hansen, Anne Kirstine; Nilsson, Holger

    Ouabain, an inhibitor of the Na+/K+-pump, has previously been shown to disturb intercellular communication. Here we test the hypothesis that the communication between vascular smooth muscle cells (SMCs) is regulated through an interaction between the Na+/K+-pump and the Na+/Ca2+-exchanger...... in restricted spaces near the plasma membrane. The intracellular Ca2+ concentration ([Ca2+]i) in individual SMCs was imaged simultaneously with isometric force in rat mesenteric small arteries. Paired cultured rat aortic smooth muscle cells (A7r5) were used as a model for electrical coupling of SMC by measuring...

  16. Development of a membrane electrode assembly process for proton exchange membrane fuel cell (PEMFC)

    International Nuclear Information System (INIS)

    Baldo, Wilians Roberto


    In this work, a Membrane Electrode Assembly (MEA) producing process was developed, involving simple steps, aiming cost reduction and good reproducibility for Proton Exchange Membrane Fuel Cell (PEMFC) commercial applications. The electrodes were produced by spraying ink into both sides of the polymeric membrane, building the catalytic layers, followed by hot pressing of Gas Diffusion Layers (GDL), forming the MEA. This new producing method was called 'Spray and hot pressing hybrid method'. Concerning that all the parameters of spray and hot pressing methods are interdependent, a statistical procedure were used in order to study the mutual variables influences and to optimize the method. This study was earned out in two distinct steps: the first one, where seven variables were considered for the analysis and the second one, where only the variables that interfered in the process performance in the first step were considered for analysis. The results showed that the developed process was adequate, including only simple steps, reaching MEA's performance of 651 m A cm -2 at a potential of 600 mV for catalysts loading of 0,4 mg cm -2 Pt at the anode and 0,6 mg cm -2 Pt at the cathode. This result is compared to available commercial MEA's, with the same fuel cell operations conditions. (author)

  17. Solid Polymer Fuel Cells. Electrode and membrane performance studies

    Energy Technology Data Exchange (ETDEWEB)

    Moeller-Holst, S.


    This doctoral thesis studies aspects of fuel cell preparation and performance. The emphasis is placed on preparation and analysis of low platinum-loading solid polymer fuel cell (SPEC) electrodes. A test station was built and used to test cells within a wide range of real operating conditions, 40-150{sup o}C and 1-10 bar. Preparation and assembling equipment for single SPFCs was designed and built, and a new technique of spraying the catalyst layer directly onto the membrane was successfully demonstrated. Low Pt-loading electrodes (0.1 mg Pt/cm{sup 2}) prepared by the new technique exhibited high degree of catalyst utilization. The performance of single cells holding these electrodes is comparable to state-of-the-art SPFCs. Potential losses in single cell performance are ascribed to irreversibilities by analysing the efficiency of the Solid Oxide Fuel Cell by means of the second law of thermodynamics. The water management in membranes is discussed for a model system and the results are relevant to fuel cell preparation and performance. The new spray deposition technique should be commercially interesting as it involves few steps as well as techniques that are adequate for larger scale production. 115 refs., 43 figs., 18 tabs.

  18. Embelin-Induced Phosphatidylserine Translocation in the Erythrocyte Cell Membrane

    Directory of Open Access Journals (Sweden)

    Ghada Bouguerra


    Full Text Available Background/Aims: The antihelminthic, contraceptive, anti-inflammatory and anticancer phytochemical embelin is at least in part effective against malignancy by inducing suicidal death or apoptosis of tumor cells. Erythrocytes are similarly able to enter suicidal death or eryptosis, which is characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Signaling of eryptosis includes increase of cytosolic Ca2+-activity ([Ca2+]i, ceramide formation, oxidative stress as well as activation of p38 kinase and protein kinase C (PKC. The present study tested, whether and how embelin induces eryptosis. Methods: Phosphatidylserine exposure at the cell surface was estimated from annexin V binding, cell volume from forward scatter, [Ca2+]i from Fluo3-fluorescence, ceramide abundance utilizing specific antibodies and reactive oxygen species (ROS from 2′,7′-dichlorodihydrofluorescein diacetate (DCFDA fluorescence. Results: A 48 hours exposure of human erythrocytes to embelin (≥25 µM significantly increased the percentage of annexin-V-binding cells and hemolysis. Embelin did not significantly modify [Ca2+]i. The effect of embelin on annexin-V-binding was not blunted by removal of extracellular Ca2+, by p38 kinase inhibitor SB203580 (2 µM or by PKC inhibitor staurosporine (1 µM. Embelin did, however, significantly increase the ceramide abundance. Conclusions: Embelin stimulates phospholipid scrambling of the erythrocyte cell membrane, an effect involving ceramide formation.

  19. Proton exchange membrane fuel cell technology for transportation applications

    Energy Technology Data Exchange (ETDEWEB)

    Swathirajan, S. [General Motors R& D Center, Warren, MI (United States)


    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.

  20. Regulation of plant plasma membrane H+- and Ca2+-ATPases by terminal domains

    DEFF Research Database (Denmark)

    Bækgaard, Lone; Fuglsang, Anja Thoe; Palmgren, Michael Gjedde


    In the last few years, major progress has been made to elucidate the structure, function, and regulation of P-type plasma membrane H(+)-and Ca(2+)-ATPases. Even though a number of regulatory proteins have been identified, many pieces are still lacking in order to understand the complete regulator...... mechanisms of these pumps. In plant plasma membrane H(+)- and Ca(2+)-ATPases, autoinhibitory domains are situated in the C- and N-terminal domains, respectively. A model for a common mechanism of autoinhibition is discussed....

  1. Factors Determining the Oxygen Permeability of Biological Membranes: Oxygen Transport Across Eye Lens Fiber-Cell Plasma Membranes. (United States)

    Subczynski, Witold Karol; Widomska, Justyna; Mainali, Laxman


    Electron paramagnetic resonance (EPR) spin-label oximetry allows the oxygen permeability coefficient to be evaluated across homogeneous lipid bilayer membranes and, in some cases, across coexisting membrane domains without their physical separation. The most pronounced effect on oxygen permeability is observed for cholesterol, which additionally induces the formation of membrane domains. In intact biological membranes, integral proteins induce the formation of boundary and trapped lipid domains with a low oxygen permeability. The effective oxygen permeability coefficient across the intact biological membrane is affected not only by the oxygen permeability coefficients evaluated for each lipid domain but also by the surface area occupied by these domains in the membrane. All these factors observed in fiber cell plasma membranes of clear human eye lenses are reviewed here.

  2. A laser microsurgical method of cell wall removal allows detection of large-conductance ion channels in the guard cell plasma membrane (United States)

    Miedema, H.; Henriksen, G. H.; Assmann, S. M.; Evans, M. L. (Principal Investigator)


    Application of patch clamp techniques to higher-plant cells has been subject to the limitation that the requisite contact of the patch electrode with the cell membrane necessitates prior enzymatic removal of the plant cell wall. Because the wall is an integral component of plant cells, and because cell-wall-degrading enzymes can disrupt membrane properties, such enzymatic treatments may alter ion channel behavior. We compared ion channel activity in enzymatically isolated protoplasts of Vicia faba guard cells with that found in membranes exposed by a laser microsurgical technique in which only a tiny portion of the cell wall is removed while the rest of the cell remains intact within its tissue environment. "Laser-assisted" patch clamping reveals a new category of high-conductance (130 to 361 pS) ion channels not previously reported in patch clamp studies on plant plasma membranes. These data indicate that ion channels are present in plant membranes that are not detected by conventional patch clamp techniques involving the production of individual plant protoplasts isolated from their tissue environment by enzymatic digestion of the cell wall. Given the large conductances of the channels revealed by laser-assisted patch clamping, we hypothesize that these channels play a significant role in the regulation of ion content and electrical signalling in guard cells.

  3. Highly durable direct hydrazine hydrate anion exchange membrane fuel cell (United States)

    Sakamoto, Tomokazu; Serov, Alexey; Masuda, Teruyuki; Kamakura, Masaki; Yoshimoto, Koji; Omata, Takuya; Kishi, Hirofumi; Yamaguchi, Susumu; Hori, Akihiro; Horiuchi, Yousuke; Terada, Tomoaki; Artyushkova, Kateryna; Atanassov, Plamen; Tanaka, Hirohisa


    The factors influenced on degradation of direct hydrazine hydrate fuel cells (DHFCs) under operation conditions are analyzed by in situ soft X-ray radiography. A durability of DHFCs is significantly improved by multi-step reaction DHFCs (MSR-DHFCs) approach designed to decrease the crossover of liquid fuel. An open circuit voltage (OCV) as well as cell voltage at 5 mA cm-2 of MSR-DHFC construct with commercial anion exchange membrane (AEM) maintained for over of 3500 h at 60 °C. Furthermore, the commercial proton exchange membrane (PEM) is integrated into AEM of MSR-DHFCs resulting in stable power output of MSR-DHFCs for over than 2800 h at 80 °C.

  4. In vitro membrane reconstitution of the T-cell receptor proximal signaling network. (United States)

    Hui, Enfu; Vale, Ronald D


    T-cell receptor (TCR) phosphorylation is controlled by a complex network that includes Lck, a Src family kinase (SFK), the tyrosine phosphatase CD45 and the Lck-inhibitory kinase Csk. How these competing phosphorylation and dephosphorylation reactions are modulated to produce T-cell triggering is not fully understood. Here we reconstituted this signaling network using purified enzymes on liposomes, recapitulating the membrane environment in which they normally interact. We demonstrate that Lck's enzymatic activity can be regulated over an ~10-fold range by controlling its phosphorylation state. By varying kinase and phosphatase concentrations, we constructed phase diagrams that reveal ultrasensitivity in the transition from the quiescent to the phosphorylated state and demonstrate that co-clustering TCR and Lck or detaching Csk from the membrane can trigger TCR phosphorylation. Our results provide insight into the mechanism of TCR signaling as well as other signaling pathways involving SFKs.

  5. Regulation of the cell cycle by irradiation

    International Nuclear Information System (INIS)

    Akashi, Makoto


    The molecular mechanism of cell proliferation is extremely complex; deregulation results in neoplastic transformation. In eukaryotes, proliferation of cells is finely regulated through the cell cycle. Studies have shown that the cell cycle is regulated by s series of enzymes known as cyclin-dependent kinases (CDKs). The activities of CDKs are controlled by their association with regulatory subunits, cyclins; the expression of cyclins and the activation of the different cyclin-CDK complexes are required for the cell to cycle. Thus, the cell cycle is regulated by activating and inhibiting phosphorylation of the CDK subunits and this program has internal check points at different stages of the cell cycle. When cells are exposed to external insults such as DNA damaging agents, negative regulation of the cell cycle occurs; arrest in either G1 or G2 stage is induced to prevent the cells from prematurely entering into the next stage before DNA is repaired. Recently, a potent inhibitor of CDKs, which inhibits the phosphorylation of retinoblastoma susceptibility (Rb) gene product by cyclin A-CDK2, cyclin E-CDK2, cyclin D1-CDK4, and cyclin D2-CDK4 complexes has been identified. This protein named WAF1, Sdi1, Cip1, or p21 (a protein of Mr 21,000) contains a p53-binding site in its promoter and studies have reported that the expression of WAF1 was directly regulated by p53; cells with loss of p53 activity due to mutational alteration were unable to induce WAF1. This chapter will be focused on the mechanisms of the cell cycle including inhibitors of CDKs, and the induction of WAF1 by irradiation through a pathway independent of p53 will be also described. (author)

  6. Regulation of the cell cycle by irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Akashi, Makoto [National Inst. of Radiological Sciences, Chiba (Japan)


    The molecular mechanism of cell proliferation is extremely complex; deregulation results in neoplastic transformation. In eukaryotes, proliferation of cells is finely regulated through the cell cycle. Studies have shown that the cell cycle is regulated by s series of enzymes known as cyclin-dependent kinases (CDKs). The activities of CDKs are controlled by their association with regulatory subunits, cyclins; the expression of cyclins and the activation of the different cyclin-CDK complexes are required for the cell to cycle. Thus, the cell cycle is regulated by activating and inhibiting phosphorylation of the CDK subunits and this program has internal check points at different stages of the cell cycle. When cells are exposed to external insults such as DNA damaging agents, negative regulation of the cell cycle occurs; arrest in either G1 or G2 stage is induced to prevent the cells from prematurely entering into the next stage before DNA is repaired. Recently, a potent inhibitor of CDKs, which inhibits the phosphorylation of retinoblastoma susceptibility (Rb) gene product by cyclin A-CDK2, cyclin E-CDK2, cyclin D1-CDK4, and cyclin D2-CDK4 complexes has been identified. This protein named WAF1, Sdi1, Cip1, or p21 (a protein of Mr 21,000) contains a p53-binding site in its promoter and studies have reported that the expression of WAF1 was directly regulated by p53; cells with loss of p53 activity due to mutational alteration were unable to induce WAF1. This chapter will be focused on the mechanisms of the cell cycle including inhibitors of CDKs, and the induction of WAF1 by irradiation through a pathway independent of p53 will be also described. (author)

  7. The keratin-binding protein Albatross regulates polarization of epithelial cells


    Sugimoto, Masahiko; Inoko, Akihito; Shiromizu, Takashi; Nakayama, Masanori; Zou, Peng; Yonemura, Shigenobu; Hayashi, Yuko; Izawa, Ichiro; Sasoh, Mikio; Uji, Yukitaka; Kaibuchi, Kozo; Kiyono, Tohru; Inagaki, Masaki


    The keratin intermediate filament network is abundant in epithelial cells, but its function in the establishment and maintenance of cell polarity is unclear. Here, we show that Albatross complexes with Par3 to regulate formation of the apical junctional complex (AJC) and maintain lateral membrane identity. In nonpolarized epithelial cells, Albatross localizes with keratin filaments, whereas in polarized epithelial cells, Albatross is primarily localized in the vicinity of the AJC. Knockdown o...

  8. Binding of 18F by cell membranes and cell walls of Streptococcus mutans

    International Nuclear Information System (INIS)

    Yotis, W.W.; Zeb, M.; McNulty, J.; Kirchner, F.; Reilly, C.; Glendenin, L.


    The binding of 18 F to isolated cell membranes and cell walls of Streptococcus mutans GS-5 or other bacteria was assayed. The attachment of 18 F to these cell envelopes proceeded slowly and reached equilibrium within 60 min. 18 F binding was stimulated by Ca 2+ (1 mM). The binding of 18 F to cellular components was dependent upon the pH, as well as the amount of 18 F and dose of the binder employed. The binding of 18 F by cell walls prepared from fluoride-sensitive and fluoride-resistant cells of S. salivarius and S. mutans did not differ significantly. The pretreatment of cell walls or cell membranes for 60 min at 30 degrees C with 1 mg of RNase, DNase, or trypsin per ml did not influence the binding of 18 F by the walls and membranes of S. mutans GS-5. However, prior exposure of cell membranes to sodium dodecyl sulfate caused a significant reduction in the number of 18 F atoms bound by the membranes. In saturated assay systems, cell membranes of S. mutans GS-5 bound 10(15) to 10(16) atoms of 18 F per mg (dry weight), whereas cell walls from S. mutans GS-5, FA-1, and HS-6 or Actinomyces viscosus T14V and T14AV bound 10(12) to 10(13) atoms of 18 F per mg (dry weight). 18 F in this quantity (10(12) to 10(13) atoms) cannot be detected with the fluoride electrode. The data provide, for the first time, a demonstration of 18 F binding by cell membranes and walls of oral flora

  9. Polybenzimidazole membranes for zero gap alkaline electrolysis cells

    DEFF Research Database (Denmark)

    Kraglund, Mikkel Rykær; Aili, David; Christensen, Erik

    Membranes of m-PBI doped in KOH (aq), 15-35 wt%, show high ionic conductivity in the temperature range 20-80 ºC. In electrolysis cells with nickel foam electrodes m-PBI membranesprovide low internal resistance. With a 60 µm membraneat 80ºC in 20 wt% KOH,1000 mA/cm2 is achieved at 2.25....

  10. Proteomic analysis of isolated membrane fractions from superinvasive cancer cells


    Dowling, Paul; Meleady, Paula; Dowd, Andrew; Henry, Michael; Glynn, Sharon; Clynes, Martin


    The superinvasive phenotype exhibited by paclitaxel-selected variants of an in vitro invasive clonal population of the human cancer cell line, MDA-MB-435S were examined using DIGE (Fluorescence 2-D Difference Gel Electrophoresis) and mass spectrometry. Isolation of membrane proteins from the MDA-MB-435S-F/Taxol-10p4p and parental populations was performed by temperature-dependent phase partitioning using the detergent Triton X-114. Subsequent DIGE-generated data analysed using Decyde...

  11. The effect of propofol on plasma membrane ultrastructure in the intact cells (United States)

    Jin, Weixiang; Pralle, Arnd

    The mechanism of general anesthesia is still unknown. One drug used for human anesthesia, propofol, has been shown to interact with some ligand gated ion-channels, but also easily dissolves in the lipid bilayer and alters fluidity. Which mechanism dominates or even how anesthesia arises are unclear. We study the influence of propofol on plasma membrane (PM) ultrastructure in intact cells. In the PM, transient submicroscopic nanodomains form by interactions between lipid-acyl-chains or lipid head groups, stabilized by cholesterol. In addition, membrane cytoskeleton further regulates the nanodomains, which then regulate signaling. We study transient propofol effects on these domains from low to clinically relevant propofol concentrations by analyzing diffusion of GFP-tagged outer leaflet/inner leaflet membrane proteins. Using bimFCS we measure diffusion on multiple length scales simultaneously. We observe that at low propofol concentrations, the nanodomains trap GPI-mGFP less, consistent with studies showing that propofol decreases the phase transition temperature of membrane derived vesicles. Interestingly, at clinical relevant concentrations of propofol, the nanodomains trap GPI-mGFP more strongly. This is only observed at 37C. By inhibiting myosin activity or actin filaments (de-)polymerization, we find that the activity of actin filaments further alters the behavior of cholesterol nanodomains due to propofol. We compare the effect of propofol and its analog confirming specificity.

  12. The extracellular membrane-proximal domain of membrane-bound IgE restricts B cell activation by limiting B cell antigen receptor surface expression. (United States)

    Vanshylla, Kanika; Opazo, Felipe; Gronke, Konrad; Wienands, Jürgen; Engels, Niklas


    Immunoglobulin E (IgE) antibodies are key mediators of allergic reactions. Due to their potentially harmful anaphylactic properties, their production is tightly regulated. The membrane-bound isoform of IgE (mIgE), which is an integral component of the B cell antigen receptor, has been shown to be critical for the regulation of IgE responses in mice. In primate species including humans, mIgE can be expressed in two isoforms that are produced by alternative splicing of the primary ε Ig heavy chain transcript, and differ in the absence or presence of an extracellular membrane-proximal domain (EMPD) consisting of 52 amino acids. However, the function of the EMPD remains unclear. Here, we demonstrate that the EMPD restricts surface expression of mIgE-containing BCRs in human and murine B cells. The EMPD does not interfere with BCR assembly but acts as an autonomous endoplasmic reticulum retention domain. Limited surface expression of EMPD-containing mIgE-BCRs caused impaired activation of intracellular signaling cascades and hence represents a regulatory mechanism that may control the production of potentially anaphylactic IgE antibodies in primate species. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Collective cell behavior on basement membranes floating in space (United States)

    Ellison, Sarah; Bhattacharjee, Tapomoy; Morley, Cameron; Sawyer, W.; Angelini, Thomas

    The basement membrane is an essential part of the polarity of endothelial and epithelial tissues. In tissue culture and organ-on-chip devices, monolayer polarity can be established by coating flat surfaces with extracellular matrix proteins and tuning the trans-substrate permeability. In epithelial 3D culture, spheroids spontaneously establish inside-out polarity, morphing into hollow shell-like structures called acini, generating their own basement membrane on the inner radius of the shell. However, 3D culture approaches generally lack the high degree of control provided by the 2D culture plate or organ-on-chip devices, making it difficult to create more faithful in vitro tissue models with complex surface curvature and morphology. Here we present a method for 3D printing complex basement membranes covered in cells. We 3D print collagen-I and Matrigel into a 3D growth medium made from jammed microgels. This soft, yielding material allows extracellular matrix to be formed as complex surfaces and shapes, floating in space. We then distribute MCF10A epithelial cells across the polymerized surface. We envision employing this strategy to study 3D collective cell behavior in numerous model tissue layers, beyond this simple epithelial model.

  14. Fluorine NMR-based screening on cell membrane extracts. (United States)

    Veronesi, Marina; Romeo, Elisa; Lambruschini, Chiara; Piomelli, Daniele; Bandiera, Tiziano; Scarpelli, Rita; Garau, Gianpiero; Dalvit, Claudio


    The possibility of measuring the action of inhibitors of specific enzymatic reactions in intact cells, cell lysates or membrane preparations represents a major advance in the lead discovery process. Despite the relevance of assaying in physiological conditions, only a small number of biophysical techniques, often requiring complex set-up, are applicable to these sample types. Here, we demonstrate the first application of n-fluorine atoms for biochemical screening (n-FABS), a homogeneous and versatile assay based on (19) F NMR spectroscopy, to the detection of high- and low-affinity inhibitors of a membrane enzyme in cell extracts and determination of their IC50 values. Our approach can allow the discovery of novel binding fragments against targets known to be difficult to purify or where membrane-association is required for activity. These results pave the way for future applications of the methodology to these relevant and complex biological systems. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Water Soluble Polymers as Proton Exchange Membranes for Fuel Cells

    Directory of Open Access Journals (Sweden)

    Bing-Joe Hwang


    Full Text Available The relentless increase in the demand for useable power from energy-hungry economies continues to drive energy-material related research. Fuel cells, as a future potential power source that provide clean-at-the-point-of-use power offer many advantages such as high efficiency, high energy density, quiet operation, and environmental friendliness. Critical to the operation of the fuel cell is the proton exchange membrane (polymer electrolyte membrane responsible for internal proton transport from the anode to the cathode. PEMs have the following requirements: high protonic conductivity, low electronic conductivity, impermeability to fuel gas or liquid, good mechanical toughness in both the dry and hydrated states, and high oxidative and hydrolytic stability in the actual fuel cell environment. Water soluble polymers represent an immensely diverse class of polymers. In this comprehensive review the initial focus is on those members of this group that have attracted publication interest, principally: chitosan, poly (ethylene glycol, poly (vinyl alcohol, poly (vinylpyrrolidone, poly (2-acrylamido-2-methyl-1-propanesulfonic acid and poly (styrene sulfonic acid. The paper then considers in detail the relationship of structure to functionality in the context of polymer blends and polymer based networks together with the effects of membrane crosslinking on IPN and semi IPN architectures. This is followed by a review of pore-filling and other impregnation approaches. Throughout the paper detailed numerical results are given for comparison to today’s state-of-the-art Nafion® based materials.

