Sample records for cell rbc membrane

  1. The study on the parallel processing based time series correlation analysis of RBC membrane flickering in quantitative phase imaging

    NASA Astrophysics Data System (ADS)

    Lee, Minsuk; Won, Youngjae; Park, Byungjun; Lee, Seungrag

    2017-02-01

    Not only static characteristics but also dynamic characteristics of the red blood cell (RBC) contains useful information for the blood diagnosis. Quantitative phase imaging (QPI) can capture sample images with subnanometer scale depth resolution and millisecond scale temporal resolution. Various researches have been used QPI for the RBC diagnosis, and recently many researches has been developed to decrease the process time of RBC information extraction using QPI by the parallel computing algorithm, however previous studies are interested in the static parameters such as morphology of the cells or simple dynamic parameters such as root mean square (RMS) of the membrane fluctuations. Previously, we presented a practical blood test method using the time series correlation analysis of RBC membrane flickering with QPI. However, this method has shown that there is a limit to the clinical application because of the long computation time. In this study, we present an accelerated time series correlation analysis of RBC membrane flickering using the parallel computing algorithm. This method showed consistent fractal scaling exponent results of the surrounding medium and the normal RBC with our previous research.

  2. Red blood cell (RBC) membrane proteomics--Part I: Proteomics and RBC physiology.

    PubMed

    Pasini, Erica M; Lutz, Hans U; Mann, Matthias; Thomas, Alan W

    2010-01-03

    Membrane proteomics is concerned with accurately and sensitively identifying molecules involved in cell compartmentalisation, including those controlling the interface between the cell and the outside world. The high lipid content of the environment in which these proteins are found often causes a particular set of problems that must be overcome when isolating the required material before effective HPLC-MS approaches can be performed. The membrane is an unusually dynamic cellular structure since it interacts with an ever changing environment. A full understanding of this critical cell component will ultimately require, in addition to proteomics, lipidomics, glycomics, interactomics and study of post-translational modifications. Devoid of nucleus and organelles in mammalian species other than camelids, and constantly in motion in the blood stream, red blood cells (RBCs) are the sole mammalian oxygen transporter. The fact that mature mammalian RBCs have no internal membrane-bound organelles, somewhat simplifies proteomics analysis of the plasma membrane and the fact that it has no nucleus disqualifies microarray based methods. Proteomics has the potential to provide a better understanding of this critical interface, and thereby assist in identifying new approaches to diseases. (c) 2009 Elsevier B.V. All rights reserved.

  3. Autoantibodies against mouse bromelain-modified RBC are specifically inhibited by a common membrane phospholipid, phosphatidylcholine.

    PubMed Central

    Cox, K O; Hardy, S J

    1985-01-01

    Sera from mice injected 4 days earlier with lipopolysaccharide lysed mouse RBC treated with bromelain (brom). This lytic activity was totally inhibited by including phosphatidylcholine at final concentrations of about 2 micrograms/ml, or more, in the lytic mixtures. In contrast, the lytic activity of antibodies against rat RBC was not inhibited, even at concentrations of phosphatidylcholine up to 2.5 mg/ml. Various components of the phosphatidylcholine molecule, and other lipids including the closely-related molecule dipalmitoyl phosphatidyl-dimethyl-ethanolamine which is identical to dipalmitoyl phosphatidylcholine, except for the absence of a CH2 group on the polar head group, did not inhibit lysis by the autoantibodies. Autoantibodies against mouse brom RBC, but not antibodies against rat RBC, bound to, and could be eluted from, phosphatidylcholine molecules attached to an insoluble matrix. Liposomes of phosphatidylcholine prepared in the presence of phosphatidic acid or phosphatidylinositol did not inhibit the lysis of mouse brom RBC by autoantibodies to the same extent as liposomes of only phosphatidylcholine. This suggests that phosphatidylcholine is recognized by the autoantibodies only if presented in a certain configuration. We suggest that the function of these autoantibodies may be to facilitate the removal of membrane-damaged cells from the body. Such cells may arise by the process of ageing, or because of the effects of infectious agents such as viruses. PMID:4007927

  4. The Molecular Structure of Human Red Blood Cell Membranes from Highly Oriented, Solid Supported Multi-Lamellar Membranes

    PubMed Central

    Himbert, Sebastian; Alsop, Richard J.; Rose, Markus; Hertz, Laura; Dhaliwal, Alexander; Moran-Mirabal, Jose M.; Verschoor, Chris P.; Bowdish, Dawn M. E.; Kaestner, Lars; Wagner, Christian; Rheinstädter, Maikel C.

    2017-01-01

    We prepared highly oriented, multi-lamellar stacks of human red blood cell (RBC) membranes applied on silicon wafers. RBC ghosts were prepared by hemolysis and applied onto functionalized silicon chips and annealed into multi-lamellar RBC membranes. High resolution X-ray diffraction was used to determine the molecular structure of the stacked membranes. We present direct experimental evidence that these RBC membranes consist of nanometer sized domains of integral coiled-coil peptides, as well as liquid ordered (lo) and liquid disordered (ld) lipids. Lamellar spacings, membrane and hydration water layer thicknesses, areas per lipid tail and domain sizes were determined. The common drug aspirin was added to the RBC membranes and found to interact with RBC membranes and preferably partition in the head group region of the lo domain leading to a fluidification of the membranes, i.e., a thinning of the bilayers and an increase in lipid tail spacing. Our results further support current models of RBC membranes as patchy structures and provide unprecedented structural details of the molecular organization in the different domains. PMID:28045119

  5. The Molecular Structure of Human Red Blood Cell Membranes from Highly Oriented, Solid Supported Multi-Lamellar Membranes

    NASA Astrophysics Data System (ADS)

    Himbert, Sebastian; Alsop, Richard J.; Rose, Markus; Hertz, Laura; Dhaliwal, Alexander; Moran-Mirabal, Jose M.; Verschoor, Chris P.; Bowdish, Dawn M. E.; Kaestner, Lars; Wagner, Christian; Rheinstädter, Maikel C.

    2017-01-01

    We prepared highly oriented, multi-lamellar stacks of human red blood cell (RBC) membranes applied on silicon wafers. RBC ghosts were prepared by hemolysis and applied onto functionalized silicon chips and annealed into multi-lamellar RBC membranes. High resolution X-ray diffraction was used to determine the molecular structure of the stacked membranes. We present direct experimental evidence that these RBC membranes consist of nanometer sized domains of integral coiled-coil peptides, as well as liquid ordered (lo) and liquid disordered (ld) lipids. Lamellar spacings, membrane and hydration water layer thicknesses, areas per lipid tail and domain sizes were determined. The common drug aspirin was added to the RBC membranes and found to interact with RBC membranes and preferably partition in the head group region of the lo domain leading to a fluidification of the membranes, i.e., a thinning of the bilayers and an increase in lipid tail spacing. Our results further support current models of RBC membranes as patchy structures and provide unprecedented structural details of the molecular organization in the different domains.

  6. RBC Distribution Width: Biomarker for Red Cell Dysfunction and Critical Illness Outcome?

    PubMed

    Said, Ahmed S; Spinella, Philip C; Hartman, Mary E; Steffen, Katherine M; Jackups, Ronald; Holubkov, Richard; Wallendorf, Mike; Doctor, Allan

    2017-02-01

    RBC distribution width is reported to be an independent predictor of outcome in adults with a variety of conditions. We sought to determine if RBC distribution width is associated with morbidity or mortality in critically ill children. Retrospective observational study. Tertiary PICU. All admissions to St. Louis Children's Hospital PICU between January 1, 2005, and December 31, 2012. We collected demographics, laboratory values, hospitalization characteristics, and outcomes. We calculated the relative change in RBC distribution width from admission RBC distribution width to the highest RBC distribution width during the first 7 days of hospitalization. Our primary outcome was ICU mortality or use of extracorporeal membrane oxygenation as a composite. Secondary outcomes were ICU- and ventilator-free days. We identified 3,913 eligible subjects with an estimated mortality (by Pediatric Index of Mortality 2) of 2.94% ± 9.25% and an actual ICU mortality of 2.91%. For the study cohort, admission RBC distribution width was 14.12% ± 1.89% and relative change in RBC distribution width was 2.63% ± 6.23%. On univariate analysis, both admission RBC distribution width and relative change in RBC distribution width correlated with mortality or the use of extracorporeal membrane oxygenation (odds ratio, 1.19 [95% CI, 1.12-1.27] and odds ratio, 1.06 [95% CI, 1.04-1.08], respectively; p < 0.001). After adjusting for confounding variables, including severity of illness, both admission RBC distribution width (odds ratio, 1.13; 95% CI, 1.03-1.24) and relative change in RBC distribution width (odds ratio, 1.04; 95% CI, 1.01-1.07) remained independently associated with ICU mortality or the use of extracorporeal membrane oxygenation. Admission RBC distribution width and relative change in RBC distribution width both weakly correlated with fewer ICU- (r = 0.038) and ventilator-free days (r = 0.05) (p < 0.001). Independent of illness severity in critically ill children, admission RBC

  7. The study on RBC characteristic in paroxysmal nocturnal hemoglobinuria (PNH) patients using common path interferometric quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Park, Byung Jun; Won, Youngjae; Kim, Byungyeon; Lee, Seungrag

    2016-03-01

    We have studied the RBC membrane properties between a normal RBC and a RBC in Paroxysrnal nocturnal hemoglobinuria (PNH) patient using common path interferometric quantitative phase microscopy (CPIQPM). CPIQPM system has provided the subnanometer optical path length sensitivity on a millisecond. We have measured the dynamic thickness fluctuations of a normal RBC membrane and a RBC membrane in PNH patient over the whole cell surface with CPIQPM. PNH is a rare and serious disease of blood featured by destruction of red blood cells (RBCs). This destruction happens since RBCs show the defect of protein which protects RBCs from the immune system. We have applied CPIQPM to study the characteristic of RBC membrane in PNH patient. We have shown the morphological shape, volume, and projected surface for both different RBC types. The results have showed both RBCs had the similar shape with donut, but membrane fluctuations in PNH patient was shown to reveal the difference of temporal properties compared with a normal RBC. In order to demonstrate the practical tool of the CPIQPM technique, we have also obtained the time series thickness fluctuation outside a cell.