  16. Rupturing Giant Plasma Membrane Vesicles to Form Micron-sized Supported Cell Plasma Membranes with Native Transmembrane Proteins. (United States)

    Chiang, Po-Chieh; Tanady, Kevin; Huang, Ling-Ting; Chao, Ling


    Being able to directly obtain micron-sized cell blebs, giant plasma membrane vesicles (GPMVs), with native membrane proteins and deposit them on a planar support to form supported plasma membranes could allow the membrane proteins to be studied by various surface analytical tools in native-like bilayer environments. However, GPMVs do not easily rupture on conventional supports because of their high protein and cholesterol contents. Here, we demonstrate the possibility of using compression generated by the air-water interface to efficiently rupture GPMVs to form micron-sized supported membranes with native plasma membrane proteins. We demonstrated that not only lipid but also a native transmembrane protein in HeLa cells, Aquaporin 3 (AQP3), is mobile in the supported membrane platform. This convenient method for generating micron-sized supported membrane patches with mobile native transmembrane proteins could not only facilitate the study of membrane proteins by surface analytical tools, but could also enable us to use native membrane proteins for bio-sensing applications.

  17. Proton Exchange Membrane Fuel Cell Reversal: A Review

    Directory of Open Access Journals (Sweden)

    Congwei Qin


    Full Text Available The H2/air-fed proton exchange membrane fuel cell (PEMFC has two major problems: cost and durability, which obstruct its pathway to commercialization. Cell reversal, which would create irreversible damage to the fuel cell and shorten its lifespan, is caused by reactant starvation, load change, low catalyst performance, and so on. This paper will summarize the causes, consequences, and mitigation strategies of cell reversal of PEMFC in detail. A description of potential change in the anode and cathode and the differences between local starvation and overall starvation are reviewed, which gives a framework for comprehending the origins of cell reversal. According to the root factor of cell starvation, i.e., fuel cells do not satisfy the requirements of electrons and protons of normal anode and cathode chemical reactions, we will introduce specific methods to mitigate or prevent fuel cell damage caused by cell reversal in the view of system management strategies and component material modifications. Based on a comprehensive understanding of cell reversal, it is beneficial to operate a fuel cell stack and extend its lifetime.

  18. New insights into circulating FABP4: Interaction with cytokeratin 1 on endothelial cell membranes. (United States)

    Saavedra, Paula; Girona, Josefa; Bosquet, Alba; Guaita, Sandra; Canela, Núria; Aragonès, Gemma; Heras, Mercedes; Masana, Lluís


    Fatty acid-binding protein 4 (FABP4) is an adipose tissue-secreted adipokine that is involved in the regulation of energetic metabolism and inflammation. Increased levels of circulating FABP4 have been detected in individuals with cardiovascular risk factors. Recent studies have demonstrated that FABP4 has a direct effect on peripheral tissues, specifically promoting vascular dysfunction; however, its mechanism of action is unknown. The objective of this work was to assess the specific interactions between exogenous FABP4 and the plasma membranes of endothelial cells. Immunofluorescence assays showed that exogenous FABP4 localized along the plasma membranes of human umbilical vein endothelial cells (HUVECs), interacting specifically with plasma membrane proteins. Anti-FABP4 immunoblotting revealed two covalent protein complexes containing FABP4 and its putative receptor; these complexes were approximately 108 kDa and 77 kDa in size. Proteomics and mass spectrometry experiments revealed that cytokeratin 1 (CK1) was the FABP4-binding protein. An anti-CK1 immunoblot confirmed the presence of CK1. FABP4-CK1 complexes were also detected in HAECs, HCASMCs, HepG2 cells and THP-1 cells. Pharmacological FABP4 inhibition by BMS309403 results in a slight decrease in the formation of these complexes, indicating that fatty acids may play a role in FABP4 functionality. In addition, we demonstrated that exogenous FABP4 crosses the plasma membrane to enter the cytoplasm and nucleus in HUVECs. These findings indicate that exogenous FABP4 interacts with plasma membrane proteins, specifically CK1. These data contribute to our current knowledge regarding the mechanism of action of circulating FABP4.

  19. Dynamic analysis of magnetic nanoparticles crossing cell membrane

    Energy Technology Data Exchange (ETDEWEB)

    Pedram, Maysam Z. [Department of Mechanical Engineering, Sharif University of Tech., Azadi Ave., Tehran (Iran, Islamic Republic of); Shamloo, Amir, E-mail: [Department of Mechanical Engineering, Sharif University of Tech., Azadi Ave., Tehran (Iran, Islamic Republic of); Ghafar-Zadeh, Ebrahim [Biologically-Inspired Sensors and Actuators Laboratory, Department of Electrical Engineering and Computer science, York University, Keel Street, Toronto (Canada); Alasty, Aria, E-mail: [Department of Mechanical Engineering, Sharif University of Tech., Azadi Ave., Tehran (Iran, Islamic Republic of)


    Nowadays, nanoparticles (NPs) are used in a variety of biomedical applications including brain disease diagnostics and subsequent treatments. Among the various types of NPs, magnetic nanoparticles (MNPs) have been implemented by many research groups for an array of life science applications. In this paper, we studied MNPs controlled delivery into the endothelial cells using a magnetic field. Dynamics equations of MNPs were defined in the continuous domain using control theory methods and were applied to crossing the cell membrane. This study, dedicated to clinical and biomedical research applications, offers a guideline for the generation of a magnetic field required for the delivery of MNPs.

  20. Hydrocarbon-based fuel cell membranes: Sulfonated crosslinked poly(1,3-cyclohexadiene) membranes for high temperature polymer electrolyte fuel cells


    Deng, Suxiang; Hassan, Mohammad K.; Mauritz, Kenneth A.; Mays, Jimmy W.


    High temperature fuel cell membranes based on poly(1,3-cyclohexadiene) were prepared by a Polymerization-Crosslinking-Sulfonation (PCS) approach, and a broad range of membrane compositions were achieved using various sulfonating reagents and reaction conditions. Membranes were characterized for their proton conductivity and thermal degradation behavior. Some of the membranes showed up to a 68% increase in proton conductivity as compared to Nafion under the same conditions (100% relative humid...

  1. Experimental study on the membrane electrode assembly of a proton exchange membrane fuel cell: effects of microporous layer, membrane thickness and gas diffusion layer hydrophobic treatment

    International Nuclear Information System (INIS)

    Ferreira, Rui B.; Falcão, D.S.; Oliveira, V.B.; Pinto, A.M.F.R.


    Highlights: • EIS is employed to investigate the MEA design of a PEM fuel cell. • Effects of MPL, membrane thickness and GDL hydrophobic treatment are studied. • MPL increases cell output at low to medium currents but reduces it at high currents. • Better results are obtained when employing a thinner Nafion membrane. • GDL hydrophobic treatment improves the cell performance. - Abstract: In this study, electrochemical impedance spectroscopy (EIS) is employed to analyze the influence of microporous layer (MPL), membrane thickness and gas diffusion layer (GDL) hydrophobic treatment in the performance of a proton exchange membrane (PEM) fuel cell. Results show that adding a MPL increases cell performance at low to medium current densities. Because lower ohmic losses are observed when applying a MPL, such improvement is attributed to a better hydration state of the membrane. The MPL creates a pressure barrier for water produced at the cathode, forcing it to travel to the anode side, therefore increasing the water content in the membrane. However, at high currents, this same phenomenon seems to have intensified liquid water flooding in the anode gas channels, increasing mass transfer losses and reducing the cell performance. Decreasing membrane thickness results into considerably higher performances, due to a decrease in ohmic resistance. Moreover, at low air humidity operation, a rapid recovery from dehydration is observed when a thinner membrane is employed. The GDL hydrophobic treatment significantly improves the cell performance. Untreated GDLs appear to act as water-traps that not only hamper reactants transport to the reactive sites but also impede the proper humidification of the cell. From the different designs tested, the highest maximum power density is obtained from that containing a MPL, a thinner membrane and treated GDLs.

  2. Cell Surface and Membrane Engineering: Emerging Technologies and Applications

    Directory of Open Access Journals (Sweden)

    Christopher T. Saeui


    Full Text Available Membranes constitute the interface between the basic unit of life—a single cell—and the outside environment and thus in many ways comprise the ultimate “functional biomaterial”. To perform the many and often conflicting functions required in this role, for example to partition intracellular contents from the outside environment while maintaining rapid intake of nutrients and efflux of waste products, biological membranes have evolved tremendous complexity and versatility. This article describes how membranes, mainly in the context of living cells, are increasingly being manipulated for practical purposes with drug discovery, biofuels, and biosensors providing specific, illustrative examples. Attention is also given to biology-inspired, but completely synthetic, membrane-based technologies that are being enabled by emerging methods such as bio-3D printers. The diverse set of applications covered in this article are intended to illustrate how these versatile technologies—as they rapidly mature—hold tremendous promise to benefit human health in numerous ways ranging from the development of new medicines to sensitive and cost-effective environmental monitoring for pathogens and pollutants to replacing hydrocarbon-based fossil fuels.

  3. Calcium-regulated anion channels in the plasma membrane of Lilium longiflorum pollen protoplasts. (United States)

    Tavares, Bárbara; Dias, Pedro Nuno; Domingos, Patrícia; Moura, Teresa Fonseca; Feijó, José Alberto; Bicho, Ana


    • Currents through anion channels in the plasma membrane of Lilium longiflorum pollen grain protoplasts were studied under conditions of symmetrical anionic concentrations by means of patch-clamp whole-cell configuration. • With Cl(-) -based intra- and extracellular solutions, three outward-rectifying anion conductances, I(Cl1) , I(Cl2) and I(Cl3) , were identified. These three activities were discriminated by differential rundown behaviour and sensitivity to 5-nitro-2-(phenylpropylamino)-benzoate (NPPB), which could not be attributed to one or more channel types. All shared strong outward rectification, activated instantaneously and displayed a slow time-dependent activation for positive potentials. All showed modulation by intracellular calcium ([Ca(2+) ](in) ), increasing intensity from 6.04 nM up to 0.5 mM (I(Cl1) ), or reaching a maximum value with 8.50 μM (I(Cl2) and I(Cl3) ). • After rundown, the anionic currents measured using NO(3) (-) -based solutions were indistinguishable, indicating that the permeabilities of the channels for Cl(-) and NO(3) (-) are similar. Additionally, unitary anionic currents were measured from outside-out excised patches, confirming the presence of individual anionic channels. • This study shows for the first time the presence of a large anionic conductance across the membrane of pollen protoplasts, resulting from the presence of Ca(2+) -regulated channels. A similar conductance was also found in germinated pollen. We hypothesize that these putative channels may be responsible for the large anionic fluxes previously detected by means of self-referencing vibrating probes. © 2011 The Authors. New Phytologist © 2011 New Phytologist Trust.

  4. Lowering the platinum loading of high temperature polymer electrolyte membrane fuel cells with acid doped polybenzimidazole membranes

    DEFF Research Database (Denmark)

    Fernandez, Santiago Martin; Li, Qingfeng; Jensen, Jens Oluf


    Membrane electrode assemblies (MEAs) with ultra-low Pt loading electrodes were prepared for high temperature polymer electrolyte membrane fuel cells (HT-PEMFCs) based on acid doped polybenzimidazole. With no electrode binders or ionomers, the triple phase boundary of the catalyst layer was establ...

  5. A hybrid microbial fuel cell membrane bioreactor with a conductive ultrafiltration membrane biocathode for wastewater treatment

    KAUST Repository

    Malaeb, Lilian


    A new hybrid, air-biocathode microbial fuel cell-membrane bioreactor (MFC-MBR) system was developed to achieve simultaneous wastewater treatment and ultrafiltration to produce water for direct reclamation. The combined advantages of this system were achieved by using an electrically conductive ultrafiltration membrane as both the cathode and the membrane for wastewater filtration. The MFC-MBR used an air-biocathode, and it was shown to have good performance relative to an otherwise identical cathode containing a platinum catalyst. With 0.1 mm prefiltered domestic wastewater as the feed, the maximum power density was 0.38 W/m2 (6.8 W/m3) with the biocathode, compared to 0.82 W/m2 (14.5 W/m3) using the platinum cathode. The permeate quality from the biocathode reactor was comparable to that of a conventional MBR, with removals of 97% of the soluble chemical oxygen demand, 97% NH3-N, and 91% of total bacteria (based on flow cytometry). The permeate turbidity was <0.1 nephelometric turbidity units. These results show that a biocathode MFC-MBR system can achieve high levels of wastewater treatment with a low energy input due to the lack of a need for wastewater aeration. © 2013 American Chemical Society.

  6. Cell cycle regulation by the bacterial nucleoid


    Adams, David William; Wu, Ling Juan; Errington, Jeff


    Division site selection presents a fundamental challenge to all organisms. Bacterial cells are small and the chromosome (nucleoid) often fills most of the cell volume. Thus, in order to maximise fitness and avoid damaging the genetic material, cell division must be tightly co-ordinated with chromosome replication and segregation. To achieve this, bacteria employ a number of different mechanisms to regulate division site selection. One such mechanism, termed nucleoid occlusion, allows the nucl...

  7. Better Proton-Conducting Polymers for Fuel-Cell Membranes (United States)

    Narayan, Sri; Reddy, Prakash


    Polyoxyphenylene triazole sulfonic acid has been proposed as a basis for development of improved proton-conducting polymeric materials for solid-electrolyte membranes in hydrogen/air fuel cells. Heretofore, the proton-conducting membrane materials of choice have been exemplified by a family of perfluorosulfonic acid-based polymers (Nafion7 or equivalent). These materials are suitable for operation in the temperature of 75 to 85 C, but in order to reduce the sizes and/or increase the energy-conversion efficiencies of fuel-cell systems, it would be desirable to increase temperatures to as high as 120 C for transportation applications, and to as high as 180 C for stationary applications. However, at 120 C and at relative humidity values below 50 percent, the loss of water from perfluorosulfonic acid-based polymer membranes results in fuel-cell power densities too low to be of practical value. Therefore, membrane electrolyte materials that have usefully high proton conductivity in the temperature range of 180 C at low relative humidity and that do not rely on water for proton conduction at 180 C would be desirable. The proposed polyoxyphenylene triazole sulfonic acid-based materials have been conjectured to have these desirable properties. These materials would be free of volatile or mobile acid constituents. The generic molecular structure of these materials is intended to exploit the fact, demonstrated in previous research, that materials that contain ionizable acid and base groups covalently attached to thermally stable polymer backbones exhibit proton conduction even in the anhydrous state.

  8. A direct ascorbate fuel cell with an anion exchange membrane (United States)

    Muneeb, Omar; Do, Emily; Tran, Timothy; Boyd, Desiree; Huynh, Michelle; Ghosn, Gregory; Haan, John L.


    Ascorbic Acid (Vitamin C) is investigated as a renewable alternative fuel for alkaline direct liquid fuel cells (DLFCs). The environmentally- and biologically-friendly compound, L-ascorbic acid (AA) has been modeled and studied experimentally under acidic fuel cell conditions. In this work, we demonstrate that ascorbic acid is a more efficient fuel in alkaline media than in acidic media. An operating direct ascorbate fuel cell is constructed with the combination of L-ascorbic acid and KOH as the anode fuel, air or oxygen as the oxidant, a polymer anion exchange membrane, metal or carbon black anode materials and metal cathode catalyst. Operation of the fuel cell at 60 °C using 1 M AA and 1 M KOH as the anode fuel and electrolyte, respectively, and oxygen gas at the cathode, produces a maximum power density of 73 mW cm-2, maximum current density of 497 mA cm-2 and an open circuit voltage of 0.90 V. This performance is significantly greater than that of an ascorbic acid fuel cell with a cation exchange membrane, and it is competitive with alkaline DLFCs fueled by alcohols.

  9. Noninvasive microelectrode ion flux estimation technique (MIFE) for the study of the regulation of root membrane transport by cyclic nucleotides

    KAUST Repository

    Ordoñez, Natalia Maria


    Changes in ion permeability and subsequently intracellular ion concentrations play a crucial role in intracellular and intercellular communication and, as such, confer a broad array of developmental and adaptive responses in plants. These changes are mediated by the activity of plasma-membrane based transport proteins many of which are controlled by cyclic nucleotides and/or other signaling molecules. The MIFE technique for noninvasive microelectrode ion flux measuring allows concurrent quantification of net fluxes of several ions with high spatial (μm range) and temporal (ca. 5 s) resolution, making it a powerful tool to study various aspects of downstream signaling events in plant cells. This chapter details basic protocols enabling the application of the MIFE technique to study regulation of root membrane transport in general and cyclic nucleotide mediated transport in particular. © Springer Science+Business Media New York 2013.

  10. New proton conducting membranes for fuel cell applications

    Energy Technology Data Exchange (ETDEWEB)

    Sukumar, P.R.


    In order to synthesize proton-conducting materials which retain acids in the membrane during fuel cell operating conditions, the synthesis of poly(vinylphosphonic acid) grafted polybenzimidazole (PVPA grafted PBI) and the fabrication of multilayer membranes are mainly focussed in this dissertation. Synthesis of PVPA grafted PBI membrane can be done according to ''grafting through'' method. In ''grafting through'' method (or macromonomer method), monomer (e.g., vinylphosphonic acid) is radically copolymerized with olefin group attached macromonomer (e.g., allyl grafted PBI and vinylbenzyl grafted PBI). This approach is inherently limited to synthesize graft-copolymer with well-defined architectural and structural parameters. The incorporation of poly(vinylphosphonic acid) into PBI lead to improvements in proton conductivity up to 10-2 S/cm. Regarding multilayer membranes, the proton conducting layer-by-layer (LBL) assembly of polymers by various strong acids such as poly(vinylphosphonic acid), poly(vinylsulfonic acid) and poly(styrenesulfonic acid) paired with basic polymers such as poly(4-vinylimidazole) and poly(benzimidazole), which are appropriate for Proton Exchange Membrane Fuel Cell applications have been described. Proton conductivity increases with increasing smoothness of the film and the maximum measured conductivity was 10-4 S/cm at 25A C. Recently, anhydrous proton-conducting membranes with flexible structural backbones, which show proton-conducting properties comparable to Nafion have been focus of current research. The flexible backbone of polymer chains allow for a high segmental mobility and thus, a sufficiently low glass transition temperature (Tg), which is an essential factor to reach highly conductive systems. Among the polymers with a flexible chain backbone, poly(vinylphosphonic acid), poly(vinylbenzylphosphonic acid), poly(2-vinylbenzimidazole), poly(4-styrenesulfonic acid), poly(4-vinylimidazole), poly

  11. Angiogenic sprouting is regulated by endothelial cell expression of Slug. (United States)

    Welch-Reardon, Katrina M; Ehsan, Seema M; Wang, Kehui; Wu, Nan; Newman, Andrew C; Romero-Lopez, Monica; Fong, Ashley H; George, Steven C; Edwards, Robert A; Hughes, Christopher C W


    The Snail family of zinc-finger transcription factors are evolutionarily conserved proteins that control processes requiring cell movement. Specifically, they regulate epithelial-to-mesenchymal transitions (EMT) where an epithelial cell severs intercellular junctions, degrades basement membrane and becomes a migratory, mesenchymal-like cell. Interestingly, Slug expression has been observed in angiogenic endothelial cells (EC) in vivo, suggesting that angiogenic sprouting may share common attributes with EMT. Here, we demonstrate that sprouting EC in vitro express both Slug and Snail, and that siRNA-mediated knockdown of either inhibits sprouting and migration in multiple in vitro angiogenesis assays. We find that expression of MT1-MMP, but not of VE-Cadherin, is regulated by Slug and that loss of sprouting as a consequence of reduced Slug expression can be reversed by lentiviral-mediated re-expression of MT1-MMP. Activity of MMP2 and MMP9 are also affected by Slug expression, likely through MT1-MMP. Importantly, we find enhanced expression of Slug in EC in human colorectal cancer samples compared with normal colon tissue, suggesting a role for Slug in pathological angiogenesis. In summary, these data implicate Slug as an important regulator of sprouting angiogenesis, particularly in pathological settings.

  12. Thermal Regulation of Membrane Lipid Fluidity by a Two-Component System in "Bacillus Subtilis" (United States)

    Bredeston, L. M.; Marciano, D.; Albanesi, D.; De Mendoza, D.; Delfino, J. M.


    This article describes a simple and robust laboratory exercise on the regulation of membrane unsaturated fatty acid composition in bacteria by a decrease in growth temperature. We take advantage of the well characterized Des pathway of "Bacillus subtilis", composed of a [delta]5-desaturase (encoded by the "des" gene) and the canonical…

  13. Molecular mechanism of α-tocopheryl-phosphate transport across the cell membrane

    International Nuclear Information System (INIS)

    Negis, Yesim; Meydani, Mohsen; Zingg, Jean-Marc; Azzi, Angelo


    α-Tocopheryl-phosphate (α-TP) is synthesized and hydrolyzed in animal cells and tissues where it modulates several functions. α-TP is more potent than α-T in inhibiting cell proliferation, down-regulating CD36 transcription, inhibiting atherosclerotic plaque formation. Administration of α-TP to cells or animals requires its transfer through membranes, via a transporter. We show here that α-TP is passing the plasma membrane via a system that is inhibited by glibenclamide and probenecid, inhibitors of a number of transporters. Glibenclamide and probenecid prevent dose-dependently α-TP inhibition of cell proliferation. The two inhibitors act on ATP binding cassette (ABC) and organic anion transporters (OAT). Since ABC transporters function to export solutes and α-TP is transported into cells, it may be concluded that α-TP transport may occur via an OAT family member. Due to the protection by glibenclamide and probenecid on the α-TP induced cell growth inhibition it appears that α-TP acts after its uptake inside cells

  14. H+ channels in embryonic Biomphalaria glabrata cell membranes: Putative roles in snail host-schistosome interactions.

    Directory of Open Access Journals (Sweden)

    Brandon J Wright


    Full Text Available The human blood fluke Schistosoma mansoni causes intestinal schistosomiasis, a widespread neglected tropical disease. Infection of freshwater snails Biomphalaria spp. is an essential step in the transmission of S. mansoni to humans, although the physiological interactions between the parasite and its obligate snail host that determine success or failure are still poorly understood. In the present study, the B. glabrata embryonic (Bge cell line, a widely used in vitro model for hemocyte-like activity, was used to investigate membrane properties, and assess the impact of larval transformation proteins (LTP on identified ion channels. Whole-cell patch clamp recordings from Bge cells demonstrated that a Zn2+-sensitive H+ channel serves as the dominant plasma membrane conductance. Moreover, treatment of Bge cells with Zn2+ significantly inhibited an otherwise robust production of reactive oxygen species (ROS, thus implicating H+ channels in the regulation of this immune function. A heat-sensitive component of LTP appears to target H+ channels, enhancing Bge cell H+ current over 2-fold. Both Bge cells and B. glabrata hemocytes express mRNA encoding a hydrogen voltage-gated channel 1 (HVCN1-like protein, although its function in hemocytes remains to be determined. This study is the first to identify and characterize an H+ channel in non-neuronal cells of freshwater molluscs. Importantly, the involvement of these channels in ROS production and their modulation by LTP suggest that these channels may function in immune defense responses against larval S. mansoni.

  15. Preparation of Highly Sulfonated Ultra-Thin Proton-Exchange Polymer Membranes for Proton Exchange Membrane Fuel Cells (United States)

    Jiang, Zhongqing; Meng, Yuedong; Jiang, Zhong-Jie; Shi, Yicai

    Sulfonated ultra-thin proton-exchange polymer membrane carrying pyridine groups was made from a plasma polymerization of styrene, 2-vinylpyridine, and trifluoromethanesulfonic acid by after-glow capacitively coupled discharge technique. Pyridine groups tethered to the polymer backbone acts as a medium through the basic nitrogen for transfer of protons between the sulfonic acid groups of proton exchange membrane. It shows that the method using present technology could effectively depress the degradation of monomers during the plasma polymerization. Spectroscopic analyses reveal that the obtained membranes are highly functionalized with proton exchange groups and have higher proton conductivity. Thus, the membranes are expected to be used in direct methanol fuel cells.

  16. Modeling Of Proton Exchange Membrane Fuel Cell Systems

    DEFF Research Database (Denmark)

    Nielsen, Mads Pagh

    The objective of this doctoral thesis was to develop reliable steady-state and transient component models suitable to asses-, develop- and optimize proton exchange membrane (PEM) fuel cell systems. Several components in PEM fuel cell systems were characterized and modeled. The developed component......- and system models match experimental data from the literature. However, limited data were available for verification so further work is necessary to confirm detailed aspects of the models. It is nonetheless expected that the developed models will be useful for system modeling and optimization of PEM fuel...... cell systems. Consequences of indirectly fueling PEM stacks with hydrocarbons using reforming technology were investigated using a PEM stack model including CO poisoning kinetics and a transient Simulink steam reforming system model. Aspects regarding the optimization of PEM fuel cell systems...

  17. Protonic conductors for proton exchange membrane fuel cells: An overview

    Directory of Open Access Journals (Sweden)

    Jurado Ramon Jose


    Full Text Available At present, Nation, which is a perfluorinated polymer, is one of the few materials that deliver the set of chemical and mechanical properties required to perform as a good electrolyte in proton exchange membrane fuel cells (PEMFCs. However, Nation presents some disadvantages, such as limiting the operational temperature of the fuel system (So°C, because of its inability to retain water at higher temperatures and also suffers chemical crossover. In addition to these restrictions, Nation membranes are very expensive. Reducing costs and using environmentally friendly materials are good reasons to make a research effort in this field in order to achieve similar or even better fuel-cell performances. Glass materials of the ternary system SiO2-ZrO2-P2O5, hybrid materials based on Nation, and nanopore ceramic membranes based on SiO2 TiO2, Al2O3, etc. are considered at present, as promising candidates to replace Nation as the electrolyte in PEMFCs. These types of materials are generally prepared by sol-gel processes in order to tailor their channel-porous structure and pore size. In this communication, the possible candidates in the near future as electrolytes (including other polymers different than Nation in PEMFCs are briefly reviewed. Their preparation methods, their electrical transport properties and conduction mechanisms are considered. The advantages and disadvantages of these materials with respect to Nation are also discussed.

  18. Trans-cis isomerization of lipophilic dyes probing membrane microviscosity in biological membranes and in live cells. (United States)

    Chmyrov, Volodymyr; Spielmann, Thiemo; Hevekerl, Heike; Widengren, Jerker


    Membrane environment and fluidity can modulate the dynamics and interactions of membrane proteins and can thereby strongly influence the function of cells and organisms in general. In this work, we demonstrate that trans-cis isomerization of lipophilic dyes is a useful parameter to monitor packaging and fluidity of biomembranes. Fluorescence fluctuations, generated by trans-cis isomerization of the thiocarbocyanine dye Merocyanine 540 (MC540), were first analyzed by fluorescence correlation spectroscopy (FCS) in different alcohol solutions. Similar isomerization kinetics of MC540 in lipid vesicles could then also be monitored, and the influence of lipid polarity, membrane curvature, and cholesterol content was investigated. While no influence of membrane curvature and lipid polarity could be observed, a clear decrease in the isomerization rates could be observed with increasing cholesterol contents in the vesicle membranes. Finally, procedures to spatially map photoinduced and thermal isomerization rates on live cells by transient state (TRAST) imaging were established. On the basis of these procedures, MC540 isomerization was studied on live MCF7 cells, and TRAST images of the cells at different temperatures were found to reliably detect differences in the isomerization parameters. Our studies indicate that trans-cis isomerization is a useful parameter for probing membrane dynamics and that the TRAST imaging technique can provide spatial maps of photoinduced isomerization as well as both photoinduced and thermal back-isomerization, resolving differences in local membrane microviscosity in live cells.

  19. Performance of proton exchange membrane fuel cells at elevated temperature

    International Nuclear Information System (INIS)

    Shyu, Jin-Cherng; Hsueh, Kan-Lin; Tsau, Fanghei


    Highlights: → At 1 atm, cell has best performance (∼1300 mA/cm at 0.6 V) at 100 deg. C and RH = 100%. → The A value in Eq. increased with increases in the back pressure and RH. →R i dramatically decreased at back pressure of 1 atm. → At each RH, R i decreased and then increased as cell temperature increased at 1 atm. - Abstract: The polarization curves of a single PEMFC having a Nafion membrane fed with H 2 /O 2 with relative humidity (RH) of 35%, 70% and 100% were measured at cell temperatures ranging from 65 deg. C to 120 deg. C at back pressures of 0 atm and 1 atm, respectively. Measured results showed that the best cell performance at 0.6 V operated within 65-120 deg. C at zero back pressure was 1000 mA cm -2 at 65 deg. C and RH = 100%, while the best cell performance at 1 atm back pressure was 1300 mA cm -2 at 100 deg. C and RH = 100%. Based on the analysis of impedance data measured at anode and cathode humidification temperatures of 90 deg. C and cell temperature of 100 deg. C at back pressures of 0 and 1 atm (90-100p0 and 90-100p1), it could be found that the membrane resistance was reduced and the catalyst became more active as the back pressure increases. The present results showed that increasing back pressure was able to dramatically improve cell performance and the effect of the back pressure surpassed that of humidification in the internal resistance of cell.

  20. Porous polybenzimidazole membranes doped with phosphoric acid: Preparation and application in high-temperature proton-exchange-membrane fuel cells

    International Nuclear Information System (INIS)

    Li, Jin; Li, Xiaojin; Yu, Shuchun; Hao, Jinkai; Lu, Wangting; Shao, Zhigang; Yi, Baolian


    Highlights: • Porous polybenzimidazole membrane was prepared with glucose as porogen. • Phosphoric acid content was as high as 15.7 mol H 3 PO 4 per PBI repeat unit. • 200 h Constant current density test was carried out at 150 °C. • Degradation was due to the gap between membrane and catalyst layer. - Abstract: In this paper, the preparation and characterization of porous polybenzimidazole membranes doped with phosphoric acid were reported. For the preparation of porous polybenzimidazole membranes, glucose and saccharose were selected as porogen and added into PBI resin solution before solvent casting. The prepared porous PBI membranes had high proton conductivity and high content of acid doping at room temperature with 15.7 mol H 3 PO 4 per PBI repeat unit, much higher than pure PBI membrane at the same condition. Further, the performance and stability of the porous PBI membrane in high-temperature proton-exchange-membrane fuel cells was tested. It was found that the cell performance remained stable during 200 h stability test under a constant current discharge of 0.5 A cm −2 except for the last fifty hours. The decay in the last fifty hours was ascribed to the delamination between the catalyst layer and membrane increasing the charge-transfer resistance

  1. Cyclic guanosine monophosphate in the regulation of the cell function

    Directory of Open Access Journals (Sweden)

    Małgorzata Zbrojkiewicz


    Full Text Available Intracellular concentration of cGMP depends on the activity of guanylate cyclase, responsible for its synthesis, on the activity of cyclic nucleotide degrading enzymes - phosphodiesterases (PDEs. There are two forms of guanylate cyclase: the membrane-bound cyclase and the soluble form. The physiological activators of the membrane guanylate cyclase are natriuretic peptides (NPs, and of the cytosolic guanylate cyclase - nitric oxide (NO and carbon monoxide (CO. Intracellular cGMP signaling pathways arise from its direct effect on the activity of G protein kinases, phosphodiesterases and cyclic nucleotide dependent cation channels. It has been shown in recent years that cGMP can also affect other signal pathways in cell signaling activity involving Wnt proteins and sex hormones. The increased interest in the research on the role of cGMP, resulted also in the discovery of its role in the regulation of phototransduction in the eye, neurotransmission, calcium homeostasis, platelet aggregation, heartbeat, bone remodeling, lipid metabolism and the activity of the cation channels. Better understanding of the mechanisms of action of cGMP in the regulation of cell function can create new opportunities for the cGMP affecting drugs use in the pharmacotherapy.

  2. MARCKS-related protein regulates cytoskeletal organization at cell-cell and cell-substrate contacts in epithelial cells. (United States)

    Van Itallie, Christina M; Tietgens, Amber Jean; Aponte, Angel; Gucek, Marjan; Cartagena-Rivera, Alexander X; Chadwick, Richard S; Anderson, James M


    Treatment of epithelial cells with interferon-γ and TNF-α (IFN/TNF) results in increased paracellular permeability. To identify relevant proteins mediating barrier disruption, we performed proximity-dependent biotinylation (BioID) of occludin and found that tagging of MARCKS-related protein (MRP; also known as MARCKSL1) increased ∼20-fold following IFN/TNF administration. GFP-MRP was focused at the lateral cell membrane and its overexpression potentiated the physiological response of the tight junction barrier to cytokines. However, deletion of MRP did not abrogate the cytokine responses, suggesting that MRP is not required in the occludin-dependent IFN/TNF response. Instead, our results reveal a key role for MRP in epithelial cells in control of multiple actin-based structures, likely by regulation of integrin signaling. Changes in focal adhesion organization and basal actin stress fibers in MRP-knockout (KO) cells were reminiscent of those seen in FAK-KO cells. In addition, we found alterations in cell-cell interactions in MRP-KO cells associated with increased junctional tension, suggesting that MRP may play a role in focal adhesion-adherens junction cross talk. Together, our results are consistent with a key role for MRP in cytoskeletal organization of cell contacts in epithelial cells. © 2018. Published by The Company of Biologists Ltd.

  3. Determine equilibrium dissociation constant of drug-membrane receptor affinity using the cell membrane chromatography relative standard method. (United States)

    Ma, Weina; Yang, Liu; Lv, Yanni; Fu, Jia; Zhang, Yanmin; He, Langchong


    The equilibrium dissociation constant (K D ) of drug-membrane receptor affinity is the basic parameter that reflects the strength of interaction. The cell membrane chromatography (CMC) method is an effective technique to study the characteristics of drug-membrane receptor affinity. In this study, the K D value of CMC relative standard method for the determination of drug-membrane receptor affinity was established to analyze the relative K D values of drugs binding to the membrane receptors (Epidermal growth factor receptor and angiotensin II receptor). The K D values obtained by the CMC relative standard method had a strong correlation with those obtained by the frontal analysis method. Additionally, the K D values obtained by CMC relative standard method correlated with pharmacological activity of the drug being evaluated. The CMC relative standard method is a convenient and effective method to evaluate drug-membrane receptor affinity. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Effects of cholesterol depletion on membrane nanostructure in MCF-7 cells by atomic force microscopy (United States)

    Wang, Yuhua; Jiang, Ningcheng; Shi, Aisi; Zheng, Liqin; Yang, Hongqin; Xie, Shusen


    The cell membrane is composed of phospholipids, glycolipids, cholesterol and proteins that are dynamic and heterogeneous distributed in the bilayer structure and many researches have showed that the plasma membrane in eukaryotic cells contains microdomains termed "lipid raft" in which cholesterol, sphingolipids and specific membrane proteins are enriched. Cholesterol extraction induced lipid raft disruption is one of the most widely used methods for lipid raft research and MβCD is a type of solvent to extract the cholesterol from cell membranes. In this study, the effect of MβCD treatment on the membrane nanostructure in MCF-7 living cells was investigated by atomic force microscopy. Different concentrations of MβCD were selected to deplete cholesterol for 30 min and the viability of cells was tested by MTT assay to obtain the optimal concentration. Then the nanostructure of the cell membrane was detected. The results show that an appropriate concentration of MβCD can induce the alteration of cell membranes nanostructure and the roughness of membrane surface decreases significantly. This may indicate that microdomains of the cell membrane disappear and the cell membrane appears more smoothly. Cholesterol can affect nanostructure and inhomogeneity of the plasma membrane in living cells.

  5. Nanocomposite membranes based on polybenzimidazole and ZrO2 for high-temperature proton exchange membrane fuel cells. (United States)

    Nawn, Graeme; Pace, Giuseppe; Lavina, Sandra; Vezzù, Keti; Negro, Enrico; Bertasi, Federico; Polizzi, Stefano; Di Noto, Vito


    Owing to the numerous benefits obtained when operating proton exchange membrane fuel cells at elevated temperature (>100 °C), the development of thermally stable proton exchange membranes that demonstrate conductivity under anhydrous conditions remains a significant goal for fuel cell technology. This paper presents composite membranes consisting of poly[2,2'-(m-phenylene)-5,5'-bibenzimidazole] (PBI4N) impregnated with a ZrO2 nanofiller of varying content (ranging from 0 to 22 wt %). The structure-property relationships of the acid-doped and undoped composite membranes have been studied using thermogravimetric analysis, differential scanning calorimetry, dynamic mechanical analysis, wide-angle X-ray scattering, infrared spectroscopy, and broadband electrical spectroscopy. Results indicate that the level of nanofiller has a significant effect on the membrane properties. From 0 to 8 wt %, the acid uptake as well as the thermal and mechanical properties of the membrane increase. As the nanofiller level is increased from 8 to 22 wt % the opposite effect is observed. At 185 °C, the ionic conductivity of [PBI4N(ZrO2 )0.231 ](H3 PO4 )13 is found to be 1.04×10(-1)  S cm(-1) . This renders membranes of this type promising candidates for use in high-temperature proton exchange membrane fuel cells. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Influence of estrogenic pesticides on membrane integrity and membrane transfer of monosaccharide into the human red cell

    International Nuclear Information System (INIS)

    Ingermann, R.L.


    Some natural and synthetic estrogens inhibit carrier-mediated transport of glucose into human red blood cells and membrane vesicles from the placenta. The inhibitory action of these estrogens on transport appears to be a direct effect at the membrane and does not involve receptor binding and protein synthesis. It is not clear, however, whether such inhibition is a common feature among estrogenic agents. Several chlorinated hydrocarbon pesticides have been shown to possess estrogenic activity. These pesticides could have inhibitory effects on the human sodium-independent glucose transporter. Owing to the apparent importance of this membrane transporter in human tissues, direct interaction of hormones and xenobiotics with the glucose transporter is of fundamental significance. Some pesticides have been shown to alter membrane structure directly and alter the passive permeability of membranes. Whether the estrogenic pesticides influence passive diffusion of sugars across membranes has not been established. Finally, preliminary observations have suggested that some estrogens and pesticides have lytic effects on intact cells. Consequently, this study focuses on the ability of several estrogens and estrogenic pesticides to disrupt the cell membrane, influence the monosaccharide transporter, and alter the rate of monosaccharide permeation through the membrane by simple diffusion

  7. Redox regulation of plant stem cell fate. (United States)

    Zeng, Jian; Dong, Zhicheng; Wu, Haijun; Tian, Zhaoxia; Zhao, Zhong


    Despite the importance of stem cells in plant and animal development, the common mechanisms of stem cell maintenance in both systems have remained elusive. Recently, the importance of hydrogen peroxide (H 2 O 2 ) signaling in priming stem cell differentiation has been extensively studied in animals. Here, we show that different forms of reactive oxygen species (ROS) have antagonistic roles in plant stem cell regulation, which were established by distinct spatiotemporal patterns of ROS-metabolizing enzymes. The superoxide anion (O2·-) is markedly enriched in stem cells to activate WUSCHEL and maintain stemness, whereas H 2 O 2 is more abundant in the differentiating peripheral zone to promote stem cell differentiation. Moreover, H 2 O 2 negatively regulates O2·- biosynthesis in stem cells, and increasing H 2 O 2 levels or scavenging O2·- leads to the termination of stem cells. Our results provide a mechanistic framework for ROS-mediated control of plant stem cell fate and demonstrate that the balance between O2·- and H 2 O 2 is key to stem cell maintenance and differentiation. © 2017 The Authors.

  8. Tip cells: master regulators of tubulogenesis? (United States)

    Weavers, Helen; Skaer, Helen


    The normal development of an organ depends on the coordinated regulation of multiple cell activities. Focusing on tubulogenesis, we review the role of specialised cells or groups of cells that are selected from within tissue primordia and differentiate at the outgrowing tips or leading edge of developing tubules. Tip or leading cells develop distinctive patterns of gene expression that enable them to act both as sensors and transmitters of intercellular signalling. This enables them to explore the environment, respond to both tissue intrinsic signals and extrinsic cues from surrounding tissues and to regulate the behaviour of their neighbours, including the setting of cell fate, patterning cell division, inducing polarity and promoting cell movement and cell rearrangements by neighbour exchange. Tip cells are also able to transmit mechanical tension to promote tissue remodelling and, by interacting with the extracellular matrix, they can dictate migratory pathways and organ shape. Where separate tubular structures fuse to form networks, as in the airways of insects or the vascular system of vertebrates, specialised fusion tip cells act to interconnect disparate elements of the developing network. Finally, we consider their importance in the maturation of mature physiological function and in the development of disease. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  9. Thyroid Hormones as Renal Cell Cancer Regulators


    Szyma?ski, ?ukasz; Matak, Damian; Bartnik, Ewa; Szczylik, Cezary; Czarnecka, Anna M.


    It is known that thyroid hormone is an important regulator of cancer development and metastasis. What is more, changes across the genome, as well as alternative splicing, may affect the activity of the thyroid hormone receptors. Mechanism of action of the thyroid hormone is different in every cancer; therefore in this review thyroid hormone and its receptor are presented as a regulator of renal cell carcinoma.

  10. Rho kinase regulates fragmentation and phagocytosis of apoptotic cells

    International Nuclear Information System (INIS)

    Orlando, Kelly A.; Stone, Nicole L.; Pittman, Randall N.


    During the execution phase of apoptosis, a cell undergoes cytoplasmic and nuclear changes that prepare it for death and phagocytosis. The end-point of the execution phase is condensation into a single apoptotic body or fragmentation into multiple apoptotic bodies. Fragmentation is thought to facilitate phagocytosis; however, mechanisms regulating fragmentation are unknown. An isoform of Rho kinase, ROCK-I, drives membrane blebbing through its activation of actin-myosin contraction; this raises the possibility that ROCK-I may regulate other execution phase events, such as cellular fragmentation. Here, we show that COS-7 cells fragment into a number of small apoptotic bodies during apoptosis; treating with ROCK inhibitors (Y-27632 or H-1152) prevents fragmentation. Latrunculin B and blebbistatin, drugs that interfere with actin-myosin contraction, also inhibit fragmentation. During apoptosis, ROCK-I is cleaved and activated by caspases, while ROCK-II is not activated, but rather translocates to a cytoskeletal fraction. siRNA knock-down of ROCK-I but not ROCK-II inhibits fragmentation of dying cells, consistent with ROCK-I being required for apoptotic fragmentation. Finally, cells dying in the presence of the ROCK inhibitor Y-27632 are not efficiently phagocytized. These data show that ROCK plays an essential role in fragmentation and phagocytosis of apoptotic cells

  11. Triggering of Erythrocyte Cell Membrane Scrambling by Emodin

    Directory of Open Access Journals (Sweden)

    Morena Mischitelli


    Full Text Available Background/Aims: The natural anthraquinone derivative emodin (1,3,8-trihydroxy-6-methylanthraquinone is a component of several Chinese medicinal herbal preparations utilized for more than 2000 years. The substance has been used against diverse disorders including malignancy, inflammation and microbial infection. The substance is effective in part by triggering suicidal death or apoptosis. Similar to apoptosis of nucleated cells erythrocytes may enter suicidal erythrocyte death or eryptosis, characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Signaling involved in the triggering of eryptosis include increase of cytosolic Ca2+ activity ([Ca2+]i, oxidative stress and ceramide. The present study aimed to test, whether emodin induces eryptosis and, if so, to elucidate underlying cellular mechanisms. Methods: Phosphatidylserine abundance at the cell surface was estimated from annexin-V-binding, cell volume from forward scatter, [Ca2+]i from Fluo3-fluorescence, ROS formation from DCFDA dependent fluorescence, and ceramide abundance utilizing specific antibodies. Results: Exposure of human erythrocytes for 48 hours to emodin (≥ 10 µM significantly increased the percentage of annexin-V-binding cells, and at higher concentrations (≥ 50 µM significantly increased forward scatter. Emodin significantly increased Fluo3-fluorescence (≥ 10 µM, DCFDA fluorescence (75 µM and ceramide abundance (75 µM. The effect of emodin on annexin-V-binding was significantly blunted but not abolished by removal of extracellular Ca2+. Conclusions: Emodin triggers phospholipid scrambling of the erythrocyte cell membrane, an effect at least in part due to stimulation of Ca2+ entry and paralleled by oxidative stress and ceramide appearance at the erythroctye surface.