  8. Diagnostic tool for red blood cell membrane disorders: Assessment of a new generation ektacytometer☆

    PubMed Central

    Da Costa, Lydie; Suner, Ludovic; Galimand, Julie; Bonnel, Amandine; Pascreau, Tiffany; Couque, Nathalie; Fenneteau, Odile; Mohandas, Narla

    2016-01-01

    Inherited red blood cell (RBC) membrane disorders, such as hereditary spherocytosis, elliptocytosis and hereditary ovalocytosis, result from mutations in genes encoding various RBC membrane and skeletal proteins. The RBC membrane, a composite structure composed of a lipid bilayer linked to a spectrin/actin-based membrane skeleton, confers upon the RBC unique features of deformability and mechanical stability. The disease severity is primarily dependent on the extent of membrane surface area loss. RBC membrane disorders can be readily diagnosed by various laboratory approaches that include RBC cytology, flow cytometry, ektacytometry, electrophoresis of RBC membrane proteins and genetics. The reference technique for diagnosis of RBC membrane disorders is the osmotic gradient ektacytometry. However, in spite of its recognition as the reference technique, this technique is rarely used as a routine diagnosis tool for RBC membrane disorders due to its limited availability. This may soon change as a new generation of ektacytometer has been recently engineered. In this review, we describe the workflow of the samples shipped to our Hematology laboratory for RBC membrane disorder analysis and the data obtained for a large cohort of French patients presenting with RBC membrane disorders using a newly available version of the ektacytomer. PMID:26603718

  9. Diagnostic tool for red blood cell membrane disorders: Assessment of a new generation ektacytometer.

    PubMed

    Da Costa, Lydie; Suner, Ludovic; Galimand, Julie; Bonnel, Amandine; Pascreau, Tiffany; Couque, Nathalie; Fenneteau, Odile; Mohandas, Narla

    2016-01-01

    Inherited red blood cell (RBC) membrane disorders, such as hereditary spherocytosis, elliptocytosis and hereditary ovalocytosis, result from mutations in genes encoding various RBC membrane and skeletal proteins. The RBC membrane, a composite structure composed of a lipid bilayer linked to a spectrin/actin-based membrane skeleton, confers upon the RBC unique features of deformability and mechanical stability. The disease severity is primarily dependent on the extent of membrane surface area loss. RBC membrane disorders can be readily diagnosed by various laboratory approaches that include RBC cytology, flow cytometry, ektacytometry, electrophoresis of RBC membrane proteins and genetics. The reference technique for diagnosis of RBC membrane disorders is the osmotic gradient ektacytometry. However, in spite of its recognition as the reference technique, this technique is rarely used as a routine diagnosis tool for RBC membrane disorders due to its limited availability. This may soon change as a new generation of ektacytometer has been recently engineered. In this review, we describe the workflow of the samples shipped to our Hematology laboratory for RBC membrane disorder analysis and the data obtained for a large cohort of French patients presenting with RBC membrane disorders using a newly available version of the ektacytomer. Copyright © 2015 Elsevier Inc. All rights reserved.

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

    PubMed

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

    2014-10-01

    The normal red blood cell (RBC) shape is a biconcave discocyte. An intercalation of a drug in the outer half of the membrane lipid bilayer leads to echinocytosis, an intercalation in the inner half to stomatocytosis. We have used the shape transforming capacity of RBCs as a model to analyse the membrane interaction potential of various neurotropic drugs. Chlorpromazine, clomipramine, citalopram, clonazepam, and diazepam induced a reversible stomatocytosis, phenytoin induced echinocytosis, while the anticonvulsants levetiracetam, valproic acid and phenobarbital had no effect. This diversity of RBC shape transformations suggests that the pharmacological action is not linked to the membrane interaction. We conclude that this simple RBC shape transformation assay could be a useful tool to screen for potential drug interactions with cell membranes. Copyright © 2014. Published by Elsevier Ltd.

  11. Transformation of membrane nanosurface of red blood cells under hemin action

    NASA Astrophysics Data System (ADS)

    Kozlova, Elena; Chernysh, Alexander; Moroz, Victor; Gudkova, Olga; Sergunova, Victoria; Kuzovlev, Artem

    2014-08-01

    Hemin is the product of hemoglobin oxidation. Some diseases may lead to a formation of hemin. The accumulation of hemin causes destruction of red blood cells (RBC) membranes. In this study the process of development of topological defects of RBC membranes within the size range from nanoscale to microscale levels is shown. The formation of the grain-like structures in the membrane (``grains'') with typical sizes of 120-200 nm was experimentally shown. The process of formation of ``grains'' was dependent on the hemin concentration and incubation time. The possible mechanism of membrane nanostructure alterations is proposed. The kinetic equations of formation and transformation of small and medium topological defects were analyzed. This research can be used to study the cell intoxication and analyze the action of various agents on RBC membranes.

  12. Effect of additive solutions on red blood cell (RBC) membrane properties of stored RBCs prepared from whole blood held for 24 hours at room temperature.

    PubMed

    Veale, Margaret F; Healey, Gerry; Sparrow, Rosemary L

    2011-01-01

    The quality of RBC components is influenced by collection, processing and storage conditions. Regulations require that whole blood (WB) units be refrigerated within 8 hours and processed into RBCs within 24 hours of collection. Overnight room temperature hold of WB has logistical advantages, but the effect on RBC quality has not been fully investigated. RBC additive solutions were compared for their ability to provide improved quality of RBCs prepared from WB held at room temperature for 24 hours. Leukocyte-reduced RBCs were prepared from WB held at 20°C on cooling plates for 24 hours prior to processing. RBCs were stored in additive solutions, SAG-M (control), Erythrosol-4, and PAGGSM, under standard blood banking conditions and sampled during 49 days of storage. Stored RBCs were evaluated for RBC shape and microparticle (MP) accumulation using flow cytometry. Osmotic fragility, adhesion of RBCs to endothelium under shear stress conditions (0.5 dyne/cm(2) ), and routine RBC quality parameters were assessed. RBCs stored in Erythrosol-4 and PAGGSM had decreased cell size, reduced osmotic fragility, and decreased accumulation of glycophorin A-positive MPs and annexin V-binding MPs compared with RBCs stored in SAG-M. RBCs stored in erythrosol-4 had increased adherence to endothelium at days 42 and 49 compared with RBCs stored in SAG-M or PAGGSM. RBCs stored in PAGGSM or Erythrosol-4 had improved retention of RBC membrane and osmotic resilience. The development of new additive solutions may offer improved quality of RBC components prepared from WB held overnight at room temperature. © 2010 American Association of Blood Banks.

  13. Myosin IIA interacts with the spectrin-actin membrane skeleton to control red blood cell membrane curvature and deformability.

    PubMed

    Smith, Alyson S; Nowak, Roberta B; Zhou, Sitong; Giannetto, Michael; Gokhin, David S; Papoin, Julien; Ghiran, Ionita C; Blanc, Lionel; Wan, Jiandi; Fowler, Velia M

    2018-05-08

    The biconcave disk shape and deformability of mammalian RBCs rely on the membrane skeleton, a viscoelastic network of short, membrane-associated actin filaments (F-actin) cross-linked by long, flexible spectrin tetramers. Nonmuscle myosin II (NMII) motors exert force on diverse F-actin networks to control cell shapes, but a function for NMII contractility in the 2D spectrin-F-actin network of RBCs has not been tested. Here, we show that RBCs contain membrane skeleton-associated NMIIA puncta, identified as bipolar filaments by superresolution fluorescence microscopy. MgATP disrupts NMIIA association with the membrane skeleton, consistent with NMIIA motor domains binding to membrane skeleton F-actin and contributing to membrane mechanical properties. In addition, the phosphorylation of the RBC NMIIA heavy and light chains in vivo indicates active regulation of NMIIA motor activity and filament assembly, while reduced heavy chain phosphorylation of membrane skeleton-associated NMIIA indicates assembly of stable filaments at the membrane. Treatment of RBCs with blebbistatin, an inhibitor of NMII motor activity, decreases the number of NMIIA filaments associated with the membrane and enhances local, nanoscale membrane oscillations, suggesting decreased membrane tension. Blebbistatin-treated RBCs also exhibit elongated shapes, loss of membrane curvature, and enhanced deformability, indicating a role for NMIIA contractility in promoting membrane stiffness and maintaining RBC biconcave disk cell shape. As structures similar to the RBC membrane skeleton exist in many metazoan cell types, these data demonstrate a general function for NMII in controlling specialized membrane morphology and mechanical properties through contractile interactions with short F-actin in spectrin-F-actin networks.