  12. Bleb Expansion in Migrating Cells Depends on Supply of Membrane from Cell Surface Invaginations. (United States)

    Goudarzi, Mohammad; Tarbashevich, Katsiaryna; Mildner, Karina; Begemann, Isabell; Garcia, Jamie; Paksa, Azadeh; Reichman-Fried, Michal; Mahabaleshwar, Harsha; Blaser, Heiko; Hartwig, Johannes; Zeuschner, Dagmar; Galic, Milos; Bagnat, Michel; Betz, Timo; Raz, Erez


    Cell migration is essential for morphogenesis, organ formation, and homeostasis, with relevance for clinical conditions. The migration of primordial germ cells (PGCs) is a useful model for studying this process in the context of the developing embryo. Zebrafish PGC migration depends on the formation of cellular protrusions in form of blebs, a type of protrusion found in various cell types. Here we report on the mechanisms allowing the inflation of the membrane during bleb formation. We show that the rapid expansion of the protrusion depends on membrane invaginations that are localized preferentially at the cell front. The formation of these invaginations requires the function of Cdc42, and their unfolding allows bleb inflation and dynamic cell-shape changes performed by migrating cells. Inhibiting the formation and release of the invaginations strongly interfered with bleb formation, cell motility, and the ability of the cells to reach their target. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. Process for recycling components of a PEM fuel cell membrane electrode assembly (United States)

    Shore, Lawrence [Edison, NJ


    The membrane electrode assembly (MEA) of a PEM fuel cell can be recycled by contacting the MEA with a lower alkyl alcohol solvent which separates the membrane from the anode and cathode layers of the assembly. The resulting solution containing both the polymer membrane and supported noble metal catalysts can be heated under mild conditions to disperse the polymer membrane as particles and the supported noble metal catalysts and polymer membrane particles separated by known filtration means.

  14. High-Efficiency Artificial Photosynthesis Using a Novel Alkaline Membrane Cell (United States)

    Narayan, Sri; Haines, Brennan; Blosiu, Julian; Marzwell, Neville


    A new cell designed to mimic the photosynthetic processes of plants to convert carbon dioxide into carbonaceous products and oxygen at high efficiency, has an improved configuration using a polymer membrane electrolyte and an alkaline medium. This increases efficiency of the artificial photosynthetic process, achieves high conversion rates, permits the use of inexpensive catalysts, and widens the range of products generated by this type of process. The alkaline membrane electrolyte allows for the continuous generation of sodium formate without the need for any additional separation system. The electrolyte type, pH, electrocatalyst type, and cell voltage were found to have a strong effect on the efficiency of conversion of carbon dioxide to formate. Indium electrodes were found to have higher conversion efficiency compared to lead. Bicarbonate electrolyte offers higher conversion efficiency and higher rates than water solutions saturated with carbon dioxide. pH values between 8 and 9 lead to the maximum values of efficiency. The operating cell voltage of 2.5 V, or higher, ensures conversion of the carbon dioxide to formate, although the hydrogen evolution reaction begins to compete strongly with the formate production reaction at higher cell voltages. Formate is produced at indium and lead electrodes at a conversion efficiency of 48 mg of CO2/kilojoule of energy input. This efficiency is about eight times that of natural photosynthesis in green plants. The electrochemical method of artificial photosynthesis is a promising approach for the conversion, separation and sequestration of carbon dioxide for confined environments as in space habitats, and also for carbon dioxide management in the terrestrial context. The heart of the reactor is a membrane cell fabricated from an alkaline polymer electrolyte membrane and catalyst- coated electrodes. This cell is assembled and held in compression in gold-plated hardware. The cathode side of the cell is supplied with carbon

  15. Comparison of gas membrane separation cascades using conventional separation cell and two-unit separation cells

    International Nuclear Information System (INIS)

    Ohno, Masayoshi; Morisue, Tetsuo; Ozaki, Osamu; Miyauchi, Terukatsu.


    The adoption of two-unit separation cells in radioactive rare gas membrane separation equipment enhances the separation factor, but increases the required membrane area and compressive power. An analytical economic evaluation was undertaken to compare the conventional separation cell with the two-unit separation cells, adopting as parameters the number of cascade stages, the membrane area and the operating power requirements. This paper describes the models used for evaluating the separation performance and the economics of cascade embodying these different concepts of separation cell taken up for study, and the results obtained for the individual concepts are mutually compared. It proved that, in respect of the number required of cascade stages, of operating power requirements and of the annual expenditure, better performance could always be expected of the two-unit separation cells as compared with the conventional separation cell, at least in the range of parameters adopted in this study. As regards the minimum membrane area, the conventional separation cell and the series-type separation cell yielded almost the same values, with the parallel-type separation cell falling somewhat behind. (auth.)

  16. Cell dualism: presence of cells with alternative membrane potentials in growing populations of bacteria and yeasts. (United States)

    Ivanov, Volodymyr; Rezaeinejad, Saeid; Chu, Jian


    It is considered that all growing cells, for exception of acidophilic bacteria, have negatively charged inside cytoplasmic membrane (Δψ⁻-cells). Here we show that growing populations of microbial cells contain a small portion of cells with positively charged inside cytoplasmic membrane (Δψ⁺-cells). These cells were detected after simultaneous application of the fluorescent probes for positive membrane potential (anionic dye DIBAC⁻) and membrane integrity (propidium iodide, PI). We found in exponentially growing cell populations of Escherichia coli and Saccharomyces cerevisiae that the content of live Δψ⁻-cells was 93.6 ± 1.8 % for bacteria and 90.4 ± 4.0 % for yeasts and the content of live Δψ⁺-cells was 0.9 ± 0.3 % for bacteria and 2.4 ± 0.7 % for yeasts. Hypothetically, existence of Δψ⁺-cells could be due to short-term, about 1 min for bacteria and 5 min for yeasts, change of membrane potential from negative to positive value during the cell cycle. This change has been shown by the reversions of K⁺, Na⁺, and Ca²⁺ ions fluxes across the cell membrane during synchronous yeast culture. The transformation of Δψ(⁻-cells to Δψ⁺-cells can be explained by slow influx of K⁺ ions into Δψ⁻-cell to the trigger level of K⁺ concentration ("compression of potassium spring"), which is forming "alternative" Δψ⁺-cell for a short period, following with fast efflux of K⁺ ions out of Δψ⁺-cell ("release of potassium spring") returning cell to normal Δψ⁻ state. We anticipate our results to be a starting point to reveal the biological role of cell dualism in form of Δψ⁻- and Δψ⁺- cells.

  17. Regulation of Murine Natural Killer Cell Commitment

    Directory of Open Access Journals (Sweden)

    Nicholas D Huntington


    Full Text Available NK cells can derive from the same precursors as B and T cells, however to achieve lineage specificity, several transcription factors need to be activated or annulled. While a few important transcription factors have identified for NK genesis the mechanisms of how this is achieved is far from resolved. Adding to the complexity of this, NK cells are found and potentially develop in diverse locations in vivo and it remains to be addressed if a common NK cell precursor seeds diverse niches and how transcription factors may differentially regulate NK cell commitment in distinct microenvironments. Here we will summarise some recent findings in NK cell commitment and discuss how a NK cell transcriptional network might be organised, while addressing some misconceptions and anomalies along the way.

  18. Nuclear and Membrane Actions of Estrogen Receptor Alpha: Contribution to the Regulation of Energy and Glucose Homeostasis. (United States)

    Guillaume, Maeva; Montagner, Alexandra; Fontaine, Coralie; Lenfant, Françoise; Arnal, Jean-François; Gourdy, Pierre


    Estrogen receptor alpha (ERα) has been demonstrated to play a key role in reproduction but also to exert numerous functions in nonreproductive tissues. Accordingly, ERα is now recognized as a key regulator of energy homeostasis and glucose metabolism and mediates the protective effects of estrogens against obesity and type 2 diabetes. This chapter attempts to summarize our current understanding of the mechanisms of ERα activation and their involvement in the modulation of energy balance and glucose metabolism. We first focus on the experimental studies that constitute the basis of the understanding of ERα as a nuclear receptor and more specifically on the key roles played by its two activation functions (AFs). We depict the consequences of the selective inactivation of these AFs in mouse models, which further underline the prominent role of nuclear ERα in the prevention of obesity and diabetes, as on the reproductive tract and the vascular system. Besides these nuclear actions, a fraction of ERα is associated with the plasma membrane and activates nonnuclear signaling from this site. Such rapid effects, called membrane-initiated steroid signals (MISS), have been characterized in a variety of cell lines and in particular in endothelial cells. The development of selective pharmacological tools that specifically activate MISS as well as the generation of mice expressing an ERα protein impeded for membrane localization has just begun to unravel the physiological role of MISS in vivo and their contribution to ERα-mediated metabolic protection. Finally, we discuss novel perspectives for the design of tissue-selective ER modulators.

  19. New insights into the regulation of cholesterol efflux from the sperm membrane

    Directory of Open Access Journals (Sweden)

    Tamara Leahy


    Full Text Available Cholesterol is an essential component of the mammalian plasma membrane because it promotes membrane stability without comprising membrane fluidity. Given this important cellular role, cholesterol levels are tightly controlled at multiple levels. It has been clearly shown that cholesterol redistribution and depletion from the sperm membrane is a key part of the spermatozoon′s preparation for fertilization. Some factors that regulate these events are described (e.g., bicarbonate, calcium but the mechanisms underlying cholesterol export are poorly understood. How does a hydrophobic cholesterol molecule inserted in the sperm plasma membrane enter the energetically unfavorable aqueous surroundings? This review will provide an overview of knowledge in this area and highlight our gaps in understanding. The overall aim is to better understand cholesterol redistribution in the sperm plasma membrane, its relation to the possible activation of a cholesterol transporter and the role of cholesterol acceptors. Armed with such knowledge, sperm handling techniques can be adapted to better prepare spermatozoa for in vitro and in vivo fertilization.

  20. Regulation of PutA-membrane associations by flavin adenine dinucleotide reduction. (United States)

    Zhang, Weimin; Zhou, Yuzhen; Becker, Donald F


    Proline utilization A (PutA) from Escherichia coli is a multifunctional flavoprotein that is both a transcriptional repressor of the proline utilization (put) genes and a membrane-associated enzyme which catalyzes the 4-electron oxidation of proline to glutamate. Previously, proline was shown to induce PutA-membrane binding and alter the intracellular location and function of PutA. To distinguish the roles of substrate binding and FAD reduction in the mechanism of how PutA changes from a DNA-binding protein to a membrane-bound enzyme, the kinetic parameters of PutA-membrane binding were measured under different conditions using model lipid bilayers and surface plasmon resonance (SPR). The effects of proline, FAD reduction, and proline analogues on PutA-membrane associations were determined. Oxidized PutA shows no binding to E. coli polar lipid vesicles. In contrast, proline and sodium dithionite induce tight binding of PutA to the lipid bilayer with indistinguishable kinetic parameters and an estimated dissociation constant (K(D)) of PutA-lipid complex. Proline analogues such as L-THFA and DL-P5C also stimulate PutA binding to E. coli polar lipid vesicles with K(D) values ranging from approximately 3.6 to 34 nM (pH 7.4) for the PutA-lipid complex. The greater PutA-membrane binding affinity (>300-fold) generated by FAD reduction relative to the nonreducing ligands demonstrates that FAD reduction controls PutA-membrane associations. On the basis of SPR kinetic analysis with differently charged lipid bilayers, the driving force for PutA-membrane binding is primarily hydrophobic. In the SPR experiments membrane-bound PutA did not bind put control DNA, confirming that the membrane-binding and DNA-binding activities of PutA are mutually exclusive. A model for the regulation of PutA is described in which the overall translocation of PutA from the cytoplasm to the membrane is driven by FAD reduction and the subsequent energy difference ( approximately 24 kJ/mol) between

  1. Noncontact microsurgery of cell membranes using femtosecond laser pulses for optoinjection of specified substances into cells (United States)

    Il'ina, I. V.; Ovchinnikov, A. V.; Chefonov, O. V.; Sitnikov, D. S.; Agranat, Mikhail B.; Mikaelyan, A. S.


    IR femtosecond laser pulses were used for microsurgery of a cell membrane aimed at local and short-duration change in its permeability and injection of specified extracellular substances into the cells. The possibility of noncontact laser delivery of the propidium iodide fluorescent dye and the pEGFP plasmid, encoding the green fluorescent protein, into the cells with preservation of the cell viability was demonstrated.

  2. Comparative Studies of Polymer Electrolyte Membrane Fuel Cell Stacks and Single Cells (United States)


    in the Catalyst Layer and Effects of Both Perfluorosulfonate Ionomer and PTFE-Loaded Carbon on the Catalyst Layer of Polymer Electrolyte Fuel Cells support of this project. 12 References 1. T. F. Fuller, "Is a Fuel Cell in Your Future?" 77K Electrochemical Society Interface (Fall...ARMY RESEARCH LABORATORY mm^ n Comparative Studies of Polymer Electrolyte Membrane Fuel Cell Stacks and Single Cells Deryn Chu and Rongzhong

  3. Synthesis and characterisation of alkaline anionic-exchange membranes for direct alcohol fuel cells

    CSIR Research Space (South Africa)

    Nonjola, P


    Full Text Available , but the most important being proton exchange membrane fuel cell (PEMFC), which uses an acidic membrane like Nafion (sulfonated fluorocarbon polymers) as an electrolyte. The use of polymer electrolytes represents an interesting path to pursue...

  4. The structure and function of cell membranes studied by atomic force microscopy. (United States)

    Shi, Yan; Cai, Mingjun; Zhou, Lulu; Wang, Hongda


    The cell membrane, involved in almost all communications of cells and surrounding matrix, is one of the most complicated components of cells. Lack of suitable methods for the detection of cell membranes in vivo has sparked debates on the biochemical composition and structure of cell membranes over half a century. The development of single molecule techniques, such as AFM, SMFS, and TREC, provides a versatile platform for imaging and manipulating cell membranes in biological relevant environments. Here, we discuss the latest developments in AFM and the progress made in cell membrane research. In particular, we highlight novel structure models and dynamic processes, including the mechanical properties of the cell membranes. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Transient response of a proton exchange membrane fuel cell (United States)

    Weydahl, Helge; Møller-Holst, Steffen; Hagen, Georg; Børresen, Børre

    The transient response of a proton exchange membrane fuel cell (PEMFC) supplied with pure hydrogen and oxygen was investigated by load step measurements assisted by electrochemical impedance spectroscopy and chronoamperometry. Using an in-house designed resistance board, the uncontrolled response in both cell voltage and current upon step changes in a resistive load was observed. The PEMFC was found to respond quickly and reproducibly to load changes. The transient PEMFC response was limited by a cathodic charge transfer process with a potential-dependent response time. For load steps to high-current densitities, a second transient process with a constant response time was observed. This transient was offset from the charge transfer transient by a temporarily stable plateau. Results from chronoamperometry indicated that the second transient could be related to a diffusion process. Transient paths were plotted in the V- i diagram, matching a predicted pattern with overshooting cell voltage and current during a load step.

  6. High performance direct methanol fuel cell with thin electrolyte membrane (United States)

    Wan, Nianfang


    A high performance direct methanol fuel cell is achieved with thin electrolyte membrane. 320 mW cm-2 of peak power density and over 260 mW cm-2 at 0.4 V are obtained when working at 90 °C with normal pressure air supply. It is revealed that the increased anode half-cell performance with temperature contributes primarily to the enhanced performance at elevated temperature. From the comparison of iR-compensated cathode potential of methanol/air with that of H2/air fuel cell, the impact of methanol crossover on cathode performance decreases with current density and becomes negligible at high current density. Current density is found to influence fuel efficiency and methanol crossover significantly from the measurement of fuel efficiency at different current density. At high current density, high fuel efficiency can be achieved even at high temperature, indicating decreased methanol crossover.

  7. Calcium pumps of plasma membrane and cell interior

    DEFF Research Database (Denmark)

    Strehler, Emanuel E; Treiman, Marek


    Calcium entering the cell from the outside or from intracellular organelles eventually must be returned to the extracellular milieu or to intracellular storage organelles. The two major systems capable of pumping Ca2+ against its large concentration gradient out of the cell or into the sarco....../endoplasmatic reticulum are the plasma membrane Ca2+ ATPases (PMCAs) and the sarco/endoplasmic reticulum Ca2+ ATPases (SERCAs), respectively. In mammals, multigene families code for these Ca2+ pumps and additional isoform subtypes are generated via alternative splicing. PMCA and SERCA isoforms show developmental-, tissue......- and cell type-specific patterns of expression. Different PMCA and SERCA isoforms are characterized by different regulatory and kinetic properties that likely are optimized for the distinct functional tasks fulfilled by each pump in setting resting cytosolic or intra-organellar Ca2+ levels, and in shaping...

  8. Percolation in a Proton Exchange Membrane Fuel Cell Catalyst Layer

    Energy Technology Data Exchange (ETDEWEB)

    Stacy, Stephen; Allen, Jeffrey


    Water management in the catalyst layers of proton exchange membrane fuel cells (PEMFC) is confronted by two issues, flooding and dry out, both of which result in improper functioning of the fuel cell and lead to poor performance and degradation. At the present time, the data that has been reported about water percolation and wettability within a fuel cell catalyst layer is limited. A method and apparatus for measuring the percolation pressure in the catalyst layer has been developed based upon an experimental apparatus used to test water percolation in porous transport layers (PTL). The experimental setup uses a pseudo Hele-Shaw type testing where samples are compressed and a fluid is injected into the sample. Testing the samples gives percolation pressure plots which show trends in increasing percolation pressure with an increase in flow rate. A decrease in pressure was seen as percolation occurred in one sample, however the pressure only had a rising effect in the other sample.

  9. A Simple Alkaline Method for Decellularizing Human Amniotic Membrane for Cell Culture (United States)

    Saghizadeh, Mehrnoosh; Winkler, Michael A.; Kramerov, Andrei A.; Hemmati, David M.; Ghiam, Chantelle A.; Dimitrijevich, Slobodan D.; Sareen, Dhruv; Ornelas, Loren; Ghiasi, Homayon; Brunken, William J.; Maguen, Ezra; Rabinowitz, Yaron S.; Svendsen, Clive N.; Jirsova, Katerina; Ljubimov, Alexander V.


    Human amniotic membrane is a standard substratum used to culture limbal epithelial stem cells for transplantation to patients with limbal stem cell deficiency. Various methods were developed to decellularize amniotic membrane, because denuded membrane is poorly immunogenic and better supports repopulation by dissociated limbal epithelial cells. Amniotic membrane denuding usually involves treatment with EDTA and/or proteolytic enzymes; in many cases additional mechanical scraping is required. Although ensuring limbal cell proliferation, these methods are not standardized, require relatively long treatment times and can result in membrane damage. We propose to use 0.5 M NaOH to reliably remove amniotic cells from the membrane. This method was used before to lyse cells for DNA isolation and radioactivity counting. Gently rubbing a cotton swab soaked in NaOH over the epithelial side of amniotic membrane leads to nearly complete and easy removal of adherent cells in less than a minute. The denuded membrane is subsequently washed in a neutral buffer. Cell removal was more thorough and uniform than with EDTA, or EDTA plus mechanical scraping with an electric toothbrush, or n-heptanol plus EDTA treatment. NaOH-denuded amniotic membrane did not show any perforations compared with mechanical or thermolysin denuding, and showed excellent preservation of immunoreactivity for major basement membrane components including laminin α2, γ1-γ3 chains, α1/α2 and α6 type IV collagen chains, fibronectin, nidogen-2, and perlecan. Sodium hydroxide treatment was efficient with fresh or cryopreserved (10% dimethyl sulfoxide or 50% glycerol) amniotic membrane. The latter method is a common way of membrane storage for subsequent grafting in the European Union. NaOH-denuded amniotic membrane supported growth of human limbal epithelial cells, immortalized corneal epithelial cells, and induced pluripotent stem cells. This simple, fast and reliable method can be used to standardize

  10. A simple alkaline method for decellularizing human amniotic membrane for cell culture.

    Directory of Open Access Journals (Sweden)

    Mehrnoosh Saghizadeh

    Full Text Available Human amniotic membrane is a standard substratum used to culture limbal epithelial stem cells for transplantation to patients with limbal stem cell deficiency. Various methods were developed to decellularize amniotic membrane, because denuded membrane is poorly immunogenic and better supports repopulation by dissociated limbal epithelial cells. Amniotic membrane denuding usually involves treatment with EDTA and/or proteolytic enzymes; in many cases additional mechanical scraping is required. Although ensuring limbal cell proliferation, these methods are not standardized, require relatively long treatment times and can result in membrane damage. We propose to use 0.5 M NaOH to reliably remove amniotic cells from the membrane. This method was used before to lyse cells for DNA isolation and radioactivity counting. Gently rubbing a cotton swab soaked in NaOH over the epithelial side of amniotic membrane leads to nearly complete and easy removal of adherent cells in less than a minute. The denuded membrane is subsequently washed in a neutral buffer. Cell removal was more thorough and uniform than with EDTA, or EDTA plus mechanical scraping with an electric toothbrush, or n-heptanol plus EDTA treatment. NaOH-denuded amniotic membrane did not show any perforations compared with mechanical or thermolysin denuding, and showed excellent preservation of immunoreactivity for major basement membrane components including laminin α2, γ1-γ3 chains, α1/α2 and α6 type IV collagen chains, fibronectin, nidogen-2, and perlecan. Sodium hydroxide treatment was efficient with fresh or cryopreserved (10% dimethyl sulfoxide or 50% glycerol amniotic membrane. The latter method is a common way of membrane storage for subsequent grafting in the European Union. NaOH-denuded amniotic membrane supported growth of human limbal epithelial cells, immortalized corneal epithelial cells, and induced pluripotent stem cells. This simple, fast and reliable method can be used to

  11. Activation of the B cell receptor leads to increased membrane proximity of the Igα cytoplasmic domain.

    Directory of Open Access Journals (Sweden)

    Wing-Yiu Lee

    Full Text Available Binding of antigen to the B cell receptor (BCR induces conformational changes in BCR's cytoplasmic domains that are concomitant with phosphorylation of the immunoreceptor tyrosine-based activation motifs (ITAMs. Recently, reversible folding of the CD3ε and ξ chain ITAMs into the plasma membrane has been suggested to regulate T cell receptor signaling. Here we show that the Igα and Igβ cytoplasmic domains of the BCR do not associate with plasma membrane in resting B cells. However, antigen binding and ITAM phosphorylation specifically increased membrane proximity of Igα, but not Igβ. Thus, BCR activation is accompanied by asymmetric conformational changes, possibly promoting the binding of Igα and Igβ to differently localized signaling complexes.

  12. Phosphoric acid doped polybenzimidazole membranes: Physiochemical characterization and fuel cell applications [PEM fuel cells

    DEFF Research Database (Denmark)

    Qingfeng, Li; Hjuler, Hans Aage; Bjerrum, Niels


    A polymer electrolyte membrane fuel cell operational at temperatures around 150-200 degrees C is desirable for fast electrode kinetics and high tolerance to fuel impurities. For this purpose polybenzimidazole (PBI) membranes have been prepared and H/sub 3/PO/sub 4/-doped in a doping range from 300...... to 1600 mol%. Physiochemical properties of the membrane electrolyte have been investigated by measurements of water uptake, acid doping level, electric conductivity, mechanical strength and water drag coefficient. Electrical conductivity is found to be insensitive to humidity but dependent on the acid...... doping level. At 160 degrees C a conductivity as high as 0.13 S cm/sup -1/ is obtained for membranes of high doping levels. Mechanical strength measurements show, however, that a high acid doping level results in poor mechanical properties. At operational temperatures up to 190 degrees C, fuel cells...

  13. Plasma membrane protein trafficking in plant-microbe interactions: a plant cell point of view

    Directory of Open Access Journals (Sweden)

    Nathalie eLeborgne-Castel


    Full Text Available In order to ensure their physiological and cellular functions, plasma membrane (PM proteins must be properly conveyed from their site of synthesis, i.e. the endoplasmic reticulum, to their final destination, the PM, through the secretory pathway. PM protein homeostasis also relies on recycling and/or degradation, two processes that are initiated by endocytosis. Vesicular membrane trafficking events to and from the PM have been shown to be altered when plant cells are exposed to mutualistic or pathogenic microbes. In this review, we will describe the fine-tune regulation of such alterations, and their consequence in PM protein activity. We will consider the formation of intracellular perimicrobial compartments, the PM protein trafficking machinery of the host, and the delivery or retrieval of signaling and transport proteins such as pattern-recognition receptors, producers of reactive oxygen species, and sugar transporters.

  14. Phactr3/scapinin, a member of protein phosphatase 1 and actin regulator (phactr family, interacts with the plasma membrane via basic and hydrophobic residues in the N-terminus.

    Directory of Open Access Journals (Sweden)

    Akihiro Itoh

    Full Text Available Proteins that belong to the protein phosphatase 1 and actin regulator (phactr family are involved in cell motility and morphogenesis. However, the mechanisms that regulate the actin cytoskeleton are poorly understood. We have previously shown that phactr3, also known as scapinin, localizes to the plasma membrane, including lamellipodia and membrane ruffles. In the present study, experiments using deletion and point mutants showed that the basic and hydrophobic residues in the N-terminus play crucial roles in the localization to the plasma membrane. A BH analysis ( is a program developed to identify membrane-binding domains that comprise basic and hydrophobic residues in membrane proteins. We applied this program to phactr3. The results of the BH plot analysis agreed with the experimentally determined region that is responsible for the localization of phactr3 to the plasma membrane. In vitro experiments showed that the N-terminal itself binds to liposomes and acidic phospholipids. In addition, we showed that the interaction with the plasma membrane via the N-terminal membrane-binding sequence is required for phactr3-induced morphological changes in Cos7 cells. The membrane-binding sequence in the N-terminus is highly conserved in all members of the phactr family. Our findings may provide a molecular basis for understanding the mechanisms that allow phactr proteins to regulate cell morphogenesis.

  15. Alterations of red cell membrane properties in neuroacanthocytosis.

    Directory of Open Access Journals (Sweden)

    Claudia Siegl

    Full Text Available Neuroacanthocytosis (NA refers to a group of heterogenous, rare genetic disorders, namely chorea acanthocytosis (ChAc, McLeod syndrome (MLS, Huntington's disease-like 2 (HDL2 and pantothenate kinase associated neurodegeneration (PKAN, that mainly affect the basal ganglia and are associated with similar neurological symptoms. PKAN is also assigned to a group of rare neurodegenerative diseases, known as NBIA (neurodegeneration with brain iron accumulation, associated with iron accumulation in the basal ganglia and progressive movement disorder. Acanthocytosis, the occurrence of misshaped erythrocytes with thorny protrusions, is frequently observed in ChAc and MLS patients but less prevalent in PKAN (about 10% and HDL2 patients. The pathological factors that lead to the formation of the acanthocytic red blood cell shape are currently unknown. The aim of this study was to determine whether NA/NBIA acanthocytes differ in their functionality from normal erythrocytes. Several flow-cytometry-based assays were applied to test the physiological responses of the plasma membrane, namely drug-induced endocytosis, phosphatidylserine exposure and calcium uptake upon treatment with lysophosphatidic acid. ChAc red cell samples clearly showed a reduced response in drug-induced endovesiculation, lysophosphatidic acid-induced phosphatidylserine exposure, and calcium uptake. Impaired responses were also observed in acanthocyte-positive NBIA (PKAN red cells but not in patient cells without shape abnormalities. These data suggest an "acanthocytic state" of the red cell where alterations in functional and interdependent membrane properties arise together with an acanthocytic cell shape. Further elucidation of the aberrant molecular mechanisms that cause this acanthocytic state may possibly help to evaluate the pathological pathways leading to neurodegeneration.

  16. [Interaction of FABP4 with plasma membrane proteins of endothelial cells]. (United States)

    Saavedra, Paula; Girona, Josefa; Aragonès, Gemma; Cabré, Anna; Guaita, Sandra; Heras, Mercedes; Masana, Lluís


    Fatty acid binding protein (FABP4) is an adipose tissue-secreted adipokine implicated in the regulation of the energetic metabolism and inflammation. High levels of circulating FABP4 have been described in people with obesity, atherogenic dyslipidemia, diabetes and metabolic syndrome. Recent studies have demonstrated that FABP4 could have a direct effect on peripheral tissues and, specifically, on vascular function. It is still unknown how the interaction between FABP4 and the endothelial cells is produced to prompt these effects on vascular function. The objective of this work is studying the interaction between FABP4 and the plasma membrane proteins of endothelial cells. HUVEC cells were incubated with and without FABP4 (100 ng/ml) for 5 minutes. Immunolocalization of FABP4 was studied by confocal microscopy. The results showed that FABP4 colocalizates with CD31, a membrane protein marker. A strategy which combines 6XHistidine-tag FABP4 (FABP4-His), incubations with or without FABP4-His (100 ng/ml), formaldehyde cross-linking, cellular membrane protein extraction and western blot, was designed to study the FABP4 interactions with membrane proteins of HUVECs. The results showed different western blot profiles depending of the incubation with or without FABP4-His. The immunoblot revelead three covalent protein complexes of about 108, 77 and 33 kDa containing FAPB4 and its putative receptor. The existence of a specific binding protein complex able to bind FABP4 to endothelial cells is supported by these results. The obtained results will permit us advance in the molecular knowledge of FABP4 effects as well as use this protein and its receptor as therapeutic target to prevent cardiovascular. Copyright © 2014 Sociedad Española de Arteriosclerosis. Published by Elsevier España. All rights reserved.

  17. Tandem cathode for proton exchange membrane fuel cells

    DEFF Research Database (Denmark)

    Siahrostami, Samira; Björketun, Mårten E.; Strasser, Peter


    The efficiency of proton exchange membrane fuel cells is limited mainly by the oxygen reduction reaction at the cathode. The large cathodic overpotential is caused by correlations between binding energies of reaction intermediates in the reduction of oxygen to water. This work introduces a novel...... reaction intermediate each, and they occur on different catalyst surfaces. As a result they can be optimized independently and the fundamental problem associated with the four-electron catalysis is avoided. A combination of density functional theory calculations and published experimental data is used...

  18. 2011 Alkaline Membrane Fuel Cell Workshop Final Report

    Energy Technology Data Exchange (ETDEWEB)

    Pivovar, B.


    A workshop addressing the current state-of-the-art in alkaline membrane fuel cells (AMFCs) was held May 8-9, 2011, at the Crystal Gateway Marriott in Arlington, Virginia. This workshop was the second of its kind, with the first being held December 11-13, 2006, in Phoenix, Arizona. The 2011 workshop and associated workshop report were created to assess the current state of AMFC technology (taking into account recent advances), investigate the performance potential of AMFC systems across all possible power ranges and applications, and identify the key research needs for commercial competitiveness in a variety of areas.

  19. A highly-occupied, single-cell trapping microarray for determination of cell membrane permeability. (United States)

    Weng, Lindong; Ellett, Felix; Edd, Jon; Wong, Keith H K; Uygun, Korkut; Irimia, Daniel; Stott, Shannon L; Toner, Mehmet


    Semi- and selective permeability is a fundamentally important characteristic of the cell membrane. Membrane permeability can be determined by monitoring the volumetric change of cells following exposure to a non-isotonic environment. For this purpose, several microfluidic perfusion chambers have been developed recently. However, these devices only allow the observation of one single cell or a group of cells that may interact with one another in an uncontrolled way. Some of these devices have integrated on-chip temperature control to investigate the temperature-dependence of membrane permeability, but they inevitably require sophisticated fabrication and assembly, and delicate temperature and pressure calibration. Therefore, it is highly desirable to design a simple single-cell trapping device that allows parallel monitoring of multiple separate, individual cells subjected to non-isotonic exposure at various temperatures. In this study, we developed a pumpless, single-layer microarray with high trap occupancy of single cells. The benchmark performance of the device was conducted by targeting spherical particles of 18.8 μm in diameter as a model, yielding trap occupancy of up to 86.8% with a row-to-row shift of 10-30 μm. It was also revealed that in each array the particles larger than a corresponding critical size would be excluded by the traps in a deterministic lateral displacement mode. Demonstrating the utility of this approach, we used the single-cell trapping device to determine the membrane permeability of rat hepatocytes and patient-derived circulating tumor cells (Brx-142) at 4, 22 and 37 °C. The membrane of rat hepatocytes was found to be highly permeable to water and small molecules such as DMSO and glycerol, via both lipid- and aquaporin-mediated pathways. Brx-142 cells, however, displayed lower membrane permeability than rat hepatocytes, which was associated with strong coupling of water and DMSO transport but less interaction between water and

  20. Regulation of cell division in higher plants

    Energy Technology Data Exchange (ETDEWEB)

    Jacobs, T.W.


    Cell division is arguably the most fundamental of all developmental processes. In higher plants, mitotic activity is largely confined to foci of patterned cell divisions called meristems. From these perpetually embryonic tissues arise the plant's essential organs of light capture, support, protection and reproduction. Once an adequate understanding of plant cell mitotic regulation is attained, unprecedented opportunities will ensue for analyzing and genetically controlling diverse aspects of development, including plant architecture, leaf shape, plant height, and root depth. The mitotic cycle in a variety of model eukaryotic systems in under the control of a regulatory network of striking evolutionary conservation. Homologues of the yeast cdc2 gene, its catalytic product, p34, and the cyclin regulatory subunits of the MPF complex have emerged as ubiquitous mitotic regulators. We have cloned cdc2-like and cyclin genes from pea. As in other eukaryotic model systems, p34 of Pisum sativum is a subunit of a high molecular weight complex which binds the fission yeast p13 protein and displays histone H1 kinase activity in vitro. Our primary objective in this study is to gain baseline information about the regulation of this higher plant cell division control complex in non-dividing, differentiated cells as well as in synchronous and asynchronous mitotic cells. We are investigating cdc2 and cyclin expression at the levels of protein abundance, protein phosphorylation and quaternary associations.

  1. Gastrointestinal Hormone Cholecystokinin Increases P-Glycoprotein Membrane Localization and Transport Activity in Caco-2 Cells. (United States)

    Yano, Kentaro; Shimizu, Saori; Tomono, Takumi; Ogihara, Takuo


    It was reported that stimulation of taste receptor type 2 member 38 by a bitter substance, phenylthiocarbamide (PTC), increased P-glycoprotein (P-gp) mRNA level and transport activity via release of the gastrointestinal hormone cholecystokinin-8 (CCK-8) at 9 h. Therefore, we hypothesized that CCK-8 and PTC might also regulate P-gp activity more rapidly via a different mechanism. As a result, we found that the pretreatment of human colon adenocarcinoma (Caco-2) cells with 10-mM PTC significantly decreased the intracellular accumulation of P-gp substrate rhodamine 123 (Rho123) compared with the control after 90-min incubation. Moreover, CCK-8 treatments significantly reduced the accumulation of Rho123 within 30 min, compared with the control. On the other hand, when Caco-2 cells were pretreated with PTC, the efflux ratio of Rho123 was significantly increased compared with control. The efflux ratio of Rho123 in CCK-8 treatment cells was also significantly increased compared with control. Furthermore, CCK-8 increased the phosphorylation of the scaffold proteins ezrin, radixin, and moesin, which regulate translocation of P-gp to the plasma membrane. Therefore, our results indicate that PTC induced release of CCK-8, which in turn induced the phosphorylation of ezrin, radixin, and moesin proteins, leading to upregulation of P-gp transport activity via increased membrane localization of P-gp. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

  2. Aprediction study for the behaviour of fuel cell membrane subjected to hygro and thermal stresses in running PEM fuel cell


    Maher A.R. Sadiq Al-Baghdadi


    A three-dimensional, multi–phase, non-isothermal computational fluid dynamics model of a proton exchange membrane fuel cell has been used and developed to investigate the hygro and thermal stresses in polymer membrane, which developed during the cell operation due to the changes of temperature and relative humidity. The behaviour of the membrane during operation of a unit cell has been studied and investigated under real cell operating conditions. The results show that the non-uniform distrib...

  3. Toward mechanical manipulations of cell membranes and membrane proteins using an atomic force microscope: an invited review. (United States)

    Ikai, Atsushi; Afrin, Rehana


    Recent advances in the use of the atomic force microscope (AFM) for manipulating cell membranes and membrane proteins are reviewed. Early pioneering work on measurements of the magnitude of the force required to create indentations with defined depth on their surfaces and to separate interacting pairs of avidin-biotin, antigen-antibody, and complementary DNA pairs formed the basis of this field. The method has subsequently been applied to map the presence of cell surface receptors and polysaccharides on live cell membranes by force measurement, with promising results. Attempts to extract phospholipids and proteins from lipid bilayers and live cell surfaces have been reported, providing a new tool for the manipulation of cellular activities and biochemical analysis at the single-cell level. An increasing awareness of the effect of the pulling speed (nm/s or microm/s), or more accurately, the force loading rate (pN/s or nN/s) on the magnitude of the rupture force, has led researchers to construct energy diagrams of rupture events based on the parameters available from such studies. Information on such nature of the interplay of force and loading rate is vital for nanomanipulation of living cells and cell membranes. Some relevant work for membrane manipulation using other methods is also reviewed in relation to AFM-based methodology.

  4. Cell cycle regulation of hematopoietic stem or progenitor cells. (United States)

    Hao, Sha; Chen, Chen; Cheng, Tao


    The highly regulated process of blood production is achieved through the hierarchical organization of hematopoietic stem cell (HSC) subsets and their progenies, which differ in self-renewal and differentiation potential. Genetic studies in mice have demonstrated that cell cycle is tightly controlled by the complex interplay between extrinsic cues and intrinsic regulatory pathways involved in HSC self-renewal and differentiation. Deregulation of these cellular programs may transform HSCs or hematopoietic progenitor cells (HPCs) into disease-initiating stem cells, and can result in hematopoietic malignancies such as leukemia. While previous studies have shown roles for some cell cycle regulators and related signaling pathways in HSCs and HPCs, a more complete picture regarding the molecular mechanisms underlying cell cycle regulation in HSCs or HPCs is lacking. Based on accumulated studies in this field, the present review introduces the basic components of the cell cycle machinery and discusses their major cellular networks that regulate the dormancy and cell cycle progression of HSCs. Knowledge on this topic would help researchers and clinicians to better understand the pathogenesis of relevant blood disorders and to develop new strategies for therapeutic manipulation of HSCs.

  5. Performance Degradation Tests of Phosphoric Acid Doped Polybenzimidazole Membrane Based High Temperature Polymer Electrolyte Membrane Fuel Cells

    DEFF Research Database (Denmark)

    Zhou, Fan; Araya, Samuel Simon; Grigoras, Ionela


    Degradation tests of two phosphoric acid (PA) doped PBI membrane based HT-PEM fuel cells were reported in this paper to investigate the effects of start/stop and the presence of methanol in the fuel to the performance degradation of the HT-PEM fuel cell. Continuous tests with pure dry H2 and meth......Degradation tests of two phosphoric acid (PA) doped PBI membrane based HT-PEM fuel cells were reported in this paper to investigate the effects of start/stop and the presence of methanol in the fuel to the performance degradation of the HT-PEM fuel cell. Continuous tests with pure dry H2...... to the corrosion of carbon support in the catalyst layer and degradation of the PBI membrane. During the continuous test with methanol containing H2 as the fuel the reaction kinetic resistance and mass transfer resistance of both single cells increased, which may be caused by the adsorption of methanol...

  6. Membrane-localized ubiquitin ligase ATL15 functions in sugar-responsive growth regulation in Arabidopsis. (United States)

    Aoyama, Shoki; Terada, Saki; Sanagi, Miho; Hasegawa, Yoko; Lu, Yu; Morita, Yoshie; Chiba, Yukako; Sato, Takeo; Yamaguchi, Junji


    Ubiquitin ligases play important roles in regulating various cellular processes by modulating the protein function of specific ubiquitination targets. The Arabidopsis Tóxicos en Levadura (ATL) family is a group of plant-specific RING-type ubiquitin ligases that localize to membranes via their N-terminal transmembrane-like domains. To date, 91 ATL isoforms have been identified in the Arabidopsis genome, with several ATLs reported to be involved in regulating plant responses to environmental stresses. However, the functions of most ATLs remain unknown. This study, involving transcriptome database analysis, identifies ATL15 as a sugar responsive ATL gene in Arabidopsis. ATL15 expression was rapidly down-regulated in the presence of sugar. The ATL15 protein showed ubiquitin ligase activity in vitro and localized to plasma membrane and endomembrane compartments. Further genetic analyses demonstrated that the atl15 knockout mutants are insensitive to high glucose concentrations, whereas ATL15 overexpression depresses plant growth. In addition, endogenous glucose and starch amounts were reciprocally affected in the atl15 knockout mutants and the ATL15 overexpressors. These results suggest that ATL15 protein plays a significant role as a membrane-localized ubiquitin ligase that regulates sugar-responsive plant growth in Arabidopsis. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Copine1 regulates neural stem cell functions during brain development. (United States)

    Kim, Tae Hwan; Sung, Soo-Eun; Cheal Yoo, Jae; Park, Jae-Yong; Yi, Gwan-Su; Heo, Jun Young; Lee, Jae-Ran; Kim, Nam-Soon; Lee, Da Yong


    Copine 1 (CPNE1) is a well-known phospholipid binding protein in plasma membrane of various cell types. In brain cells, CPNE1 is closely associated with AKT signaling pathway, which is important for neural stem cell (NSC) functions during brain development. Here, we investigated the role of CPNE1 in the regulation of brain NSC functions during brain development and determined its underlying mechanism. In this study, abundant expression of CPNE1 was observed in neural lineage cells including NSCs and immature neurons in human. With mouse brain tissues in various developmental stages, we found that CPNE1 expression was higher at early embryonic stages compared to postnatal and adult stages. To model developing brain in vitro, we used primary NSCs derived from mouse embryonic hippocampus. Our in vitro study shows decreased proliferation and multi-lineage differentiation potential in CPNE1 deficient NSCs. Finally, we found that the deficiency of CPNE1 downregulated mTOR signaling in embryonic NSCs. These data demonstrate that CPNE1 plays a key role in the regulation of NSC functions through the activation of AKT-mTOR signaling pathway during brain development. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Regulation of membrane protein function by lipid bilayer elasticity-a single molecule technology to measure the bilayer properties experienced by an embedded protein

    International Nuclear Information System (INIS)

    Lundbaek, Jens August


    Membrane protein function is generally regulated by the molecular composition of the host lipid bilayer. The underlying mechanisms have long remained enigmatic. Some cases involve specific molecular interactions, but very often lipids and other amphiphiles, which are adsorbed to lipid bilayers, regulate a number of structurally unrelated proteins in an apparently non-specific manner. It is well known that changes in the physical properties of a lipid bilayer (e.g., thickness or monolayer spontaneous curvature) can affect the function of an embedded protein. However, the role of such changes, in the general regulation of membrane protein function, is unclear. This is to a large extent due to lack of a generally accepted framework in which to understand the many observations. The present review summarizes studies which have demonstrated that the hydrophobic interactions between a membrane protein and the host lipid bilayer provide an energetic coupling, whereby protein function can be regulated by the bilayer elasticity. The feasibility of this 'hydrophobic coupling mechanism' has been demonstrated using the gramicidin channel, a model membrane protein, in planar lipid bilayers. Using voltage-dependent sodium channels, N-type calcium channels and GABA A receptors, it has been shown that membrane protein function in living cells can be regulated by amphiphile induced changes in bilayer elasticity. Using the gramicidin channel as a molecular force transducer, a nanotechnology to measure the elastic properties experienced by an embedded protein has been developed. A theoretical and technological framework, to study the regulation of membrane protein function by lipid bilayer elasticity, has been established

  9. A novel membrane-less direct alcohol fuel cell (United States)

    Yi, Qingfeng; Chen, Qinghua; Yang, Zheng


    Membrane-less fuel cell possesses such advantages as simplified design and lower cost. In this paper, a membrane-less direct alcohol fuel cell is constructed by using multi-walled carbon nanotubes (MWCNT) supported Pd and ternary PdSnNi composites as the anode catalysts and Fe/C-PANI composite, produced by direct pyrolysis of Fe-doped polyaniline precursor, as the oxygen reduction reaction (ORR) catalyst. The alcohols investigated in the present study are methanol, ethanol, n-propanol, iso-propanol, n-butanol, iso-butanol and sec-butanol. The cathode catalyst Fe/C-PANI is electrochemically inactive to oxidation of the alcohols. The performance of the cell with various alcohols in 1 mol L-1 NaOH solution on either Pd/MWCNT or PdSnNi/MWCNT catalyst has been evaluated. In any case, the performance of the cell using the anode catalyst PdSnNi/MWCNT is considerably better than Pd/MWCNT. For the PdSnNi/MWCNT, the maximum power densities of the cell using methanol (0.5 mol L-1), ethanol (0.5 mol L-1), n-propanol (0.5 mol L-1), iso-propanol (0.5 mol L-1), n-butanol (0.2 mol L-1), iso-butanol (0.2 mol L-1) and sec-butanol (0.2 mol L-1) are 0.34, 1.03, 1.07, 0.44, 0.50, 0.31 and 0.15 mW cm-2, respectively.