  14. Red blood cell membrane water permeability increases with length of ex vivo storage.

    PubMed

    Alshalani, Abdulrahman; Acker, Jason P

    2017-06-01

    Water transport across the red blood cell (RBC) membrane is an essential cell function that needs to be preserved during ex vivo storage. Progressive biochemical depletion during storage can result in significant conformational and compositional changes to the membrane. Characterizing the changes to RBC water permeability can help in evaluating the quality of stored blood products and aid in the development of improved methods for the cryopreservation of red blood cells. This study aimed to characterize the water permeability (L p ), osmotically inactive fraction (b), and Arrhenius activation energy (E a ) at defined storage time-points throughout storage and to correlate the observed results with other in vitro RBC quality parameters. RBCs were collected from age- and sex-matched blood donors. A stopped flow spectrophotometer was used to determine L p and b by monitoring changes in hemoglobin autofluorescence when RBCs were exposed to anisotonic solutions. Experimental values of L p were characterized at three different temperatures (4, 20 and 37 °C) to determine the E a . Results showed that L p , b, and E a of stored RBCs significantly increase by day 21 of storage. Degradation of the RBC membrane with length of storage was seen as an increase in hemolysis and supernatant potassium, and a decrease in deformability, mean corpuscular hemoglobin concentration and supernatant sodium. RBC osmotic characteristics were shown to change with storage and correlate with changes in RBC membrane quality metrics. Monitoring water parameters is a predictor of membrane damage and loss of membrane integrity in ex vivo stored RBCs. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Investigation of membrane mechanics using spring networks: application to red-blood-cell modelling.

    PubMed

    Chen, Mingzhu; Boyle, Fergal J

    2014-10-01

    In recent years a number of red-blood-cell (RBC) models have been proposed using spring networks to represent the RBC membrane. Some results predicted by these models agree well with experimental measurements. However, the suitability of these membrane models has been questioned. The RBC membrane, like a continuum membrane, is mechanically isotropic throughout its surface, but the mechanical properties of a spring network vary on the network surface and change with deformation. In this work spring-network mechanics are investigated in large deformation for the first time via an assessment of the effect of network parameters, i.e. network mesh, spring type and surface constraint. It is found that a spring network is conditionally equivalent to a continuum membrane. In addition, spring networks are employed for RBC modelling to replicate the optical tweezers test. It is found that a spring network is sufficient for modelling the RBC membrane but strain-hardening springs are required. Moreover, the deformation profile of a spring network is presented for the first time via the degree of shear. It is found that spring-network deformation approaches continuous as the mesh density increases. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. Continuous Change in Membrane and Membrane-Skeleton Organization During Development From Proerythroblast to Senescent Red Blood Cell

    PubMed Central

    Minetti, Giampaolo; Achilli, Cesare; Perotti, Cesare; Ciana, Annarita

    2018-01-01

    Within the context of erythropoiesis and the possibility of producing artificial red blood cells (RBCs) in vitro, a most critical step is the final differentiation of enucleated erythroblasts, or reticulocytes, to a fully mature biconcave discocyte, the RBC. Reviewed here is the current knowledge about this fundamental maturational process. By combining literature data with our own experimental evidence we propose that the early phase in the maturation of reticulocytes to RBCs is driven by a membrane raft-based mechanism for the sorting of disposable membrane proteins, mostly the no longer needed transferrin receptor (TfR), to the multivesicular endosome (MVE) as cargo of intraluminal vesicles that are subsequently exocytosed as exosomes, consistently with the seminal and original observation of Johnstone and collaborators of more than 30 years ago (Pan BT, Johnstone RM. Cell. 1983;33:967-978). According to a strikingly selective sorting process, the TfR becomes cargo destined to exocytosis while other molecules, including the most abundant RBC transmembrane protein, band 3, are completely retained in the cell membrane. It is also proposed that while this process could be operating in the early maturational steps in the bone marrow, additional mechanism(s) must be at play for the final removal of the excess reticulocyte membrane that is observed to occur in the circulation. This processing will most likely require the intervention of the spleen, whose function is also necessary for the continuous remodeling of the RBC membrane all along this cell's circulatory life. PMID:29632498

  17. Red blood cell membrane viscoelasticity, agglutination and zeta potential measurements with double optical tweezers

    NASA Astrophysics Data System (ADS)

    Fontes, Adriana; Fernandes, Heloise P.; Barjas-Castro, Maria L.; de Thomaz, André A.; de Ysasa Pozzo, Liliana; Barbosa, Luiz C.; Cesar, Carlos L.

    2006-02-01

    The red blood cell (RBC) viscoelastic membrane contains proteins and glycolproteins embedded in, or attached, to a fluid lipid bilayer and are negatively charged, which creates a repulsive electric (zeta) potential between the cells and prevents their aggregation in the blood stream. There are techniques, however, to decrease the zeta potential to allow cell agglutination which are the basis of most of the tests of antigen-antibody interactions in blood banks. This report shows the use of a double optical tweezers to measure RBC membrane viscosity, agglutination and zeta potential. In our technique one of the optical tweezers trap a silica bead that binds strongly to a RBC at the end of a RBCs rouleaux and, at the same time, acts as a pico-Newton force transducer, after calibration through its displacement from the equilibrium position. The other optical tweezers trap the RBC at the other end. To measure the membrane viscosity the optical force is measured as a function of the velocity between the RBCs. To measure the adhesion the tweezers are slowly displaced apart until the RBCs disagglutination happens. The RBC zeta potential is measured in two complimentary ways, by the force on the silica bead attached to a single RBC in response to an applied electric field, and the conventional way, by the measurement of terminal velocity of the RBC after released from the optical trap. These two measurements provide information about the RBC charges and, also, electrolytic solution properties. We believe this can improve the methods of diagnosis in blood banks.

  18. Measuring dynamic membrane fluctuations in cell membrane using quantitative phase imaging (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Lee, SangYun; Kim, Kyoohyun; Park, YongKeun

    2017-02-01

    There is a strong correlation between the dynamic membrane fluctuations and the biomechanical properties of living cells. The dynamic membrane fluctuation consists of submicron displacements, and can be altered by changing the cells' pathophysiological conditions. These results have significant relevance to the understanding of RBC biophysics and pathology, as follows. RBCs must withstand large mechanical deformations during repeated passages through the microvasculature and the fenestrated walls of the splenic sinusoids. This essential ability is diminished with senescence, resulting in physiological destruction of the aging RBCs. Pathological destruction of the red cells, however, occurs in cells affected by a host of diseases such as spherocytosis, malaria, and Sickle cell disease, as RBCs depart from their normal discoid shape and lose their deformability. Therefore, quantifying the RBC deformability insight into a variety of problems regarding the interplay of cell structure, dynamics, and function. Furthermore, the ability to monitor mechanical properties of RBCs is of vital interest in monitoring disease progression or response to treatment as molecular and pharmaceutical approaches for treatment of chronic diseases. Here, we present the measurements of dynamic membrane fluctuations in live cells using quantitative phase imaging techniques. Measuring both the 3-D refractive index maps and the dynamic phase images of live cells are simultaneously measured, from which dynamic membrane fluctuation and deformability of cells are precisely calculated. We also present its applications to various diseases ranging from sickle cell diseases, babesiosis, and to diabetes.

  19. Red blood cell membrane-camouflaged melanin nanoparticles for enhanced photothermal therapy.

    PubMed

    Jiang, Qin; Luo, Zimiao; Men, Yongzhi; Yang, Peng; Peng, Haibao; Guo, Ranran; Tian, Ye; Pang, Zhiqing; Yang, Wuli

    2017-10-01

    Photothermal therapy (PTT) has represented a promising noninvasive approach for cancer treatment in recent years. However, there still remain challenges in developing non-toxic and biodegradable biomaterials with high photothermal efficiency in vivo. Herein, we explored natural melanin nanoparticles extracted from living cuttlefish as effective photothermal agents and developed red blood cell (RBC) membrane-camouflaged melanin (Melanin@RBC) nanoparticles as a platform for in vivo antitumor PTT. The as-obtained natural melanin nanoparticles demonstrated strong absorption at NIR region, higher photothermal conversion efficiency (∼40%) than synthesized melanin-like polydopamine nanoparticles (∼29%), as well as favorable biocompatibility and biodegradability. It was shown that RBC membrane coating on melanin nanoparticles retained their excellent photothermal property, enhanced their blood retention and effectively improved their accumulation at tumor sites. With the guidance of their inherited photoacoustic imaging capability, optimal accumulation of Melanin@RBC at tumors was achieved around 4 h post intravenous injection. Upon irradiation by an 808-nm laser, the developed Melanin@RBC nanoparticles exhibited significantly higher PTT efficacy than that of bare melanin nanoparticles in A549 tumor-bearing mice. Given that both melanin nanoparticles and RBC membrane are native biomaterials, the developed Melanin@RBC platform could have great potential in clinics for anticancer PTT. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. The effects of membrane cholesterol and simvastatin on red blood cell deformability and ATP release.

    PubMed

    Forsyth, Alison M; Braunmüller, Susanne; Wan, Jiandi; Franke, Thomas; Stone, Howard A

    2012-05-01

    It is known that deformation of red blood cells (RBCs) is linked to ATP release from the cells. Further, membrane cholesterol has been shown to alter properties of the cell membrane such as fluidity and bending stiffness. Membrane cholesterol content is increased in some cardiovascular diseases, for example, in individuals with acute coronary syndromes and chronic stable angina, and therefore, because of the potential clinical relevance, we investigated the influence of altered RBC membrane cholesterol levels on ATP release. Because of the correlation between statins and reduced membrane cholesterol in vivo, we also investigated the effects of simvastatin on RBC deformation and ATP release. We found that reducing membrane cholesterol increases cell deformability and ATP release. We also found that simvastatin increases deformability by acting directly on the membrane in the absence of the liver, and that ATP release was increased for cells with enriched cholesterol after treatment with simvastatin. Copyright © 2012 Elsevier Inc. All rights reserved.