  10. The Drosophila actin regulator ENABLED regulates cell shape and orientation during gonad morphogenesis.

    Directory of Open Access Journals (Sweden)

    Hiroko Sano

    Full Text Available Organs develop distinctive morphologies to fulfill their unique functions. We used Drosophila embryonic gonads as a model to study how two different cell lineages, primordial germ cells (PGCs and somatic gonadal precursors (SGPs, combine to form one organ. We developed a membrane GFP marker to image SGP behaviors live. These studies show that a combination of SGP cell shape changes and inward movement of anterior and posterior SGPs leads to the compaction of the spherical gonad. This process is disrupted in mutants of the actin regulator, enabled (ena. We show that Ena coordinates these cell shape changes and the inward movement of the SGPs, and Ena affects the intracellular localization of DE-cadherin (DE-cad. Mathematical simulation based on these observations suggests that changes in DE-cad localization can generate the forces needed to compact an elongated structure into a sphere. We propose that Ena regulates force balance in the SGPs by sequestering DE-cad, leading to the morphogenetic movement required for gonad compaction.

  11. Redox regulation in cancer stem cells (United States)

    Reactive oxygen species (ROS) and ROS-dependent (redox regulation) signaling pathways and transcriptional activities are thought to be critical in stem cell self-renewal and differentiation during growth and organogenesis. Aberrant ROS burst and dysregulation of those ROS-dependent cellular processe...

  12. Eisosomes promote the ability of Sur7 to regulate plasma membrane organization in Candida albicans.

    Czech Academy of Sciences Publication Activity Database

    Wang, H.X.; Douglas, L.M.; Veselá, Petra; Rachel, R.; Malínský, Jan; Konopka, J.B.


    Roč. 27, č. 10 (2016), s. 1663-1675 ISSN 1059-1524 R&D Projects: GA ČR(CZ) GA15-10641S Institutional support: RVO:68378041 Keywords : plasma membrane * eisosome * PIL1 * SUR7 Subject RIV: EA - Cell Biology Impact factor: 3.685, year: 2016

  13. Membrane dynamics and interactions in measles virus dendritic cell infections. (United States)

    Avota, Elita; Koethe, Susanne; Schneider-Schaulies, Sibylle


    Viral entry, compartmentalization and transmission depend on the formation of membrane lipid/protein microdomains concentrating receptors and signalosomes. Dendritic cells (DCs) are prime targets for measles virus (MV) infection, and this interaction promotes immune activation and generalized immunosuppression, yet also MV transport to secondary lymphatics where transmission to T cells occurs. In addition to MV trapping, DC-SIGN interaction can enhance MV uptake by activating cellular sphingomyelinases and, thereby, vertical surface transport of its entry receptor CD150. To exploit DCs as Trojan horses for transport, MV promotes DC maturation accompanied by mobilization, and restrictions of viral replication in these cells may support this process. MV-infected DCs are unable to support formation of functional immune synapses with conjugating T cells and signalling via viral glycoproteins or repulsive ligands (such as semaphorins) plays a key role in the induction of T-cell paralysis. In the absence of antigen recognition, MV transmission from infected DCs to T cells most likely involves formation of polyconjugates which concentrate viral structural proteins, viral receptors and with components enhancing either viral uptake or conjugate stability. Because DCs barely support production of infectious MV particles, these organized interfaces are likely to represent virological synapses essential for MV transmission. © 2012 Blackwell Publishing Ltd.

  14. Vesicle-associated membrane protein 7 (VAMP-7) is essential for target cell killing in a natural killer cell line

    International Nuclear Information System (INIS)

    Marcet-Palacios, Marcelo; Odemuyiwa, Solomon O.; Coughlin, Jason J.; Garofoli, Daniella; Ewen, Catherine; Davidson, Courtney E.; Ghaffari, Mazyar; Kane, Kevin P.; Lacy, Paige; Logan, Michael R.; Befus, A. Dean; Bleackley, R. Chris; Moqbel, Redwan


    Natural killer cells recognize and induce apoptosis in foreign, transformed or virus-infected cells through the release of perforin and granzymes from secretory lysosomes. Clinically, NK-cell mediated killing is a major limitation to successful allo- and xenotransplantation. The molecular mechanisms that regulate the fusion of granzyme B-containing secretory lysosomes to the plasma membrane in activated NK cells, prior to target cell killing, are not fully understood. Using the NK cell line YT-Indy as a model, we have investigated the expression of SNAP REceptors (SNAREs), both target (t-) and vesicular (v-) SNAREs, and their function in granzyme B-mediated target cell killing. Our data showed that YT-Indy cells express VAMP-7 and SNAP-23, but not VAMP-2. VAMP-7 was associated with granzyme B-containing lysosomal granules. Using VAMP-7 small interfering RNA (siRNA), we successfully knocked down the expression of VAMP-7 protein in YT-Indy to less than 10% of untreated cells in 24 h. VAMP7-deficient YT-Indy cells activated via co-culture with Jurkat cells released <1 ng/mL of granzyme B, compared to 1.5-2.5 μg/mL from controls. Using Jurkat cells as targets, we showed a 7-fold reduction in NK cell-mediated killing by VAMP-7 deficient YT-Indy cells. Our results show that VAMP-7 is a crucial component of granzyme B release and target cell killing in the NK cell line YT-Indy. Thus, targeting VAMP-7 expression specifically with siRNA, following transplantation, may be a viable strategy for preventing NK cell-mediated transplant rejection, in vivo

  15. The Effect of Platinum Electrocatalyst on Membrane Degradation in Polymer Electrolyte Fuel Cells. (United States)

    Bodner, Merit; Cermenek, Bernd; Rami, Mija; Hacker, Viktor


    Membrane degradation is a severe factor limiting the lifetime of polymer electrolyte fuel cells. Therefore, obtaining a deeper knowledge is fundamental in order to establish fuel cells as competitive product. A segmented single cell was operated under open circuit voltage with alternating relative humidity. The influence of the catalyst layer on membrane degradation was evaluated by measuring a membrane without electrodes and a membrane-electrode-assembly under identical conditions. After 100 h of accelerated stress testing the proton conductivity of membrane samples near the anode and cathode was investigated by means of ex situ electrochemical impedance spectroscopy. The membrane sample near the cathode inlet exhibited twofold lower membrane resistance and a resulting twofold higher proton conductivity than the membrane sample near the anode inlet. The results from the fluoride ion analysis have shown that the presence of platinum reduces the fluoride emission rate; which supports conclusions drawn from the literature.

  16. The Effect of Platinum Electrocatalyst on Membrane Degradation in Polymer Electrolyte Fuel Cells (United States)

    Bodner, Merit; Cermenek, Bernd; Rami, Mija; Hacker, Viktor


    Membrane degradation is a severe factor limiting the lifetime of polymer electrolyte fuel cells. Therefore, obtaining a deeper knowledge is fundamental in order to establish fuel cells as competitive product. A segmented single cell was operated under open circuit voltage with alternating relative humidity. The influence of the catalyst layer on membrane degradation was evaluated by measuring a membrane without electrodes and a membrane-electrode-assembly under identical conditions. After 100 h of accelerated stress testing the proton conductivity of membrane samples near the anode and cathode was investigated by means of ex situ electrochemical impedance spectroscopy. The membrane sample near the cathode inlet exhibited twofold lower membrane resistance and a resulting twofold higher proton conductivity than the membrane sample near the anode inlet. The results from the fluoride ion analysis have shown that the presence of platinum reduces the fluoride emission rate; which supports conclusions drawn from the literature. PMID:26670258

  17. Difference in membrane repair capacity between cancer cell lines and a normal cell line

    DEFF Research Database (Denmark)

    Frandsen, Stine Krog; McNeil, Anna K.; Novak, Ivana


    repair was investigated by disrupting the plasma membrane using laser followed by monitoring fluorescent dye entry over time in seven cancer cell lines, an immortalized cell line, and a normal primary cell line. The kinetics of repair in living cells can be directly recorded using this technique......, providing a sensitive index of repair capacity. The normal primary cell line of all tested cell lines exhibited the slowest rate of dye entry after laser disruption and lowest level of dye uptake. Significantly, more rapid dye uptake and a higher total level of dye uptake occurred in six of the seven tested...

  18. Inositol phospholipids regulate the guanine-nucleotide-exchange factor Tiam1 by facilitating its binding to the plasma membrane and regulating GDP/GTP exchange on Rac1. (United States)

    Fleming, Ian N; Batty, Ian H; Prescott, Alan R; Gray, Alex; Kular, Gursant S; Stewart, Hazel; Downes, C Peter


    Binding of the Rac1-specific guanine-nucleotide-exchange factor, Tiam1, to the plasma membrane requires the N-terminal pleckstrin homology domain. In the present study, we show that membrane-association is mediated by binding of PtdIns(4,5)P(2) to the pleckstrin homology domain. Moreover, in 1321N1 astrocytoma cells, translocation of Tiam1 to the cytosol, following receptor-mediated stimulation of PtdIns(4,5)P(2) breakdown, correlates with decreased Rac1-GTP levels, indicating that membrane-association is required for GDP/GTP exchange on Rac1. In addition, we show that platelet-derived growth factor activates Rac1 in vivo by increasing PtdIns(3,4,5)P(3) concentrations, rather than the closely related lipid, PtdIns(3,4)P(2). Finally, the data demonstrate that PtdIns(4,5)P(2) and PtdIns(3,4,5)P(3) bind to the same pleckstrin homology domain in Tiam1 and that soluble inositol phosphates appear to compete with lipids for this binding. Together, these novel observations provide strong evidence that distinct phosphoinositides regulate different functions of this enzyme, indicating that local concentrations of signalling lipids and the levels of cytosolic inositol phosphates will play crucial roles in determining its activity in vivo.

  19. Autophagic regulation of smooth muscle cell biology

    Directory of Open Access Journals (Sweden)

    Joshua K. Salabei


    Full Text Available Autophagy regulates the metabolism, survival, and function of numerous cell types, including those comprising the cardiovascular system. In the vasculature, changes in autophagy have been documented in atherosclerotic and restenotic lesions and in hypertensive vessels. The biology of vascular smooth muscle cells appears particularly sensitive to changes in the autophagic program. Recent evidence indicates that stimuli or stressors evoked during the course of vascular disease can regulate autophagic activity, resulting in modulation of VSMC phenotype and viability. In particular, certain growth factors and cytokines, oxygen tension, and pharmacological drugs have been shown to trigger autophagy in smooth muscle cells. Importantly, each of these stimuli has a redox component, typically associated with changes in the abundance of reactive oxygen, nitrogen, or lipid species. Collective findings support the hypothesis that autophagy plays a critical role in vascular remodeling by regulating smooth muscle cell phenotype transitions and by influencing the cellular response to stress. In this graphical review, we summarize current knowledge on the role of autophagy in the biology of the smooth muscle cell in (pathophysiology.

  20. Adherence of Helicobacter pylori cells and their surface components to HeLa cell membranes. (United States)

    Fauchère, J L; Blaser, M J


    Four Helicobacter pylori strains were used to develop in vitro methods to assess adherence to HeLa cells. Using direct detection by microscopy, adhesion scores increased with the initial bacteria-to-cell ratio. The urease method assessed H. pylori bound to HeLa cells by their urease activity. The percentage of the original inoculum adhering to HeLa cells remained constant for initial ratios from 10(2) to 10(5) bacteria per cell. An ELISA using anti-H. pylori serum assessed whole bacteria or components bound to HeLa cell fractions. By all three methods, the four H. pylori strains were adherent to HeLa cells or membranes whereas Campylobacter fetus and Providencia control strains were not. The adherence of H. pylori whole cells decreased following extraction with saline, water, or glycine buffer and most of the superficial adhering material (SAM) was present in the saline or water extracts. SAM bound better to HeLa membranes than to calf fetuin or bovine serum albumin (BSA); binding was inhibited by preincubation of SAM with HeLa membranes but not with fetuin or BSA or by pretreatment of HeLa membranes with neuraminidase. These data indicate that SAM has a specific receptor on the HeLa cell membranes. By gel exclusion chromatography of bacterial extracts, the most adherent components were found in the fractions which also contained the highest urease activity; these fractions included urease subunit antigens. We conclude that adherence of H. pylori can be assessed by microtiter assays and involves bacterial surface material which co-purifies with urease and is different from the N-acetyl-neuraminyl-lactose binding hemagglutinin.

  1. Anion exchange membrane fuel cells: Current status and remaining challenges (United States)

    Gottesfeld, Shimshon; Dekel, Dario R.; Page, Miles; Bae, Chulsung; Yan, Yushan; Zelenay, Piotr; Kim, Yu Seung


    The anion exchange membrane fuel cell (AEMFC) is an attractive alternative to acidic proton exchange membrane fuel cells, which to date have required platinum-based catalysts, as well as acid-tolerant stack hardware. The AEMFC could use non-platinum-group metal catalysts and less expensive metal hardware thanks to the high pH of the electrolyte. Over the last decade, substantial progress has been made in improving the performance and durability of the AEMFC through the development of new materials and the optimization of system design and operation conditions. In this perspective article, we describe the current status of AEMFCs as having reached beginning of life performance very close to that of PEMFCs when using ultra-low loadings of Pt, while advancing towards operation on non-platinum-group metal catalysts alone. In the latter sections, we identify the remaining technical challenges, which require further research and development, focusing on the materials and operational factors that critically impact AEMFC performance and/or durability. These perspectives may provide useful insights for the development of next-generation of AEMFCs.

  2. The regulation of apoptotic cell death

    Directory of Open Access Journals (Sweden)

    Amarante-Mendes G.P.


    Full Text Available Apoptosis is a fundamental biological phenomenon in which the death of a cell is genetically and biochemically regulated. Different molecules are involved in the regulation of the apoptotic process. Death receptors, coupled to distinct members of the caspases as well as other adapter molecules, are involved in the initiation of the stress signals (The Indictment. Members of the Bcl-2 family control at the mitochondrial level the decision between life and death (The Judgement. The effector caspases are responsible for all morphological and biochemical changes related to apoptosis including the "eat-me" signals perceived by phagocytes and neighboring cells (The Execution. Finally, apoptosis would have little biological significance without the recognition and removal of the dying cells (The Burial.

  3. The regulation of apoptotic cell death

    Directory of Open Access Journals (Sweden)

    G.P. Amarante-Mendes


    Full Text Available Apoptosis is a fundamental biological phenomenon in which the death of a cell is genetically and biochemically regulated. Different molecules are involved in the regulation of the apoptotic process. Death receptors, coupled to distinct members of the caspases as well as other adapter molecules, are involved in the initiation of the stress signals (The Indictment. Members of the Bcl-2 family control at the mitochondrial level the decision between life and death (The Judgement. The effector caspases are responsible for all morphological and biochemical changes related to apoptosis including the "eat-me" signals perceived by phagocytes and neighboring cells (The Execution. Finally, apoptosis would have little biological significance without the recognition and removal of the dying cells (The Burial.

  4. Reactivation System for Proton-Exchange Membrane Fuel-Cells

    Directory of Open Access Journals (Sweden)

    Roberto Giral


    Full Text Available In recent years, Proton-Exchange Membrane Fuel Cells (PEMFCs have been the focus of very intensive researches. Manufacturers of these alternative power sources propose a rejuvenation sequence after the FC has been operating at high power for a certain period of time. These rejuvenation methods could be not appropriate for the reactivation of the FC when it has been out of operation for a long period of time or after it has been repaired. Since the developed reactivation system monitors temperature, current, and the cell voltages of the stack, it could be also useful for the diagnostic and repairing processes. The limited number of published contributions suggests that systems developing reactivation techniques are an open research field. In this paper, an automated system for reactivating PEMFCs and results of experimental testing are presented.

  5. Regulation of multispanning membrane protein topology via post-translational annealing. (United States)

    Van Lehn, Reid C; Zhang, Bin; Miller, Thomas F


    The canonical mechanism for multispanning membrane protein topogenesis suggests that protein topology is established during cotranslational membrane integration. However, this mechanism is inconsistent with the behavior of EmrE, a dual-topology protein for which the mutation of positively charged loop residues, even close to the C-terminus, leads to dramatic shifts in its topology. We use coarse-grained simulations to investigate the Sec-facilitated membrane integration of EmrE and its mutants on realistic biological timescales. This work reveals a mechanism for regulating membrane-protein topogenesis, in which initially misintegrated configurations of the proteins undergo post-translational annealing to reach fully integrated multispanning topologies. The energetic barriers associated with this post-translational annealing process enforce kinetic pathways that dictate the topology of the fully integrated proteins. The proposed mechanism agrees well with the experimentally observed features of EmrE topogenesis and provides a range of experimentally testable predictions regarding the effect of translocon mutations on membrane protein topogenesis.

  6. Reconstitution of membrane protein complexes involved in pneumococcal septal cell wall assembly.

    Directory of Open Access Journals (Sweden)

    Marjolaine Noirclerc-Savoye

    Full Text Available The synthesis of peptidoglycan, the major component of the bacterial cell wall, is essential to cell survival, yet its mechanism remains poorly understood. In the present work, we have isolated several membrane protein complexes consisting of the late division proteins of Streptococcus pneumoniae: DivIB, DivIC, FtsL, PBP2x and FtsW, or subsets thereof. We have co-expressed membrane proteins from S. pneumoniae in Escherichia coli. By combining two successive affinity chromatography steps, we obtained membrane protein complexes with a very good purity. These complexes are functional, as indicated by the retained activity of PBP2x to bind a fluorescent derivative of penicillin and to hydrolyze the substrate analogue S2d. Moreover, we have evidenced the stabilizing role of protein-protein interactions within each complex. This work paves the way for a complete reconstitution of peptidoglycan synthesis in vitro, which will be critical to the elucidation of its intricate regulation mechanisms.

  7. The RhoGEF TEM4 Regulates Endothelial Cell Migration by Suppressing Actomyosin Contractility.

    Directory of Open Access Journals (Sweden)

    Natalia Mitin

    Full Text Available Persistent cellular migration requires efficient protrusion of the front of the cell, the leading edge where the actin cytoskeleton and cell-substrate adhesions undergo constant rearrangement. Rho family GTPases are essential regulators of the actin cytoskeleton and cell adhesion dynamics. Here, we examined the role of the RhoGEF TEM4, an activator of Rho family GTPases, in regulating cellular migration of endothelial cells. We found that TEM4 promotes the persistence of cellular migration by regulating the architecture of actin stress fibers and cell-substrate adhesions in protruding membranes. Furthermore, we determined that TEM4 regulates cellular migration by signaling to RhoC as suppression of RhoC expression recapitulated the loss-of-TEM4 phenotypes, and RhoC activation was impaired in TEM4-depleted cells. Finally, we showed that TEM4 and RhoC antagonize myosin II-dependent cellular contractility and the suppression of myosin II activity rescued the persistence of cellular migration of TEM4-depleted cells. Our data implicate TEM4 as an essential regulator of the actin cytoskeleton that ensures proper membrane protrusion at the leading edge of migrating cells and efficient cellular migration via suppression of actomyosin contractility.

  8. Cell contact-dependent outer membrane exchange in myxobacteria: genetic determinants and mechanism.

    Directory of Open Access Journals (Sweden)

    Darshankumar T Pathak

    Full Text Available Biofilms are dense microbial communities. Although widely distributed and medically important, how biofilm cells interact with one another is poorly understood. Recently, we described a novel process whereby myxobacterial biofilm cells exchange their outer membrane (OM lipoproteins. For the first time we report here the identification of two host proteins, TraAB, required for transfer. These proteins are predicted to localize in the cell envelope; and TraA encodes a distant PA14 lectin-like domain, a cysteine-rich tandem repeat region, and a putative C-terminal protein sorting tag named MYXO-CTERM, while TraB encodes an OmpA-like domain. Importantly, TraAB are required in donors and recipients, suggesting bidirectional transfer. By use of a lipophilic fluorescent dye, we also discovered that OM lipids are exchanged. Similar to lipoproteins, dye transfer requires TraAB function, gliding motility and a structured biofilm. Importantly, OM exchange was found to regulate swarming and development behaviors, suggesting a new role in cell-cell communication. A working model proposes TraA is a cell surface receptor that mediates cell-cell adhesion for OM fusion, in which lipoproteins/lipids are transferred by lateral diffusion. We further hypothesize that cell contact-dependent exchange helps myxobacteria to coordinate their social behaviors.

  9. A Cell-Cell Fusion Assay to Assess Arenavirus Envelope Glycoprotein Membrane-Fusion Activity. (United States)

    York, Joanne; Nunberg, Jack H


    For many viruses that enter their target cells through pH-dependent fusion of the viral and endosomal membranes, cell-cell fusion assays can provide an experimental platform for investigating the structure-function relationships that promote envelope glycoprotein membrane-fusion activity. Typically, these assays employ effector cells expressing the recombinant envelope glycoprotein on the cell surface and target cells engineered to quantitatively report fusion with the effector cell. In the protocol described here, Vero cells are transfected with a plasmid encoding the arenavirus envelope glycoprotein complex GPC and infected with the vTF7-3 vaccinia virus expressing the bacteriophage T7 RNA polymerase. These effector cells are mixed with target cells infected with the vCB21R-lacZ vaccinia virus encoding a β-galactosidase reporter under the control of the T7 promoter. Cell-cell fusion is induced upon exposure to low-pH medium (pH 5.0), and the resultant expression of the β-galactosidase reporter is quantitated using a chemiluminescent substrate. We have utilized this robust microplate cell-cell fusion assay extensively to study arenavirus entry and its inhibition by small-molecule fusion inhibitors.