  1. Dynamic actin filaments control the mechanical behavior of the human red blood cell membrane

    PubMed Central

    Gokhin, David S.; Nowak, Roberta B.; Khoory, Joseph A.; de la Piedra, Alfonso; Ghiran, Ionita C.; Fowler, Velia M.

    2015-01-01

    Short, uniform-length actin filaments function as structural nodes in the spectrin-actin membrane skeleton to optimize the biomechanical properties of red blood cells (RBCs). Despite the widespread assumption that RBC actin filaments are not dynamic (i.e., do not exchange subunits with G-actin in the cytosol), this assumption has never been rigorously tested. Here we show that a subpopulation of human RBC actin filaments is indeed dynamic, based on rhodamine-actin incorporation into filaments in resealed ghosts and fluorescence recovery after photobleaching (FRAP) analysis of actin filament mobility in intact RBCs (∼25–30% of total filaments). Cytochalasin-D inhibition of barbed-end exchange reduces rhodamine-actin incorporation and partially attenuates FRAP recovery, indicating functional interaction between actin subunit turnover at the single-filament level and mobility at the membrane-skeleton level. Moreover, perturbation of RBC actin filament assembly/disassembly with latrunculin-A or jasplakinolide induces an approximately twofold increase or ∼60% decrease, respectively, in soluble actin, resulting in altered membrane deformability, as determined by alterations in RBC transit time in a microfluidic channel assay, as well as by abnormalities in spontaneous membrane oscillations (flickering). These experiments identify a heretofore-unrecognized but functionally important subpopulation of RBC actin filaments, whose properties and architecture directly control the biomechanical properties of the RBC membrane. PMID:25717184

  2. IgG red blood cell autoantibodies in autoimmune hemolytic anemia bind to epitopes on red blood cell membrane band 3 glycoprotein

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Victoria, E.J.; Pierce, S.W.; Branks, M.J.

    1990-01-01

    Red blood cell (RBC) autoantibodies from patients with IgG warm-type autoimmune hemolytic anemia were labeled with iodine 125 and their RBC binding behavior characterized. Epitope-bearing RBC membrane polypeptides were identified after autoantibody immunoprecipitation of labeled membranes and immunoblotting. Immunoaffinity isolation of labeled membrane proteins with 12 different IgG hemolytic autoantibodies with protein A-agarose revealed a major polypeptide at Mr 95 to 110 kd, which coelectrophoresed on sodium dodecylsulfate-polyacrylamide gel electrophoresis with a membrane component isolated with sheep IgG anti-band 3. Immunoprecipitation studies with chymotrypsinized RBCs resulted in the recovery of two labeled membrane polypeptides with molecular weights characteristically resulting frommore » the chymotryptic fragmentation of band 3. Immunoblotting with sheep IgG anti-band 3 of the immunoprecipitated polypeptides confirmed that hemolytic autoantibody binding led to recovery of band 3 or its fragments. Two 125I-labeled IgG hemolytic autoantibodies showed binding behavior consistent with epitope localization on band 3. The labeled RBC autoantibodies bound immunospecifically to all types of human RBC tested, including those of rare Rh type (Rh-null, D--) at a site density of approximately 10(6) per RBC. The 125I-IgG in two labeled autoantibodies was 84% and 92% adsorbable by human and higher nonhuman primate RBCs. Antigen-negative animal RBC bound less than 10%, consistent with immunospecific RBC binding. IgG-1 was the major subclass in five autoantibodies tested; one of six fixed complement; and autoantibody IgG appeared polyclonal by isoelectric focusing. We conclude that IgG eluted from RBCs of patients with autoimmune hemolytic anemia consists predominantly of a single totally RBC-adsorbable antibody population that binds to antigenic determinants on band 3.« less

  3. Inconsistencies in the red blood cell membrane proteome analysis: generation of a database for research and diagnostic applications

    PubMed Central

    Hegedűs, Tamás; Chaubey, Pururawa Mayank; Várady, György; Szabó, Edit; Sarankó, Hajnalka; Hofstetter, Lia; Roschitzki, Bernd; Sarkadi, Balázs

    2015-01-01

    Based on recent results, the determination of the easily accessible red blood cell (RBC) membrane proteins may provide new diagnostic possibilities for assessing mutations, polymorphisms or regulatory alterations in diseases. However, the analysis of the current mass spectrometry-based proteomics datasets and other major databases indicates inconsistencies—the results show large scattering and only a limited overlap for the identified RBC membrane proteins. Here, we applied membrane-specific proteomics studies in human RBC, compared these results with the data in the literature, and generated a comprehensive and expandable database using all available data sources. The integrated web database now refers to proteomic, genetic and medical databases as well, and contains an unexpected large number of validated membrane proteins previously thought to be specific for other tissues and/or related to major human diseases. Since the determination of protein expression in RBC provides a method to indicate pathological alterations, our database should facilitate the development of RBC membrane biomarker platforms and provide a unique resource to aid related further research and diagnostics. Database URL: http://rbcc.hegelab.org PMID:26078478

  4. A study of the dynamic properties of the human red blood cell membrane using quasi-elastic light-scattering spectroscopy.

    PubMed

    Tishler, R B; Carlson, F D

    1993-12-01

    A quasi-elastic light-scattering (QELS) microscope spectrometer was used to study the dynamic properties of the membrane/cytoskeleton of individual human red blood cells (RBCs). QELS is a spectroscopic technique that measures intensity fluctuations of laser light scattered from a sample. The intensity fluctuations were analyzed using power spectra and the intensity autocorrelation function, g(2)(tau), which was approximated with a single exponential. The value of the correlation time, Tcorr, was used for comparing results. Motion of the RBC membrane/cytoskeleton was previously identified as the source of the QELS signal from the RBC (R. B. Tishler and F. D. Carlson, 1987. Biophys. J. 51:993-997), and additional data supporting that conclusion are presented. Similar results were obtained from anucleate mammalian RBCs that have structures similar to that of the human RBC, but not for morphologically distinct, nucleated RBCs. The effect of altering the physical properties of the cytoplasm and the membrane/cytoskeleton was also studied. Osmotically increasing the cytoplasmic viscosity led to significant increases in Tcorr. Increasing the membrane cholesterol content and increasing the intracellular calcium content both led to decreased deformability of the human RBC. In both cases, the modified cells with decreased deformability showed an increase in Tcorr, demonstrating that QELS could measure biochemically induced changes of the membrane/cytoskeleton. Physiological changes were measured in studies of age-separated RBC populations which showed that Tcorr was increased in the older, less deformable cells.

  5. Refractive index tomograms and dynamic membrane fluctuations of red blood cells from patients with diabetes mellitus.

    PubMed

    Lee, SangYun; Park, HyunJoo; Kim, Kyoohyun; Sohn, YongHak; Jang, Seongsoo; Park, YongKeun

    2017-04-21

    In this paper, we present the optical characterisations of diabetic red blood cells (RBCs) in a non-invasive manner employing three-dimensional (3-D) quantitative phase imaging. By measuring 3-D refractive index tomograms and 2-D time-series phase images, the morphological (volume, surface area and sphericity), biochemical (haemoglobin concentration and content) and mechanical (membrane fluctuation) parameters were quantitatively retrieved at the individual cell level. With simultaneous measurements of individual cell properties, systematic correlative analyses on retrieved RBC parameters were also performed. Our measurements show there exist no statistically significant alterations in morphological and biochemical parameters of diabetic RBCs, compared to those of healthy (non-diabetic) RBCs. In contrast, membrane deformability of diabetic RBCs is significantly lower than that of healthy, non-diabetic RBCs. Interestingly, non-diabetic RBCs exhibit strong correlations between the elevated glycated haemoglobin in RBC cytoplasm and decreased cell deformability, whereas diabetic RBCs do not show correlations. Our observations strongly support the idea that slow and irreversible glycation of haemoglobin and membrane proteins of RBCs by hyperglycaemia significantly compromises RBC deformability in diabetic patients.

  6. Red Blood Cell Membrane as a Biomimetic Nanocoating for Prolonged Circulation Time and Reduced Accelerated Blood Clearance.

    PubMed

    Rao, Lang; Bu, Lin-Lin; Xu, Jun-Hua; Cai, Bo; Yu, Guang-Tao; Yu, Xiaolei; He, Zhaobo; Huang, Qinqin; Li, Andrew; Guo, Shi-Shang; Zhang, Wen-Feng; Liu, Wei; Sun, Zhi-Jun; Wang, Hao; Wang, Tza-Huei; Zhao, Xing-Zhong

    2015-12-01

    For decades, poly(ethylene glycol) (PEG) has been widely incorporated into nanoparticles for evading immune clearance and improving the systematic circulation time. However, recent studies have reported a phenomenon known as "accelerated blood clearance (ABC)" where a second dose of PEGylated nanomaterials is rapidly cleared when given several days after the first dose. Herein, we demonstrate that natural red blood cell (RBC) membrane is a superior alternative to PEG. Biomimetic RBC membrane-coated Fe(3)O(4) nanoparticles (Fe(3)O(4) @RBC NPs) rely on CD47, which is a "don't eat me" marker on the RBC surface, to escape immune clearance through interactions with the signal regulatory protein-alpha (SIRP-α) receptor. Fe(3)O(4) @RBC NPs exhibit extended circulation time and show little change between the first and second doses, with no ABC suffered. In addition, the administration of Fe(3)O(4) @RBC NPs does not elicit immune responses on neither the cellular level (myeloid-derived suppressor cells (MDSCs)) nor the humoral level (immunoglobulin M and G (IgM and IgG)). Finally, the in vivo toxicity of these cell membrane-camouflaged nanoparticles is systematically investigated by blood biochemistry, hematology testing, and histology analysis. These findings are significant advancements toward solving the long-existing clinical challenges of developing biomaterials that are able to resist both immune response and rapid clearance. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Decreased content of protein 4.2 in ankyrin-deficient normoblastosis (nb/nb) mouse red blood cells: evidence for ankyrin enhancement of protein 4.2 membrane binding.

    PubMed

    Rybicki, A C; Musto, S; Schwartz, R S

    1995-11-01

    Homozygous normoblastosis (nb/nb) mice, whose red blood cell (RBC) membranes are nearly completely deficient in full-length 210-kD ankyrin, were used to study interactions between ankyrin and protein 4.2 (P4.2). Although it is unclear whether or not these proteins interact in the membrane, both ankyrin and P4.2 bind to the cytoplasmic domain of band 3 (cdb3). In addition to the complete deficiency of full-length ankyrin, nb/nb RBC membranes are also partially spectrin deficient, resulting in morphologically spherocytic and mechanically fragile cells. A new finding was that nb/nb RBC membranes are severely (approximately 73%) P4.2 deficient compared with wild type (+/+) or high reticulocyte mouse RBC membranes. Metabolic labeling of nb/nb reticulocytes showed active P4.2 synthesis at levels comparable with high reticulocyte controls suggesting that the nb/nb P4.2 deficiency was not the result of defective P4.2 synthesis. Reconstitution of nb/nb inside-out vesicles (IOVs) with human RBC ankyrin restored ankyrin levels to approximately 80% of +/+ IOV levels and increased binding of exogenously added human RBC P4.2 by approximately 60%. These results suggest that ankyrin is required for normal associations of P4.2 with the RBC membrane.

  8. The Effect of Alcohols on Red Blood Cell Mechanical Properties and Membrane Fluidity Depends on Their Molecular Size

    PubMed Central

    Sonmez, Melda; Ince, Huseyin Yavuz; Yalcin, Ozlem; Ajdžanović, Vladimir; Spasojević, Ivan; Meiselman, Herbert J.; Baskurt, Oguz K.

    2013-01-01

    The role of membrane fluidity in determining red blood cell (RBC) deformability has been suggested by a number of studies. The present investigation evaluated alterations of RBC membrane fluidity, deformability and stability in the presence of four linear alcohols (methanol, ethanol, propanol and butanol) using ektacytometry and electron paramagnetic resonance (EPR) spectroscopy. All alcohols had a biphasic effect on deformability such that it increased then decreased with increasing concentration; the critical concentration for reversal was an inverse function of molecular size. EPR results showed biphasic changes of near-surface fluidity (i.e., increase then decrease) and a decreased fluidity of the lipid core; rank order of effectiveness was butanol > propanol > ethanol > methanol, with a significant correlation between near-surface fluidity and deformability (r = 0.697; p<0.01). The presence of alcohol enhanced the impairment of RBC deformability caused by subjecting cells to 100 Pa shear stress for 300 s, with significant differences from control being observed at higher concentrations of all four alcohols. The level of hemolysis was dependent on molecular size and concentration, whereas echinocytic shape transformation (i.e., biconcave disc to crenated morphology) was observed only for ethanol and propanol. These results are in accordance with available data obtained on model membranes. They document the presence of mechanical links between RBC deformability and near-surface membrane fluidity, chain length-dependence of the ability of alcohols to alter RBC mechanical behavior, and the biphasic response of RBC deformability and near-surface membrane fluidity to increasing alcohol concentrations. PMID:24086751

  9. The effect of alcohols on red blood cell mechanical properties and membrane fluidity depends on their molecular size.

    PubMed

    Sonmez, Melda; Ince, Huseyin Yavuz; Yalcin, Ozlem; Ajdžanović, Vladimir; Spasojević, Ivan; Meiselman, Herbert J; Baskurt, Oguz K

    2013-01-01

    The role of membrane fluidity in determining red blood cell (RBC) deformability has been suggested by a number of studies. The present investigation evaluated alterations of RBC membrane fluidity, deformability and stability in the presence of four linear alcohols (methanol, ethanol, propanol and butanol) using ektacytometry and electron paramagnetic resonance (EPR) spectroscopy. All alcohols had a biphasic effect on deformability such that it increased then decreased with increasing concentration; the critical concentration for reversal was an inverse function of molecular size. EPR results showed biphasic changes of near-surface fluidity (i.e., increase then decrease) and a decreased fluidity of the lipid core; rank order of effectiveness was butanol > propanol > ethanol > methanol, with a significant correlation between near-surface fluidity and deformability (r = 0.697; p<0.01). The presence of alcohol enhanced the impairment of RBC deformability caused by subjecting cells to 100 Pa shear stress for 300 s, with significant differences from control being observed at higher concentrations of all four alcohols. The level of hemolysis was dependent on molecular size and concentration, whereas echinocytic shape transformation (i.e., biconcave disc to crenated morphology) was observed only for ethanol and propanol. These results are in accordance with available data obtained on model membranes. They document the presence of mechanical links between RBC deformability and near-surface membrane fluidity, chain length-dependence of the ability of alcohols to alter RBC mechanical behavior, and the biphasic response of RBC deformability and near-surface membrane fluidity to increasing alcohol concentrations.

  10. Podocalyxin Is a Glycoprotein Ligand of the Human Pluripotent Stem Cell-Specific Probe rBC2LCN

    PubMed Central

    Tateno, Hiroaki; Matsushima, Asako; Hiemori, Keiko; Onuma, Yasuko; Ito, Yuzuru; Hasehira, Kayo; Nishimura, Ken; Ohtaka, Manami; Takayasu, Satoko; Nakanishi, Mahito; Ikehara, Yuzuru; Nakanishi, Mio; Ohnuma, Kiyoshi; Chan, Techuan; Toyoda, Masashi; Akutsu, Hidenori; Umezawa, Akihiro; Asashima, Makoto

    2013-01-01

    In comprehensive glycome analysis with a high-density lectin microarray, we have previously shown that the recombinant N-terminal domain of the lectin BC2L-C from Burkholderia cenocepacia (rBC2LCN) binds exclusively to undifferentiated human induced pluripotent stem (iPS) cells and embryonic stem (ES) cells but not to differentiated somatic cells. Here we demonstrate that podocalyxin, a heavily glycosylated type 1 transmembrane protein, is a glycoprotein ligand of rBC2LCN on human iPS cells and ES cells. When analyzed by DNA microarray, podocalyxin was found to be highly expressed in both iPS cells and ES cells. Western and lectin blotting revealed that rBC2LCN binds to podocalyxin with a high molecular weight of more than 240 kDa in undifferentiated iPS cells of six different origins and four ES cell lines, but no binding was observed in either differentiated mouse feeder cells or somatic cells. The specific binding of rBC2LCN to podocalyxin prepared from a large set of iPS cells (138 types) and ES cells (15 types) was also confirmed using a high-throughput antibody-overlay lectin microarray. Alkaline digestion greatly reduced the binding of rBC2LCN to podocalyxin, indicating that the major glycan ligands of rBC2LCN are presented on O-glycans. Furthermore, rBC2LCN was found to exhibit significant affinity to a branched O-glycan comprising an H type 3 structure (Ka, 2.5 × 104 M−1) prepared from human 201B7 iPS cells, indicating that H type 3 is a most probable potential pluripotency marker. We conclude that podocalyxin is a glycoprotein ligand of rBC2LCN on human iPS cells and ES cells. PMID:23526252

  11. Integrin-associated protein (CD47) is a putative mediator for soluble fibrinogen interaction with human red blood cells membrane.

    PubMed

    De Oliveira, S; Vitorino de Almeida, V; Calado, A; Rosário, H S; Saldanha, C

    2012-03-01

    Fibrinogen is a multifunctional plasma protein that plays a crucial role in several biological processes. Elevated fibrinogen induces erythrocyte hyperaggregation, suggesting an interaction between this protein and red blood cells (RBCs). Several studies support the concept that fibrinogen interacts with RBC membrane and this binding, due to specific and non-specific mechanisms, may be a trigger to RBC hyperaggregation in inflammation. The main goals of our work were to prove that human RBCs are able to specifically bind soluble fibrinogen, and identify membrane molecular targets that could be involved in this process. RBCs were first isolated from blood of healthy individuals and then separated in different age fractions by discontinuous Percoll gradients. After isolation RBC samples were incubated with human soluble fibrinogen and/or with a blocking antibody against CD47 followed by fluorescence confocal microscopy, flow cytometry acquisitions and zeta potential measurements. Our data show that soluble fibrinogen interacts with the human RBC membrane in an age-dependent manner, with younger RBCs interacting more with soluble fibrinogen than the older cells. Importantly, this interaction is abrogated in the presence of a specific antibody against CD47. Our results support a specific and age-dependent interaction of soluble fibrinogen with human RBC membrane; additionally we present CD47 as a putative mediator in this process. This interaction may contribute to RBC hyperaggregation in inflammation. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Studying red blood cell agglutination by measuring membrane viscosity with optical tweezers

    NASA Astrophysics Data System (ADS)

    Fernandes, Heloise P.; Fontes, Adriana; de Thomaz, André A.; Barbosa, Luiz C.; Barjas-Castro, Maria L.; Cesar, Carlos L.