  10. Novel secreted isoform of adhesion molecule ICAM-4: Potential regulator of membrane-associated ICAM-4 interactions

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Gloria; Spring, Frances A.; Parons, Stephen F.; Mankelow, Tosti J.; Peters, Luanne L.; Koury, Mark J.; Mohandas, Narla; Anstee, David J.; Chasis, Joel Anne


    ICAM-4, a newly characterized adhesion molecule, is expressed early in human erythropoiesis and functions as a ligand for binding a4b1 and aV integrin-expressing cells. Within the bone marrow, erythroblasts surround central macrophages forming erythroblastic islands. Evidence suggests that these islands are highly specialized subcompartments where cell adhesion events, in concert with cytokines, play critical roles in regulating erythropoiesis and apoptosis. Since erythroblasts express a4b1 and ICAM-4 and macrophages exhibit aV, ICAM-4 is an attractive candidate for mediating cellular interactions within erythroblastic islands. To determine whether ICAM-4 binding properties are conserved across species, we first cloned and sequenced the murine homologue. The translated amino acid sequence showed 68 percent overall identity with human ICAM-4. Using recombinant murine ICAM-4 extracellular domains, we discovered that hematopoietic a4b1-expressing HEL cells and non-hematopoietic aV-expressing FLY cells adhered to mouse ICAM-4. Cell adhesion studies showed that FLY and HEL cells bound to mouse and human proteins with similar avidity. These data strongly suggest conservation of integrin-binding properties across species. Importantly, we characterized a novel second splice cDNA that would be predicted to encode an ICAM-4 isoform, lacking the membrane-spanning domain. Erythroblasts express both isoforms of ICAM-4. COS-7 cells transfected with GFP constructs of prototypic or novel ICAM-4 cDNA showed different cellular localization patterns. Moreover, analysis of tissue culture medium revealed that the novel ICAM-4 cDNA encodes a secreted protein. We postulate that secretion of this newly described isoform, ICAM-4S, may modulate binding of membrane-associated ICAM-4 and could thus play a critical regulatory role in erythroblast molecular attachments.

  11. Modelling and validation of Proton exchange membrane fuel cell (PEMFC) (United States)

    Mohiuddin, A. K. M.; Basran, N.; Khan, A. A.


    This paper is the outcome of a small scale fuel cell project. Fuel cell is an electrochemical device that converts energy from chemical reaction to electrical work. Proton Exchange Membrane Fuel Cell (PEMFC) is one of the different types of fuel cell, which is more efficient, having low operational temperature and fast start up capability results in high energy density. In this study, a mathematical model of 1.2 W PEMFC is developed and simulated using MATLAB software. This model describes the PEMFC behaviour under steady-state condition. This mathematical modeling of PEMFC determines the polarization curve, power generated, and the efficiency of the fuel cell. Simulation results were validated by comparing with experimental results obtained from the test of a single PEMFC with a 3 V motor. The performance of experimental PEMFC is little lower compared to simulated PEMFC, however both results were found in good agreement. Experiments on hydrogen flow rate also been conducted to obtain the amount of hydrogen consumed to produce electrical work on PEMFC.

  12. Pyro-electrification of polymer membranes for cell patterning

    Energy Technology Data Exchange (ETDEWEB)

    Rega, R.; Gennari, O.; Mecozzia, L.; Grilli, S.; Pagliarulo, V.; Ferraro, P. [National Council of Research, Institute of Applied Science & Intelligent Systems (ISASI) ‘E. Caianiello’, Via Campi Flegrei 34, 80078 Pozzuoli (Italy)


    In the recent years, much attention has been devoted to the possibility of charging polymer-based materials, due to their potential in developing large-scale and inexpensive flexible thin-film technology. The availability of localized electrostatic fields is in of great interest for a huge amount of applications such as distribution of biomolecules and cells from the liquid phase. Here we report a voltage-free pyro-electrification (PE) process able to induce permanent dipoles into polymer layers; the lithium niobate (LN) crystal is the key component that plays the multi-purpose role of sustaining, heating and poling the polymer layer that is then peeled-off easily in order to have a free-standing charged membrane. The results show the fascinating application for the living cell patterning. It well known that cell behaviour is affected by chemical and topographical cues of substrate. In fact, polymers, such as polystyrene (PS) and poly(methyl methacrylate) (PMMA), are naturally cytophobic and require specific functionalization treatments in order to promote cell adhesion. Through our proposal technique, it’s possible to obtain spontaneous organization and a driven growth of SH-SY5Y cells that is solely dictated by the nature of the charge polymer surface, opening, in this way, the innovative chance to manipulate and transfer biological samples on a free-standing polymer layer [1].

  13. Vesicles mimicking normal and cancer cell membranes exhibit differential responses to the cell-penetrating peptide Pep-1. (United States)

    Almarwani, Bashiyar; Phambu, Esther Nzuzi; Alexander, Christopher; Nguyen, Ha Aimee T; Phambu, Nsoki; Sunda-Meya, Anderson


    The cell-penetrating peptide (CPP) Pep-1 presents a great potential in drug delivery due to its intrinsic property to cross plasma membrane. However, its mechanism of entry into the cell remains unresolved. In this study, we compare the selectivity of Pep-1 towards vesicles mimicking normal and cancer cell membranes. The interaction was performed in a wide range of peptide-to-lipid molar ratios using infrared (IR), fluorescence, scanning electron microscopy (SEM), thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) techniques. At low peptide concentration, fluorescence experiments show that lipid-phosphatidylserine (PS) seems to enable Pep-1 translocation into cancer cell membrane as evidenced by the blue shift of its maximal emission wavelength. DSC data show that Pep-1 induces segregation of lipids. At high peptide concentration, IR data indicate that the interaction of Pep-1 is relatively stronger with normal cell membrane than with cancer cell membrane through the phosphate groups, while the interaction is weaker with normal cell membrane than with cancer cell membrane through the carbonyl groups. TGA and DSC data reveal that vesicles of normal cell membrane are thermally more stable than vesicles of cancer cell membrane. This suggests that the additional lipid PS included in cancer cell membrane has a destabilizing effect on the membrane structure. SEM images reveal that Pep-1 form superstructures including spherical particles and fibrils in the presence of both model membranes. PS seems to enhance peptide transport across cellular membranes. The biophysical techniques in this study provide valuable insights into the properties of CPPs in drug delivery systems. Copyright © 2018 Elsevier B.V. All rights reserved.

  14. Developmental regulation of tandem promoters for the major outer membrane protein gene of Chlamydia trachomatis. (United States)

    Stephens, R S; Wagar, E A; Edman, U


    Chlamydia trachomatis has a biphasic developmental cycle which is characterized by qualitative and quantitative changes in protein expression. The molecular mechanisms that mediate these changes are unknown. Evidence for transcriptional regulation of the chlamydial major outer membrane protein gene (omp1) was found by Northern hybridization of RNA isolated sequentially during the chlamydial developmental cycle. Early in the growth cycle a single transcript was detected, which was followed hours later in the cycle by an additional transcript. Mapping of the initiating nucleotide for each transcript suggested that this gene is regulated by differential transcription from tandem promoters. Images PMID:2448291

  15. Comparison of human mesenchymal stromal cells from four neonatal tissues: Amniotic membrane, chorionic membrane, placental decidua and umbilical cord. (United States)

    Araújo, Anelise Bergmann; Salton, Gabrielle Dias; Furlan, Juliana Monteiro; Schneider, Natália; Angeli, Melissa Helena; Laureano, Álvaro Macedo; Silla, Lúcia; Passos, Eduardo Pandolfi; Paz, Ana Helena


    Mesenchymal stromal cells (MSCs) are being investigated as a potential alternative for cellular therapy. This study was designed to compare the biological characteristics of MSCs isolated from amniotic membrane (A-MSCs), chorionic membrane (C-MSCs), placental decidua (D-MSCs) and umbilical cord (UC-MSCs) to ascertain whether any one of these sources is superior to the others for cellular therapy purposes. MSCs were isolated from amniotic membrane, chorionic membrane, umbilical cord and placental decidua. Immunophenotype, differentiation ability, cell size, cell complexity, polarity index and growth kinetics of MSCs isolated from these four sources were analyzed. MSCs were successfully isolated from all four sources. Surface marker profile and differentiation ability were consistent with human MSCs. C-MSCs in suspension were the smallest cells, whereas UC-MSCs presented the greatest length and least width. A-MSCs had the lowest polarity index and UC-MSCs, as more elongated cells, the highest. C-MSCs, D-MSCs and UC-MSCs exhibited similar growth capacity until passage 8 (P8); C-MSCs presented better lifespan, whereas insignificant proliferation was observed in A-MSCs. Neonatal and maternal tissues can serve as sources of multipotent stem cells. Some characteristics of MSCs obtained from four neonatal tissues were compared and differences were observed. Amniotic membrane was the least useful source of MSCs, whereas chorionic membrane and umbilical cord were considered good options for future use in cell therapy because of the known advantages of immature cells. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  16. Influence of Silica/Sulfonated Polyether-Ether Ketone as Polymer Electrolyte Membrane for Hydrogen Fueled Proton Exchange Membrane Fuel Cells

    Directory of Open Access Journals (Sweden)

    Sri Handayani


    Full Text Available The operation of non-humidified condition of proton exchange membrane fuel cell (PEMFC using composite sPEEK-silica membrane is reported. Sulfonated membrane of PEEK is known as hydrocarbon polyelectrolyte membrane for PEMFC and direct methanol fuel cell (DMFC. The state of the art of fuel cells is based on the perluorosulfonic acid membrane (Nafion. Nafion has been the most used in both PEMFC and DMFC due to good performance although in low humidified condition showed poor current density. Here we reported the effect of silica in hydrocarbon sPEEK membrane that contributes for a better water management system inside the cell, and showed 0.16 W/cm2 of power density which is 78% higher than that of non-silica modified [Keywords: composite membrane, polyether-ether ketone, silica, proton exchange membrane fuel cell].

  17. VDAC Regulation: A Mitochondrial Target to Stop Cell Proliferation. (United States)

    Fang, Diana; Maldonado, Eduardo N


    Cancer metabolism is emerging as a chemotherapeutic target. Enhanced glycolysis and suppression of mitochondrial metabolism characterize the Warburg phenotype in cancer cells. The flux of respiratory substrates, ADP, and Pi into mitochondria and the release of mitochondrial ATP to the cytosol occur through voltage-dependent anion channels (VDACs) located in the mitochondrial outer membrane. Catabolism of respiratory substrates in the Krebs cycle generates NADH and FADH 2 that enter the electron transport chain (ETC) to generate a proton motive force that maintains mitochondrial membrane potential (ΔΨ) and is utilized to generate ATP. The ETC is also the major cellular source of mitochondrial reactive oxygen species (ROS). αβ-Tubulin heterodimers decrease VDAC conductance in lipid bilayers. High constitutive levels of cytosolic free tubulin in intact cancer cells close VDAC decreasing mitochondrial ΔΨ and mitochondrial metabolism. The VDAC-tubulin interaction regulates VDAC opening and globally controls mitochondrial metabolism, ROS formation, and the intracellular flow of energy. Erastin, a VDAC-binding molecule lethal to some cancer cell types, and erastin-like compounds identified in a high-throughput screening antagonize the inhibitory effect of tubulin on VDAC. Reversal of tubulin inhibition of VDAC increases VDAC conductance and the flux of metabolites into and out of mitochondria. VDAC opening promotes a higher mitochondrial ΔΨ and a global increase in mitochondrial metabolism leading to high cytosolic ATP/ADP ratios that inhibit glycolysis. VDAC opening also increases ROS production causing oxidative stress that, in turn, leads to mitochondrial dysfunction, bioenergetic failure, and cell death. In summary, antagonism of the VDAC-tubulin interaction promotes cell death by a "double-hit model" characterized by reversion of the proproliferative Warburg phenotype (anti-Warburg) and promotion of oxidative stress. © 2018 Elsevier Inc. All rights reserved.

  18. Water-Free Proton-Conducting Membranes for Fuel Cells (United States)

    Narayanan, Sekharipuram; Yen, Shiao-Pin


    Poly-4-vinylpyridinebisulfate (P4VPBS) is a polymeric salt that has shown promise as a water-free proton-conducting material (solid electrolyte) suitable for use in membrane/electrode assemblies in fuel cells. Heretofore, proton-conducting membranes in fuel cells have been made from perfluorinated ionomers that cannot conduct protons in the absence of water and, consequently, cannot function at temperatures >100 C. In addition, the stability of perfluorinated ionomers at temperatures >100 C is questionable. However, the performances of fuel cells of the power systems of which they are parts could be improved if operating temperatures could be raised above 140 C. What is needed to make this possible is a solid-electrolyte material, such as P4VPBS, that can be cast into membranes and that both retains proton conductivity and remains stable in the desired higher operating temperature range. A family of solid-electrolyte materials different from P4VPBS was described in Anhydrous Proton-Conducting Membranes for Fuel Cells (NPO-30493), NASA Tech Briefs, Vol. 29, No. 8 (August 2005), page 48. Those materials notably include polymeric quaternized amine salts. If molecules of such a polymeric salt could be endowed with flexible chain structures, it would be possible to overcome the deficiencies of simple organic amine salts that must melt before being able to conduct protons. However, no polymeric quaternized amine salts have yet shown to be useful in this respect. The present solid electrolyte is made by quaternizing the linear polymer poly- 4-vinylpyridine (P4VP) to obtain P4VPBS. It is important to start with P4VP having a molecular weight of 160,000 daltons because P4VPBS made from lower-molecular-weight P4VP yields brittle membranes. In an experimental synthesis, P4VP was dissolved in methanol and then reacted with an excess of sulfuric acid to precipitate P4VPBS. The precipitate was recovered, washed several times with methanol to remove traces of acid, and dried to a

  19. Materials and characterization techniques for high-temperature polymer electrolyte membrane fuel cells. (United States)

    Zeis, Roswitha


    The performance of high-temperature polymer electrolyte membrane fuel cells (HT-PEMFC) is critically dependent on the selection of materials and optimization of individual components. A conventional high-temperature membrane electrode assembly (HT-MEA) primarily consists of a polybenzimidazole (PBI)-type membrane containing phosphoric acid and two gas diffusion electrodes (GDE), the anode and the cathode, attached to the two surfaces of the membrane. This review article provides a survey on the materials implemented in state-of-the-art HT-MEAs. These materials must meet extremely demanding requirements because of the severe operating conditions of HT-PEMFCs. They need to be electrochemically and thermally stable in highly acidic environment. The polymer membranes should exhibit high proton conductivity in low-hydration and even anhydrous states. Of special concern for phosphoric-acid-doped PBI-type membranes is the acid loss and management during operation. The slow oxygen reduction reaction in HT-PEMFCs remains a challenge. Phosphoric acid tends to adsorb onto the surface of the platinum catalyst and therefore hampers the reaction kinetics. Additionally, the binder material plays a key role in regulating the hydrophobicity and hydrophilicity of the catalyst layer. Subsequently, the binder controls the electrode-membrane interface that establishes the triple phase boundary between proton conductive electrolyte, electron conductive catalyst, and reactant gases. Moreover, the elevated operating temperatures promote carbon corrosion and therefore degrade the integrity of the catalyst support. These are only some examples how materials properties affect the stability and performance of HT-PEMFCs. For this reason, materials characterization techniques for HT-PEMFCs, either in situ or ex situ, are highly beneficial. Significant progress has recently been made in this field, which enables us to gain a better understanding of underlying processes occurring during fuel cell

  20. Circulating cell membrane microparticles transfer heme to endothelial cells and trigger vasoocclusions in sickle cell disease. (United States)

    Camus, Stéphane M; De Moraes, João A; Bonnin, Philippe; Abbyad, Paul; Le Jeune, Sylvain; Lionnet, François; Loufrani, Laurent; Grimaud, Linda; Lambry, Jean-Christophe; Charue, Dominique; Kiger, Laurent; Renard, Jean-Marie; Larroque, Claire; Le Clésiau, Hervé; Tedgui, Alain; Bruneval, Patrick; Barja-Fidalgo, Christina; Alexandrou, Antigoni; Tharaux, Pierre-Louis; Boulanger, Chantal M; Blanc-Brude, Olivier P


    Intravascular hemolysis describes the relocalization of heme and hemoglobin (Hb) from erythrocytes to plasma. We investigated the concept that erythrocyte membrane microparticles (MPs) concentrate cell-free heme in human hemolytic diseases, and that heme-laden MPs have a physiopathological impact. Up to one-third of cell-free heme in plasma from 47 patients with sickle cell disease (SCD) was sequestered in circulating MPs. Erythrocyte vesiculation in vitro produced MPs loaded with heme. In silico analysis predicted that externalized phosphatidylserine (PS) in MPs may associate with and help retain heme at the cell surface. Immunohistology identified Hb-laden MPs adherent to capillary endothelium in kidney biopsies from hyperalbuminuric SCD patients. In addition, heme-laden erythrocyte MPs adhered and transferred heme to cultured endothelial cells, inducing oxidative stress and apoptosis. In transgenic SAD mice, infusion of heme-laden MPs triggered rapid vasoocclusions in kidneys and compromised microvascular dilation ex vivo. These vascular effects were largely blocked by heme-scavenging hemopexin and by the PS antagonist annexin-a5, in vitro and in vivo. Adversely remodeled MPs carrying heme may thus be a source of oxidant stress for the endothelium, linking hemolysis to vascular injury. This pathway might provide new targets for the therapeutic preservation of vascular function in SCD. © 2015 by The American Society of Hematology.

  1. Lipid-protein interactions in plasma membranes of fiber cells isolated from the human eye lens. (United States)

    Raguz, Marija; Mainali, Laxman; O'Brien, William J; Subczynski, Witold K


    The protein content in human lens membranes is extremely high, increases with age, and is higher in the nucleus as compared with the cortex, which should strongly affect the organization and properties of the lipid bilayer portion of intact membranes. To assess these effects, the intact cortical and nuclear fiber cell plasma membranes isolated from human lenses from 41- to 60-year-old donors were studied using electron paramagnetic resonance spin-labeling methods. Results were compared with those obtained for lens lipid membranes prepared from total lipid extracts from human eyes of the same age group [Mainali, L., Raguz, M., O'Brien, W. J., and Subczynski, W. K. (2013) Biochim. Biophys. Acta]. Differences were considered to be mainly due to the effect of membrane proteins. The lipid-bilayer portions of intact membranes were significantly less fluid than lipid bilayers of lens lipid membranes, prepared without proteins. The intact membranes were found to contain three distinct lipid environments termed the bulk lipid domain, boundary lipid domain, and trapped lipid domain. However, the cholesterol bilayer domain, which was detected in cortical and nuclear lens lipid membranes, was not detected in intact membranes. The relative amounts of bulk and trapped lipids were evaluated. The amount of lipids in domains uniquely formed due to the presence of membrane proteins was greater in nuclear membranes than in cortical membranes. Thus, it is evident that the rigidity of nuclear membranes is greater than that of cortical membranes. Also the permeability coefficients for oxygen measured in domains of nuclear membranes were significantly lower than appropriate coefficients measured in cortical membranes. Relationships between the organization of lipids into lipid domains in fiber cells plasma membranes and the organization of membrane proteins are discussed. Copyright © 2014 Elsevier Ltd. All rights reserved.

  2. Tuning Collective Cell Migration by Cell-Cell Junction Regulation

    NARCIS (Netherlands)

    Friedl, P.; Mayor, R.


    Collective cell migration critically depends on cell-cell interactions coupled to a dynamic actin cytoskeleton. Important cell-cell adhesion receptor systems implicated in controlling collective movements include cadherins, immunoglobulin superfamily members (L1CAM, NCAM, ALCAM), Ephrin/Eph

  3. New polymeric electrolyte membranes based on proton donor proton acceptor properties for direct methanol fuel cells

    NARCIS (Netherlands)

    Manea, G.C.; Mulder, M.H.V.


    In order to reduce the high methanol permeability of membranes in a direct methanol fuel cell application new and better materials are still required. In this paper membranes made from polybenzimidazole/sulfonated polysulfone are given and compared with homopolymer membranes made from sulfonated

  4. New Insight Into the Roles of Membrane Microdomains in Physiological Activities of Fungal Cells.

    Czech Academy of Sciences Publication Activity Database

    Malínský, Jan; Opekarová, Miroslava


    Roč. 325, mar. (2016), s. 119-180 ISSN 1937-6448 R&D Projects: GA ČR(CZ) GA15-10641S Institutional support: RVO:68378041 Keywords : membrane microdomain * membrane structure * fungi * membrane contact sites Subject RIV: EA - Cell Biology Impact factor: 3.752, year: 2015

  5. Development of proton-conducting membranes for fuel cells: a review

    Czech Academy of Sciences Publication Activity Database

    Schauer, Jan


    Roč. 10, - (2006), s. 19-26 ISSN 0972-446X R&D Projects: GA ČR GA203/05/0080 Institutional research plan: CEZ:AV0Z40500505 Keywords : proton exchange membrane * fuel cell membrane * polymer membrane Subject RIV: CD - Macromolecular Chemistry

  6. Novel composite membranes based on PBI and dicationic ionic liquids for high temperature polymer electrolyte membrane fuel cells

    International Nuclear Information System (INIS)

    Hooshyari, Khadijeh; Javanbakht, Mehran; Adibi, Mina


    Two types of innovative composite membranes based on polybenzimidazole (PBI) containing dicationic ionic liquid 1,3-di(3-methylimidazolium) propane bis (trifluoromethylsulfonyl) imide (PDC 3 ) and monocationic ionic liquid 1-hexyl-3-methylimidazolium bis (trifluoromethanesulfonyl) imide (PMC 6 ) are prepared as electrolyte for high temperature fuel cells applications under anhydrous conditions. The analyses of results display promising characteristics such as high proton conductivity and thermal stability. Moreover the fuel cell performance of PA doped PDC 3 composite membranes is enhanced in comparison with PA doped PMC 6 and PA doped PBI membranes at high temperatures. Dicationic ionic liquid with high number of charge carriers provides well-developed ionic channels which form facile pathways and considerably develop the anhydrous proton conductivity. The highest proton conductivity of 81 mS/cm is achieved for PA doped PDC 3 composite membranes with PBI/IL mole ratio: 4 at 180 °C. A power density of 0.44 W/cm 2 is obtained at 0.5 V and 180 °C for PA doped PDC 3 composite membranes, which proves that these developed composite membranes can be considered as most promising candidates for high temperature fuel cell applications with enhanced proton conductivity.

  7. Introducing catalyst in alkaline membrane for improved performance direct borohydride fuel cells (United States)

    Qin, Haiying; Lin, Longxia; Chu, Wen; Jiang, Wei; He, Yan; Shi, Qiao; Deng, Yonghong; Ji, Zhenguo; Liu, Jiabin; Tao, Shanwen


    A catalytic material is introduced into the polymer matrix to prepare a novel polymeric alkaline electrolyte membrane (AEM) which simultaneously increases ionic conductivity, reduces the fuel cross-over. In this work, the hydroxide anion exchange membrane is mainly composed of poly(vinylalcohol) and alkaline exchange resin. CoCl2 is added into the poly(vinylalcohol) and alkaline exchange resin gel before casting the membrane to introduce catalytic materials. CoCl2 is converted into CoOOH after the reaction with KOH solution. The crystallinity of the polymer matrix decreases and the ionic conductivity of the composite membrane is notably improved by the introduction of Co-species. A direct borohydride fuel cell using the composite membrane exhibits an open circuit voltage of 1.11 V at 30 °C, which is notably higher than that of cells using other AEMs. The cell using the composite membrane achieves a maximum power density of 283 mW cm-2 at 60 °C while the cell using the membrane without Co-species only reaches 117 mW cm-2 at the same conditions. The outstanding performance of the cell using the composite membrane benefits from impregnation of the catalytic Co-species in the membrane, which not only increases the ionic conductivity but also reduces electrode polarization thus improves the fuel cell performance. This work provides a new approach to develop high-performance fuel cells through adding catalysts in the electrolyte membrane.