    2007-09-01

    The red blood cell (RBC) viscoelastic membrane contains proteins and glycoproteins embedded in a fluid lipid bilayer that are responsible for cell agglutination. Manipulating RBCs rouleaux with a double optical tweezers, we observed that the cells slide easily one over the others but are strongly connected by their edges. An explanation for this behavior could be the fact that when the cells slide one over the others, proteins are dragged through the membrane. It confers to the movement a viscous characteristic that is dependent of the velocity between the RBCs and justifies why is so easy to slide them apart. Therefore, in a first step of this work, by measuring the force as a function of the relative velocity between two cells, we confirmed this assumption and used this viscous characteristic of the RBC rouleaux to determine the apparent membrane viscosity of the cell. As this behavior is related to the proteins interactions, we can use the apparent membrane viscosity to obtain a better understanding about cell agglutination. Methods related to cell agglutination induced by antigen-antibody interactions are the basis of most of tests used in transfusion centers. Then, in a second step of this work, we measured the apparent membrane viscosity using antibodies. We observed that this methodology is sensitive to different kinds of bindings between RBCs. Better comprehension of the forces and bindings between RBCs could improve the sensibility and specificity of the hemagglutination reactions and also guides the development of new potentiator substances.

  13. Storage duration and white blood cell content of red blood cell (RBC) products increases adhesion of stored RBCs to endothelium under flow conditions.

    PubMed

    Anniss, Angela M; Sparrow, Rosemary L

    2006-09-01

    Adherence of red blood cells (RBCs) to vascular endothelium impairs blood flow and decreases oxygen delivery. Although RBCs may be stored for up to 42 days before transfusion under current blood banking guidelines, little is known of how changes to RBCs during storage may affect their adherence properties. The influence of RBC product storage time and white blood cell (WBC) burden on the adherence of RBCs for transfusion to vascular endothelium under conditions of continuous flow was investigated in this study. RBC samples were collected from nonleukoreduced (S-RBC), buffy coat-poor (BCP-RBC), and leukofiltered (LF-RBC) products at fixed time points during storage. Samples were perfused, at controlled shear stress and temperature, across a confluent endothelial cell (EC) monolayer with a parallel-flow chamber mounted to an inverted microscope. RBC-EC interactions were recorded with a digital camera attached to the microscope. The number of RBCs adhering to the EC layer increased significantly with storage time in all RBC products; however, WBC reduction delayed this increase. LF-RBCs were also significantly less adherent than S-RBC or BCP-RBC products on Day 1 of storage (p < 0.05). The strength of RBC attachment to vascular endothelium was significantly stronger in S-RBC products compared to BCP-RBC and LF-RBC products. Our findings indicate that product storage time and WBC burden increase the number and strength of adhesion of RBCs to vascular endothelium. These results may lead to greater understanding of the interaction of transfused RBCs with recipient endothelium and the biologic consequences of this adherence.

  14. Eliminating Xenoantigen Expression on Swine RBC.

    PubMed

    Wang, Zheng-Yu; Martens, Gregory R; Blankenship, Ross L; Sidner, Richard A; Li, Ping; Estrada, Jose L; Tector, Matthew; Tector, A Joseph

    2017-03-01

    The rapidly improving tools of genetic engineering may make it possible to overcome the humoral immune barrier that prevents xenotransplantation. We hypothesize that levels of human antibody binding to donor tissues from swine must approximate the antibody binding occurring in allotransplantation. It is uncertain if this is an attainable goal. Here we perform an initial analysis of this issue by comparing human antibody binding to red blood cells (RBC) isolated from knockout swine and to allogeneic or autologous human RBC. Human sera were incubated with RBC isolated from various genetically engineered swine or from humans. The level of IgG and IgM binding to these cells were compared using either flow cytometry or a novel mass spectrometric assay. Mass spectroscopic quantitation of human antibody binding demonstrated that as few as 3 gene inactivations can reduce the levels human antibody binding to swine RBC that is as low as autologous human RBC. Flow cytometry showed that RBC from 2-gene knockout swine exhibited less human antibody binding than human blood group O allogeneic RBC in 22% of tested sera. Deletion of a third gene from pigs resulted in 30% of human samples having less IgG and IgM RBC xenoreactivity than alloreactivity. Xenoantigenicity of swine RBC can be eliminated via gene disruption. These results suggest that the gene knockout approach may be able reduce antigenicity in other pig tissues to levels that enable the xenotransplantation humoral barrier to be overcome.

  15. Measurement of RBC agglutination with microscopic cell image analysis in a microchannel chip.

    PubMed

    Cho, Chi Hyun; Kim, Ju Yeon; Nyeck, Agnes E; Lim, Chae Seung; Hur, Dae Sung; Chung, Chanil; Chang, Jun Keun; An, Seong Soo A; Shin, Sehyun

    2014-01-01

    Since Landsteiner's discovery of ABO blood groups, RBC agglutination has been one of the most important immunohematologic techniques for ABO and RhD blood groupings. The conventional RBC agglutination grading system for RhD blood typings relies on macroscopic reading, followed by the assignment of a grade ranging from (-) to (4+) to the degree of red blood cells clumping. However, with the new scoring method introduced in this report, microscopically captured cell images of agglutinated RBCs, placed in a microchannel chip, are used for analysis. Indeed, the cell images' pixel number first allows the differentiation of agglutinated and non-agglutinated red blood cells. Finally, the ratio of agglutinated RBCs per total RBC counts (CRAT) from 90 captured images is then calculated. During the trial, it was observed that the agglutinated group's CRAT was significantly higher (3.77-0.003) than that of the normal control (0). Based on these facts, it was established that the microchannel method was more suitable for the discrimination between agglutinated RBCs and non-agglutinated RhD negative, and thus more reliable for the grading of RBCs agglutination than the conventional method.

  16. Spectrin-ankyrin interaction mechanics: A key force balance factor in the red blood cell membrane skeleton.

    PubMed

    Saito, Masakazu; Watanabe-Nakayama, Takahiro; Machida, Shinichi; Osada, Toshiya; Afrin, Rehana; Ikai, Atsushi

    2015-01-01

    As major components of red blood cell (RBC) cytoskeleton, spectrin and F-actin form a network that covers the entire cytoplasmic surface of the plasma membrane. The cross-linked two layered structure, called the membrane skeleton, keeps the structural integrity of RBC under drastically changing mechanical environment during circulation. We performed force spectroscopy experiments on the atomic force microscope (AFM) as a means to clarify the mechanical characteristics of spectrin-ankyrin interaction, a key factor in the force balance of the RBC cytoskeletal structure. An AFM tip was functionalized with ANK1-62k and used to probe spectrin crosslinked to mica surface. A force spectroscopy study gave a mean unbinding force of ~30 pN under our experimental conditions. Two energy barriers were identified in the unbinding process. The result was related to the well-known flexibility of spectrin tetramer and participation of ankyrin 1-spectrin interaction in the overall balance of membrane skeleton dynamics. Copyright © 2015 Elsevier B.V. All rights reserved.

  17. Headspace gas chromatography of volatile lipid peroxidation products from human red blood cell membranes.

    PubMed

    Frankel, E N; Tappel, A L

    1991-06-01

    An improved headspace capillary gas chromatographic (GC) method was developed to measure the oxidative susceptibility of human red blood cell (RBC) membranes. This method analyzed volatile peroxidation products of both n-6 (hexanal and pentane) and n-3 (propanal) polyunsaturated fatty acids. Oxidative susceptibility tests were standardized by incubating in a sealed 10-mL headspace bottle 0.25 or 1 mL of human RBC membrane in 40 mM phosphate buffer for 1 hr at 37 degrees C with a mixture of Fe++, ascorbic acid and H2O2. Sodium dodecyl sulfate increased significantly the amount of hexanal measured by headspace GC. By this standard headspace method, in one series of red blood cell membranes (RBCM) samples a four-fold variation in oxidative susceptibility was observed in RBCM from blood freshly drawn from six healthy subjects. In another series of RBCM samples a sixteen-fold variation in oxidative susceptibility was noted in frozen RBCM from blood freshly drawn from five healthy subjects. Correlation between hexanal formation and polyunsaturated fatty acids (PUFA) depletion provided good evidence that under these standard conditions hexanal is exclusively derived from the oxidation of arachidonic acid. Hydroperoxides of arachidonic acid are more readily formed and decomposed than those of linoleic acid in the presence of Fe++, ascorbic acid and H2O2 to produce hexanal as the main product that can be readily analyzed by headspace GC. This method may provide a useful tool to study susceptibility toward lipid peroxidative damage in human RBC membranes.

  18. Structural and mechanical heterogeneity of the erythrocyte membrane reveals hallmarks of membrane stability.

    PubMed

    Picas, Laura; Rico, Félix; Deforet, Maxime; Scheuring, Simon

    2013-02-26

    The erythrocyte membrane, a metabolically regulated active structure that comprises lipid molecules, junctional complexes, and the spectrin network, enables the cell to undergo large passive deformations when passing through the microvascular system. Here we use atomic force microscopy (AFM) imaging and quantitative mechanical mapping at nanometer resolution to correlate structure and mechanics of key components of the erythrocyte membrane, crucial for cell integrity and function. Our data reveal structural and mechanical heterogeneity modulated by the metabolic state at unprecedented nanometer resolution. ATP-depletion, reducing skeletal junction phosphorylation in RBC cells, leads to membrane stiffening. Analysis of ghosts and shear-force opened erythrocytes show that, in the absence of cytosolic kinases, spectrin phosphorylation results in membrane stiffening at the extracellular face and a reduced junction remodeling in response to loading forces. Topography and mechanical mapping of single components at the cytoplasmic face reveal that, surprisingly, spectrin phosphorylation by ATP softens individual filaments. Our findings suggest that, besides the mechanical signature of each component, the RBC membrane mechanics is regulated by the metabolic state and the assembly of its structural elements.

  19. RBC micromotors carrying multiple cargos towards potential theranostic applications

    NASA Astrophysics Data System (ADS)

    Wu, Zhiguang; Esteban-Fernández de Ávila, Berta; Martín, Aída; Christianson, Caleb; Gao, Weiwei; Thamphiwatana, Soracha Kun; Escarpa, Alberto; He, Qiang; Zhang, Liangfang; Wang, Joseph

    2015-08-01

    Red blood cell (RBC)-based micromotors containing both therapeutic and diagnostic modalities are described as a means for potential theranostic applications. In this natural RBC-based multicargo-loaded micromotor system, quantum dots (QDs), anti-cancer drug doxorubicin (DOX), and magnetic nanoparticles (MNPs), were co-encapsulated into RBC micromotors. The fluorescent emission of both QDs and DOX provides direct visualization of their loading inside the RBC motors at two distinct wavelengths. The presence of MNPs within the RBCs allows for efficient magnetic guidance under ultrasound propulsion along with providing the potential for magnetic resonance imaging. The simultaneous encapsulation of the imaging nanoparticles and therapeutic payloads within the same RBC micromotor has a minimal effect upon its propulsion behavior. The ability of the RBC micromotors to transport imaging and therapeutic agents at high speed and spatial precision through a complex microchannel network is also demonstrated. Such ability to load and transport diagnostic imaging agents and therapeutic drugs within a single cell-based motor, in addition to a lower toxicity observed once the drug is encapsulated within the multicargo RBC motor, opens the door to the development of theranostic micromotors that may simultaneously treat and monitor diseases.Red blood cell (RBC)-based micromotors containing both therapeutic and diagnostic modalities are described as a means for potential theranostic applications. In this natural RBC-based multicargo-loaded micromotor system, quantum dots (QDs), anti-cancer drug doxorubicin (DOX), and magnetic nanoparticles (MNPs), were co-encapsulated into RBC micromotors. The fluorescent emission of both QDs and DOX provides direct visualization of their loading inside the RBC motors at two distinct wavelengths. The presence of MNPs within the RBCs allows for efficient magnetic guidance under ultrasound propulsion along with providing the potential for magnetic

  20. Erythrocyte membrane-coated gold nanocages for targeted photothermal and chemical cancer therapy

    NASA Astrophysics Data System (ADS)

    Zhu, Dao-Ming; Xie, Wei; Xiao, Yu-Sha; Suo, Meng; Zan, Ming-Hui; Liao, Qing-Quan; Hu, Xue-Jia; Chen, Li-Ben; Chen, Bei; Wu, Wen-Tao; Ji, Li-Wei; Huang, Hui-Ming; Guo, Shi-Shang; Zhao, Xing-Zhong; Liu, Quan-Yan; Liu, Wei

    2018-02-01

    Recently, red blood cell (RBC) membrane-coated nanoparticles have attracted much attention because of their excellent immune escapability; meanwhile, gold nanocages (AuNs) have been extensively used for cancer therapy due to their photothermal effect and drug delivery capability. The combination of the RBC membrane coating and AuNs may provide an effective approach for targeted cancer therapy. However, few reports have shown the utilization of combining these two technologies. Here, we design erythrocyte membrane-coated gold nanocages for targeted photothermal and chemical cancer therapy. First, anti-EpCam antibodies were used to modify the RBC membranes to target 4T1 cancer cells. Second, the antitumor drug paclitaxel (PTX) was encapsulated into AuNs. Then, the AuNs were coated with the modified RBC membranes. These new nanoparticles were termed EpCam-RPAuNs. We characterized the capability of the EpCam-RPAuNs for selective tumor targeting via exposure to near-infrared irradiation. The experimental results demonstrate that EpCam-RPAuNs can effectively generate hyperthermia and precisely deliver the antitumor drug PTX to targeted cells. We also validated the biocompatibility of the EpCam-RAuNs in vitro. By combining the molecularly modified targeting RBC membrane and AuNs, our approach provides a new way to design biomimetic nanoparticles to enhance the surface functionality of nanoparticles. We believe that EpCam-RPAuNs can be potentially applied for cancer diagnoses and therapies.

  1. Elastic behavior of a red blood cell with the membrane's nonuniform natural state: equilibrium shape, motion transition under shear flow, and elongation during tank-treading motion.

    PubMed

    Tsubota, Ken-Ichi; Wada, Shigeo; Liu, Hao

    2014-08-01

    Direct numerical simulations of the mechanics of a single red blood cell (RBC) were performed by considering the nonuniform natural state of the elastic membrane. A RBC was modeled as an incompressible viscous fluid encapsulated by an elastic membrane. The in-plane shear and area dilatation deformations of the membrane were modeled by Skalak constitutive equation, while out-of-plane bending deformation was formulated by the spring model. The natural state of the membrane with respect to in-plane shear deformation was modeled as a sphere ([Formula: see text]), biconcave disk shape ([Formula: see text]) and their intermediate shapes ([Formula: see text]) with the nonuniformity parameter [Formula: see text], while the natural state with respect to out-of-plane bending deformation was modeled as a flat plane. According to the numerical simulations, at an experimentally measured in-plane shear modulus of [Formula: see text] and an out-of-plane bending rigidity of [Formula: see text] of the cell membrane, the following results were obtained. (i) The RBC shape at equilibrium was biconcave discoid for [Formula: see text] and cupped otherwise; (ii) the experimentally measured fluid shear stress at the transition between tumbling and tank-treading motions under shear flow was reproduced for [Formula: see text]; (iii) the elongation deformation of the RBC during tank-treading motion from the simulation was consistent with that from in vitro experiments, irrespective of the [Formula: see text] value. Based on our RBC modeling, the three phenomena (i), (ii), and (iii) were mechanically consistent for [Formula: see text]. The condition [Formula: see text] precludes a biconcave discoid shape at equilibrium (i); however, it gives appropriate fluid shear stress at the motion transition under shear flow (ii), suggesting that a combined effect of [Formula: see text] and the natural state with respect to out-of-plane bending deformation is necessary for understanding details of the

  2. Cholesterol:phospholipid ratio is elevated in platelet plasma membrane in patients with hypertension.

    PubMed

    Benjamin, N; Robinson, B F; Graham, J G; Wilson, R B

    1990-06-01

    The cholesterol:phospholipid ratio was measured in platelet plasma membrane, red blood cell (RBC) membranes, low density lipoprotein (LDL) and whole plasma in patients with primary hypertension and in matched normal controls. The cholesterol:phospholipid ratio was raised in the platelet membrane from hypertensive patients compared with that from normal controls (0.65 +/- 0.03 vs 0.53 +/- 0.02: mean +/- SEM; P less than 0.01). The ratio observed in RBC membranes, LDL and whole blood was similar in the two groups. If this abnormality in the lipid composition of platelet plasma membrane is present in other cells it could account for some of the changes in cell membrane function that have been described in hypertension.

  3. Following-up changes in red blood cell deformability and membrane stability in the presence of PTFE graft implanted into the femoral artery in a canine model

    NASA Astrophysics Data System (ADS)

    Toth, Csaba; Kiss, Ferenc; Klarik, Zoltan; Gergely, Eszter; Toth, Eniko; Peto, Katalin; Vanyolos, Erzsebet; Miko, Iren; Nemeth, Norbert

    2014-05-01

    It is known that a moderate mechanical stress can even improve the red blood cells' (RBC) micro-rheological characteristics, however, a more significant stress causes deterioration in the deformability. In this study, we aimed to investigate the effect of the presence of artificial graft on the RBC deformability and membrane stability in beagles. In the Control group only anesthesia was induced and in the postoperative (p.o.) period blood samplings were carried out. In the Grafted group under general anesthesia, the left femoral artery was isolated, from which a 3.5 cm segment was resected and a PTFE graft (O.D.: 3 mm) of equal in length was implanted into the gap. On the 1st, 3rd, 5th, 7th and 14th p.o. days blood was collected the cephalic veins and RBC deformability was determined ektacytometry (LoRRca MaxSis Osmoscan). Membrane stability test consisted of two deformability measurements before and after the cells were being exposed to mechanical stress (60 or 100 Pa for 300 seconds). Compared to the Control group and the baseline values the red blood cell deformability showed significant deterioration on the 3rd, 5th and mainly on the 7th postoperative day after the graft implantation. The membrane stability of erythrocyte revealed marked inter-group difference on the 3rd, 5th and 7th day: in the Grafted group the deformability decreased and during the membrane stability test smaller difference was observed between the states before and after shearing. We concluded that the presence of a PTFE graft in the femoral artery may cause changes in RBC deformability in the first p.o. week. RBC membrane stability investigation shows a lower elongation index profile for the grafted group and a narrowed alteration in the deformability curves due to mechanical stress.