  8. Binding and Fusion of Extracellular Vesicles to the Plasma Membrane of Their Cell Targets. (United States)

    Prada, Ilaria; Meldolesi, Jacopo


    Exosomes and ectosomes, extracellular vesicles of two types generated by all cells at multivesicular bodies and the plasma membrane, respectively, play critical roles in physiology and pathology. A key mechanism of their function, analogous for both types of vesicles, is the fusion of their membrane to the plasma membrane of specific target cells, followed by discharge to the cytoplasm of their luminal cargo containing proteins, RNAs, and DNA. Here we summarize the present knowledge about the interactions, binding and fusions of vesicles with the cell plasma membrane. The sequence initiates with dynamic interactions, during which vesicles roll over the plasma membrane, followed by the binding of specific membrane proteins to their cell receptors. Membrane binding is then converted rapidly into fusion by mechanisms analogous to those of retroviruses. Specifically, proteins of the extracellular vesicle membranes are structurally rearranged, and their hydrophobic sequences insert into the target cell plasma membrane which undergoes lipid reorganization, protein restructuring and membrane dimpling. Single fusions are not the only process of vesicle/cell interactions. Upon intracellular reassembly of their luminal cargoes, vesicles can be regenerated, released and fused horizontally to other target cells. Fusions of extracellular vesicles are relevant also for specific therapy processes, now intensely investigated.

  9. The study of preparation for immobilized cells membranes of E. Coli. by radiation technique

    International Nuclear Information System (INIS)

    Cao Jin; Chen Pin; Yu Yi


    The paper described the preparation of immobilized cells membranes with E. Coli by radiation technique. The nylon 6 was grafted with HEMA, which as a matrix to prepare immobilized cells membranes with E. Coli. by radiation entrapment at low temperature. The results showed that the retentive activity possessed a maximum value for membranes with E. Coli. when the irradiation dose was at 10-12 kGy, the entrapped cells has 2.3 g/ml at 50% HEMA concentration, the optimum pH and optimum temperature for membranes with E. Coli. are as same the original cells

  10. Infectious rotavirus enters cells by direct cell membrane penetration, not by endocytosis

    Energy Technology Data Exchange (ETDEWEB)

    Kaljot, K.T.; Shaw, R.D.; Greenberg, H.B. (Stanford Univ. School of Medicine, CA (USA) Palo Alto Veterans Administration Medical Center, CA (USA)); Rubin, D.H. (Univ. of Pennsylvania, Philadelphia (USA))


    Rotaviruses are icosahedral viruses with a segmented, double-stranded RNA genome. They are the major cause of severe infantile infectious diarrhea. Rotavirus growth in tissue culture is markedly enhanced by pretreatment of virus with trypsin. Trypsin activation is associated with cleavage of the viral hemagglutinin (viral protein 3 (VP3); 88 kilodaltons) into two fragments (60 and 28 kilodaltons). The mechanism by which proteolytic cleavage leads to enhanced growth is unknown. To determine whether trypsin treatment affected rotavirus internalization, the authors studied the kinetics of entry of infectious rhesus rotavirus (RRV) into MA104 cells. Trypsin-activated RRV was internalized with a half-time of 3 to 5 min, while nonactivated virus disappeared from the cell surface with a half-time of 30 to 50 min. In contrast to trypsin-activated RRV, loss of nonactivated RRV from the cell surface did not result in the appearance of infection, as measured by plaque formation. Purified trypsin-activated RRV added to cell monolayers at pH 7.4 mediated {sup 51}Cr, ({sup 14}C)choline, and ({sup 3}H)inositol released from prelabeled MA104 cells. This release could be specifically blocked by neutralizing antibodies to VP3. These results suggest that MA104 cell infection follows the rapid entry of trypsin-activated RRV by direct cell membrane penetration. Cell membrane penetration of infectious RRV is initiated by trypsin cleavage of VP3. Neutralizing antibodies can inhibit this direct membrane penetration.

  11. Infectious rotavirus enters cells by direct cell membrane penetration, not by endocytosis

    International Nuclear Information System (INIS)

    Kaljot, K.T.; Shaw, R.D.; Greenberg, H.B.; Rubin, D.H.


    Rotaviruses are icosahedral viruses with a segmented, double-stranded RNA genome. They are the major cause of severe infantile infectious diarrhea. Rotavirus growth in tissue culture is markedly enhanced by pretreatment of virus with trypsin. Trypsin activation is associated with cleavage of the viral hemagglutinin (viral protein 3 [VP3]; 88 kilodaltons) into two fragments (60 and 28 kilodaltons). The mechanism by which proteolytic cleavage leads to enhanced growth is unknown. To determine whether trypsin treatment affected rotavirus internalization, the authors studied the kinetics of entry of infectious rhesus rotavirus (RRV) into MA104 cells. Trypsin-activated RRV was internalized with a half-time of 3 to 5 min, while nonactivated virus disappeared from the cell surface with a half-time of 30 to 50 min. In contrast to trypsin-activated RRV, loss of nonactivated RRV from the cell surface did not result in the appearance of infection, as measured by plaque formation. Purified trypsin-activated RRV added to cell monolayers at pH 7.4 mediated 51 Cr, [ 14 C]choline, and [ 3 H]inositol released from prelabeled MA104 cells. This release could be specifically blocked by neutralizing antibodies to VP3. These results suggest that MA104 cell infection follows the rapid entry of trypsin-activated RRV by direct cell membrane penetration. Cell membrane penetration of infectious RRV is initiated by trypsin cleavage of VP3. Neutralizing antibodies can inhibit this direct membrane penetration


    Energy Technology Data Exchange (ETDEWEB)

    Shamsuddin Ilias


    Fuel cells are electrochemical devices that convert the available chemical free energy directly into electrical energy, without going through heat exchange process. Of all different types of fuel cells, the Proton Exchange Membrane Fuel Cell (PEMFC) is one of the most promising power sources for stand-alone utility and electric vehicle applications. Platinum (Pt) Catalyst is used for both fuel and air electrodes in PEMFCs. However, carbon monoxide (CO) contamination of H{sub 2} greatly affects electro catalysts used at the anode of PEMFCs and decreases cell performance. The irreversible poisoning of the anode can occur even in CO concentrations as low as few parts per million (ppm). In this work, we have synthesized several novel elctrocatalysts (Pt/C, Pt/Ru/C, Pt/Mo/C, Pt/Ir and Pt/Ru/Mo) for PEMFCs. These catalysts have been tested for CO tolerance in the H{sub 2}/air fuel cell, using CO concentrations in the H{sub 2} fuel that varies from 10 to 100 ppm. The performance of the electrodes was evaluated by determining the cell potential against current density. The effects of catalyst composition and electrode film preparation method on the performance of PEM fuel cell were also studied. It was found that at 70 C and 3.5 atm pressure at the cathode, Pt-alloy catalyst (10 wt% Pt/Ru/C, 20 wt% Pt/Mo/C) were more CO tolerant than the 20 wt% Pt/C catalyst alone. It was also observed that spraying method was better than the brushing technique for the preparation of electrode film.

  13. Membrane stress is coupled to a rapid translational control of gene expression in chlorpromazine-treated cells. (United States)

    De Filippi, Loic; Fournier, Margot; Cameroni, Elisabetta; Linder, Patrick; De Virgilio, Claudio; Foti, Michelangelo; Deloche, Olivier


    Chlorpromazine (CPZ) is a small permeable cationic amphiphilic molecule that inserts into membrane bilayers and binds to anionic lipids such as poly-phosphoinositides (PIs). Since PIs play important roles in many cellular processes, including signaling and membrane trafficking pathways, it has been proposed that CPZ affects cellular growth functions by preventing the recruitment of proteins with specific PI-binding domains. In this study, we have investigated the biological effects of CPZ in the yeast Saccharomyces cerevisiae. We screened a collection of approximately 4,800 gene knockout mutants, and found that mutants defective in membrane trafficking between the late-Golgi and endosomal compartments are highly sensitive to CPZ. Microscopy and transport analyses revealed that CPZ affects membrane structure of organelles, blocks membrane transport and activates the unfolded protein response (UPR). In addition, CPZ-treatment induces phosphorylation of the translation initiation factor (eIF2alpha), which reduces the general rate of protein synthesis and stimulates the production of Gcn4p, a major transcription factor that is activated in response to environmental stresses. Altogether, our results reveal that membrane stress within the cells rapidly activates an important gene expression program, which is followed by a general inhibition of protein synthesis. Remarkably, the increase of phosphorylated eIF2alpha and protein synthesis inhibition were also detected in CPZ-treated NIH-3T3 fibroblasts, suggesting the existence of a conserved mechanism of translational regulation that operates during a membrane stress.

  14. Regulation of the membrane insertion and conductance activity of the metamorphic chloride intracellular channel protein CLIC1 by cholesterol.

    Directory of Open Access Journals (Sweden)

    Stella M Valenzuela

    Full Text Available The Chloride Intracellular ion channel protein CLIC1 has the ability to spontaneously insert into lipid membranes from a soluble, globular state. The precise mechanism of how this occurs and what regulates this insertion is still largely unknown, although factors such as pH and redox environment are known contributors. In the current study, we demonstrate that the presence and concentration of cholesterol in the membrane regulates the spontaneous insertion of CLIC1 into the membrane as well as its ion channel activity. The study employed pressure versus area change measurements of Langmuir lipid monolayer films; and impedance spectroscopy measurements using tethered bilayer membranes to monitor membrane conductance during and following the addition of CLIC1 protein. The observed cholesterol dependent behaviour of CLIC1 is highly reminiscent of the cholesterol-dependent-cytolysin family of bacterial pore-forming proteins, suggesting common regulatory mechanisms for spontaneous protein insertion into the membrane bilayer.

  15. Plasma membrane protein polarity and trafficking in RPE cells: Past, present and future (United States)

    Lehmann, Guillermo L.; Benedicto, Ignacio; Philp, Nancy J.; Rodriguez-Boulan, Enrique


    The retinal pigment epithelium (RPE) comprises a monolayer of polarized pigmented epithelial cells that is strategically interposed between the neural retina and the fenestrated choroid capillaries. The RPE performs a variety of vectorial transport functions (water, ions, metabolites, nutrients and waste products) that regulate the composition of the subretinal space and support the functions of photoreceptors (PRs) and other cells in the neural retina. To this end, RPE cells display a polarized distribution of channels, transporters and receptors in their plasma membrane (PM) that is remarkably different from that found in conventional extra-ocular epithelia, e.g. intestine, kidney, and gall bladder. This characteristic PM protein polarity of RPE cells depends on the interplay of sorting signals in the RPE PM proteins and sorting mechanisms and biosynthetic/recycling trafficking routes in the RPE cell. Although considerable progress has been made in our understanding of the RPE trafficking machinery, most available data have been obtained from immortalized RPE cell lines that only partially maintain the RPE phenotype and by extrapolation of data obtained in the prototype Madin–Darby Canine Kidney (MDCK) cell line. The increasing availability of RPE cell cultures that more closely resemble the RPE in vivo together with the advent of advanced live imaging microscopy techniques provides a platform and an opportunity to rapidly expand our understanding of how polarized protein trafficking contributes to RPE PM polarity. PMID:25152359

  16. Regulation of satellite cell function in sarcopenia

    Directory of Open Access Journals (Sweden)

    Stephen E Alway


    Full Text Available The mechanisms contributing to sarcopenia include reduced satellite cell (myogenic stem cell function that is impacted by the environment (niche of these cells. Satellite cell function is affected by oxidative stress, which is elevated in aged muscles, and this along with changes in largely unknown systemic factors, likely contribute to the manner in which satellite cells respond to stressors such as exercise, disuse or rehabilitation in sarcopenic muscles. Nutritional intervention provides one therapeutic strategy to improve the satellite cell niche and systemic factors, with the goal of improving satellite cell function in aging muscles. Although many elderly persons consume various nutraceuticals with the hope of improving health, most of these compounds have not been thoroughly tested, and the impacts that they might have on sarcopenia, and satellite cell function are not clear. This review discusses data pertaining to the satellite cell responses and function in aging skeletal muscle, and the impact that three compounds: resveratrol, green tea catechins and β-Hydroxy-β-methylbutyrate have on regulating satellite cell function and therefore contributing to reducing sarcopenia or improving muscle mass after disuse in aging. The data suggest that these nutraceutical compounds improve satellite cell function during rehabilitative loading in animal models of aging after disuse (i.e., muscle regeneration. While these compounds have not been rigorously tested in humans, the data from animal models of aging provide a strong basis for conducting additional focused work to determine if these or other nutraceuticals can offset the muscle losses, or improve regeneration in sarcopenic muscles of older humans via improving satellite cell function.

  17. Regulation of Satellite Cell Function in Sarcopenia (United States)

    Alway, Stephen E.; Myers, Matthew J.; Mohamed, Junaith S.


    The mechanisms contributing to sarcopenia include reduced satellite cell (myogenic stem cell) function that is impacted by the environment (niche) of these cells. Satellite cell function is affected by oxidative stress, which is elevated in aged muscles, and this along with changes in largely unknown systemic factors, likely contribute to the manner in which satellite cells respond to stressors such as exercise, disuse, or rehabilitation in sarcopenic muscles. Nutritional intervention provides one therapeutic strategy to improve the satellite cell niche and systemic factors, with the goal of improving satellite cell function in aging muscles. Although many elderly persons consume various nutraceuticals with the hope of improving health, most of these compounds have not been thoroughly tested, and the impacts that they might have on sarcopenia and satellite cell function are not clear. This review discusses data pertaining to the satellite cell responses and function in aging skeletal muscle, and the impact that three compounds: resveratrol, green tea catechins, and β-Hydroxy-β-methylbutyrate have on regulating satellite cell function and therefore contributing to reducing sarcopenia or improving muscle mass after disuse in aging. The data suggest that these nutraceutical compounds improve satellite cell function during rehabilitative loading in animal models of aging after disuse (i.e., muscle regeneration). While these compounds have not been rigorously tested in humans, the data from animal models of aging provide a strong basis for conducting additional focused work to determine if these or other nutraceuticals can offset the muscle losses, or improve regeneration in sarcopenic muscles of older humans via improving satellite cell function. PMID:25295003

  18. Nanoparticle-Mediated Mechanical Destruction of Cell Membranes: A Coarse-Grained Molecular Dynamics Study. (United States)

    Zhang, Liuyang; Zhao, Yiping; Wang, Xianqiao


    The effects of binding mode, shape, binding strength, and rotational speed of actively rotating nanoparticles on the integrity of cell membranes have been systematically studied using dissipative particle dynamics simulations. With theoretical analyses of lipid density, surface tension, stress distribution, and water permeation, we demonstrate that the rotation of nanoparticles can provide a strong driving force for membrane rupture. The results show that nanoparticles embedded inside a cell membrane via endocytosis are more capable of producing large membrane deformations under active rotation than nanoparticles attached on the cell membrane surface. Nanoparticles with anisotropic shapes produce larger deformation and have a higher rupture efficiency than those with symmetric shapes. Our findings provide useful design guidelines for a general strategy based on utilizing mechanical forces to rupture cell membranes and therefore destroy the integrity of cells.

  19. Measurement of the cell membrane capacitance and conductance of colonic crypt cells of the rat using the patch clamp technique

    CERN Document Server

    Schill, C


    Using the patch clamp technique the membrane capacitance and membrane conductance of colonic crypt cells of the rat was measured. The influence of the intracellular agonists Ca++, cAMP and of osmotic changes on the membrane capacitance and conductance was studied.

  20. Bcl-xL regulates mitochondrial energetics by stabilizing the inner membrane potential. (United States)

    Chen, Ying-Bei; Aon, Miguel A; Hsu, Yi-Te; Soane, Lucian; Teng, Xinchen; McCaffery, J Michael; Cheng, Wen-Chih; Qi, Bing; Li, Hongmei; Alavian, Kambiz N; Dayhoff-Brannigan, Margaret; Zou, Shifa; Pineda, Fernando J; O'Rourke, Brian; Ko, Young H; Pedersen, Peter L; Kaczmarek, Leonard K; Jonas, Elizabeth A; Hardwick, J Marie


    Mammalian Bcl-x(L) protein localizes to the outer mitochondrial membrane, where it inhibits apoptosis by binding Bax and inhibiting Bax-induced outer membrane permeabilization. Contrary to expectation, we found by electron microscopy and biochemical approaches that endogenous Bcl-x(L) also localized to inner mitochondrial cristae. Two-photon microscopy of cultured neurons revealed large fluctuations in inner mitochondrial membrane potential when Bcl-x(L) was genetically deleted or pharmacologically inhibited, indicating increased total ion flux into and out of mitochondria. Computational, biochemical, and genetic evidence indicated that Bcl-x(L) reduces futile ion flux across the inner mitochondrial membrane to prevent a wasteful drain on cellular resources, thereby preventing an energetic crisis during stress. Given that F(1)F(O)-ATP synthase directly affects mitochondrial membrane potential and having identified the mitochondrial ATP synthase β subunit in a screen for Bcl-x(L)-binding partners, we tested and found that Bcl-x(L) failed to protect β subunit-deficient yeast. Thus, by bolstering mitochondrial energetic capacity, Bcl-x(L) may contribute importantly to cell survival independently of other Bcl-2 family proteins.

  1. Cell volume regulation: physiology and pathophysiology

    DEFF Research Database (Denmark)

    Lambert, I H; Hoffmann, E K; Pedersen, Stine Helene Falsig


    not only under physiological conditions, e.g. following accumulation of nutrients, during epithelial absorption/secretion processes, following hormonal/autocrine stimulation, and during induction of apoptosis, but also under pathophysiological conditions, e.g. hypoxia, ischaemia and hyponatremia....../hypernatremia. On the other hand, it has recently become clear that an increase or reduction in cell volume can also serve as a specific signal in the regulation of physiological processes such as transepithelial transport, cell migration, proliferation and death. Although the mechanisms by which cell volume perturbations...... are sensed are still far from clear, significant progress has been made with respect to the nature of the sensors, transducers and effectors that convert a change in cell volume into a physiological response. In the present review, we summarize recent major developments in the field, and emphasize...

  2. Inflammatory signals regulate hematopoietic stem cells. (United States)

    Baldridge, Megan T; King, Katherine Y; Goodell, Margaret A


    Hematopoietic stem cells (HSCs) are the progenitors of all blood and immune cells, yet their role in immunity is not well understood. Most studies have focused on the ability of committed lymphoid and myeloid precursors to replenish immune cells during infection. Recent studies, however, have indicated that HSCs also proliferate in response to systemic infection and replenish effector immune cells. Inflammatory signaling molecules including interferons, tumor necrosis factor-α and Toll-like receptors are essential to the HSC response. Observing the biology of HSCs through the lens of infection and inflammation has led to the discovery of an array of immune-mediators that serve crucial roles in HSC regulation and function. Copyright © 2010 Elsevier Ltd. All rights reserved.

  3. Flavonoids: from cell cycle regulation to biotechnology. (United States)

    Woo, Ho-Hyung; Jeong, Byeong Ryong; Hawes, Martha C


    Flavonoids have been proposed to play diverse roles in plant growth and development, including defense, symbiosis, pollen development and male fertility, polar auxin transport, and protection against ultraviolet radiation. Recently, a new role in cell cycle regulation has emerged. Genetic alteration of glucuronide metabolism by altered expression of a Pisum sativum UDP-glucuronosyltransferase (PsUGT1) results in an altered cell cycle in pea, alfalfa, and Arabidopsis. In alfalfa, altered expression of PsUGT1 results in accumulation of a flavonoid-like compound that suppresses growth of cultured cells. The results are consistent with the hypothesis that PsUGT1 functions by controlling cellular levels of a factor controlling cell cycle (FCC).

  4. Cystic Fibrosis Transmembrane Conductance Regulator Attaches Tumor Suppressor PTEN to the Membrane and Promotes Anti Pseudomonas aeruginosa Immunity. (United States)

    Riquelme, Sebastián A; Hopkins, Benjamin D; Wolfe, Andrew L; DiMango, Emily; Kitur, Kipyegon; Parsons, Ramon; Prince, Alice


    The tumor suppressor PTEN controls cell proliferation by regulating phosphatidylinositol-3-kinase (PI3K) activity, but the participation of PTEN in host defense against bacterial infection is less well understood. Anti-inflammatory PI3K-Akt signaling is suppressed in patients with cystic fibrosis (CF), a disease characterized by hyper-inflammatory responses to airway infection. We found that Ptenl -/- mice, which lack the NH 2 -amino terminal splice variant of PTEN, were unable to eradicate Pseudomonas aeruginosa from the airways and could not generate sufficient anti-inflammatory PI3K activity, similar to what is observed in CF. PTEN and the CF transmembrane conductance regulator (CFTR) interacted directly and this interaction was necessary to position PTEN at the membrane. CF patients under corrector-potentiator therapy, which enhances CFTR transport to the membrane, have increased PTEN amounts. These findings suggest that improved CFTR trafficking could enhance P. aeruginosa clearance from the CF airway by activating PTEN-mediated anti-bacterial responses and might represent a therapeutic strategy. Published by Elsevier Inc.

  5. Regulation of polymorphonuclear cell activation by thrombopoietin.


    Brizzi, M F; Battaglia, E; Rosso, A; Strippoli, P; Montrucchio, G; Camussi, G; Pegoraro, L


    Thrombopoietin (TPO) regulates early and late stages of platelet formation as well as platelet activation. TPO exerts its effects by binding to the receptor, encoded by the protooncogene c-mpl, that is expressed in a large number of cells of hematopoietic origin. In this study, we evaluated the expression of c-Mpl and the effects of TPO on human polymorphonuclear cells (PMN). We demonstrate that PMN express the TPO receptor c-Mpl and that TPO induces STAT1 tyrosine phosphorylation and the for...

  6. Analytical model describes ion conduction in fuel cell membranes (United States)

    Herbst, Daniel; Tse, Steve; Witten, Thomas


    Many fuel cell designs employ polyelectrolyte membranes, but little is known about how to tune the parameters (water level, morphology, etc.) to maximize ion conductivity. We came up with a simple model based on a random, discrete water distribution and ion con