  4. RBC micromotors carrying multiple cargos towards potential theranostic applications.

    PubMed

    Wu, Zhiguang; Esteban-Fernández de Ávila, Berta; Martín, Aída; Christianson, Caleb; Gao, Weiwei; Thamphiwatana, Soracha Kun; Escarpa, Alberto; He, Qiang; Zhang, Liangfang; Wang, Joseph

    2015-08-28

    Red blood cell (RBC)-based micromotors containing both therapeutic and diagnostic modalities are described as a means for potential theranostic applications. In this natural RBC-based multicargo-loaded micromotor system, quantum dots (QDs), anti-cancer drug doxorubicin (DOX), and magnetic nanoparticles (MNPs), were co-encapsulated into RBC micromotors. The fluorescent emission of both QDs and DOX provides direct visualization of their loading inside the RBC motors at two distinct wavelengths. The presence of MNPs within the RBCs allows for efficient magnetic guidance under ultrasound propulsion along with providing the potential for magnetic resonance imaging. The simultaneous encapsulation of the imaging nanoparticles and therapeutic payloads within the same RBC micromotor has a minimal effect upon its propulsion behavior. The ability of the RBC micromotors to transport imaging and therapeutic agents at high speed and spatial precision through a complex microchannel network is also demonstrated. Such ability to load and transport diagnostic imaging agents and therapeutic drugs within a single cell-based motor, in addition to a lower toxicity observed once the drug is encapsulated within the multicargo RBC motor, opens the door to the development of theranostic micromotors that may simultaneously treat and monitor diseases.

  5. Immunoproteomics of Plasmodium falciparum-infected red blood cell membrane fractions

    PubMed Central

    Cabral, Fernanda J; Vianna, Luciana G; Medeiros, Marcia M; Carlos, Bianca Cechetto; Martha, Rosimeire D; Silva, Nadia Maria; da Silva, Luiz Hildebrando P; Stabeli, Rodrigo G; Wunderlich, Gerhard

    2017-01-01

    BACKGROUND The surface of infected red blood cells (iRBCs) has been widely investigated because of the molecular complexity and pathogenesis mechanisms involved. Asymptomatic individuals are important in the field because they can perpetuate transmission as natural reservoirs and present a challenge for diagnosing malaria because of their low levels of circulating parasites. Recent studies of iRBC antibody recognition have shown that responses are quantitatively similar in symptomatic and asymptomatic infections, but no studies have characterised the plasmodial proteins targeted by this response. OBJECTIVES Our main objective was to identify Plasmodium falciparum proteins associated with iRBC ghosts recognised by antibodies in the sera of symptomatic and asymptomatic individuals in the Brazilian Amazon. METHODS We collected symptomatic and asymptomatic sera from patients residing in the Brazilian Amazon and P. falciparum iRBC ghosts to identify the proteins involved in natural antibody recognition by 2D-electrophoresis, western blotting, and high- resolution mass spectrometry. FINDINGS 2D gel-based immunoproteome analysis using symptomatic and asymptomatic sera identified 11 proteins with at least one unique peptide, such as chaperones HSP70-1 and HSP70-x, which likely are components of the secretion machinery/PTEX translocon. PfEMP1 is involved in antigenic variation in symptomatic infections and we found putative membrane proteins whose functions are unknown. MAIN FINDINGS Our results suggest a potential role of old and new proteins, such as antigenic variation proteins, iRBC remodelling, and membrane proteins, with no assigned functions related to the immune response against P. falciparum, providing insights into the pathogenesis, erythrocyte remodelling, and secretion machinery important for alternative diagnosis and/or malaria therapy. PMID:29211247

  6. Dietary supplementation with docosahexanoic acid (DHA) increases red blood cell membrane flexibility in mice with sickle cell disease.

    PubMed

    Wandersee, Nancy J; Maciaszek, Jamie L; Giger, Katie M; Hanson, Madelyn S; Zheng, Suilan; Guo, YiHe; Mickelson, Barbara; Hillery, Cheryl A; Lykotrafitis, George; Low, Philip S; Hogg, Neil

    2015-02-01

    Humans and mice with sickle cell disease (SCD) have rigid red blood cells (RBCs). Omega-3 fatty acids, such as docosahexanoic acid (DHA), may influence RBC deformability via incorporation into the RBC membrane. In this study, sickle cell (SS) mice were fed natural ingredient rodent diets supplemented with 3% DHA (DHA diet) or a control diet matched in total fat (CTRL diet). After 8weeks of feeding, we examined the RBCs for: 1) stiffness, as measured by atomic force microscopy; 2) deformability, as measured by ektacytometry; and 3) percent irreversibly sickled RBCs on peripheral blood smears. Using atomic force microscopy, it is found that stiffness is increased and deformability decreased in RBCs from SS mice fed CTRL diet compared to wild-type mice. In contrast, RBCs from SS mice fed DHA diet had markedly decreased stiffness and increased deformability compared to RBCs from SS mice fed CTRL diet. Furthermore, examination of peripheral blood smears revealed less irreversibly sickled RBCs in SS mice fed DHA diet as compared to CTRL diet. In summary, our findings indicate that DHA supplementation improves RBC flexibility and reduces irreversibly sickled cells by 40% in SS mice. These results point to potential therapeutic benefits of dietary omega-3 fatty acids in SCD. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. Liposomes composed of unsaturated lipids for membrane modification of human erythrocytes.

    PubMed

    Stoll, Christoph; Holovati, Jelena L; Acker, Jason P; Wolkers, Willem F

    2011-01-01

    Previous studies have shown that certain saturated lipids protect red blood cells (RBCs) during hypothermic storage but provide little protection during freezing or freeze-drying, whereas various unsaturated lipids destabilize RBCs during hypothermic storage but protect during freezing and freeze-drying. The protective effect of liposomes has been attributed to membrane modifications. We have previously shown that cholesterol exchange and lipid transfer between liposomes composed of saturated lipids and RBCs critically depends on the length of the lipid acyl chains. In this study the effect of unsaturated lipids with differences in their number of unsaturated bonds (18:0/18:1, 18:1/18:1, 18:2/18:2) on RBC membrane properties has been studied. RBCs were incubated in the presence of liposomes and both the liposomal and RBC fraction were analyzed by Fourier transform infrared spectroscopy (FTIR) after incubation. The liposomes caused an increase in RBC membrane conformational disorder at suprazero temperatures. The fluidizing effect of the liposomes on the RBC membranes, however, was found to be similar for the different lipids irrespective of their unsaturation level. The gel to liquid crystalline phase transition temperature of the liposomes increased after incubation with RBCs. RBC membrane fluidity increased linearly during the first 8 hours of incubation in the presence of liposomes. The increase in RBC membrane fluidity was found to be temperature dependent and displayed Arrhenius behaviour between 20 and 40°C, with an activation energy of 88 kJ mol⁻¹. Taken together, liposomes composed of unsaturated lipids increase RBC membrane conformational disorder, which could explain their cryoprotective action.

  8. Apatite nanoparticles strongly improve red blood cell cryopreservation by mediating trehalose delivery via enhanced membrane permeation.

    PubMed

    Stefanic, Martin; Ward, Kevin; Tawfik, Harvey; Seemann, Ralf; Baulin, Vladimir; Guo, Yachong; Fleury, Jean-Baptiste; Drouet, Christophe

    2017-09-01

    Cryopreservation of red blood cells (RBC) is an important method for maintaining an inventory of rare RBC units and managing special transfusion circumstances. Currently, in a clinical setting, glycerol is used as cryoprotectant against freezing damage. After thawing and before transfusion, glycerol must however be removed to avoid intravascular hemolysis, via a complex and time-consuming deglycerolization process which requires specialized equipment. Improved cryopreservation methods using non-toxic agents are required to increase biocompatibility and decrease processing time. Biocompatible cryoprotectants (e.g. trehalose) were proposed, but their low permeation through RBC membranes limits their cryoprotection efficacy. Herein, we report for the first time a glycerol-free cryopreservation approach, using colloidal bioinspired apatite nanoparticles (NP) as bioactive promoters of RBC cryopreservation mediated by trehalose. Addition of apatite NP in the medium tremendously increases RBC cryosurvival, up to 91% (42% improvement compared to a control without NP) which is comparable to FDA-approved cryoprotection protocol employing glycerol. NP concentration and incubation conditions strongly modulate the NP bioactivity. Complementary experimental and computational analyses of the interaction between apatite NP and model lipid bilayers revealed complex events occurring at the NP-bilayer interface. Apatite NP do not cross the bilayer but momentarily modulate its physical status. These changes affect the membrane behavior, and promote the permeation of trehalose and a model fluorescent molecule (FITC). This approach is a new alternative to using toxic glycerol for cells cryopreservation, and the identification of this enhancing no-pore permeation mechanism of apatite NP appears as an original delivery pathway for cryoprotectant agents and beyond. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Metabolomics of AS-1 RBC storage

    PubMed Central

    Roback, John D.; Josephson, Cassandra D.; Waller, Edmund K.; Newman, James L.; Karatela, Sulaiman; Uppal, Karan; Jones, Dean; Zimring, James C.; Dumont, Larry J.

    2014-01-01

    Background Population based investigations suggest that red blood cells (RBCs) are therapeutically effective when collected, processed and stored for up to 42 days under validated conditions prior to transfusion. However, some retrospective clinical studies have shown worse patient outcomes when transfused RBCs have been stored for the longest times. Furthermore, studies of RBC persistence in the circulation after transfusion have suggested that considerable donor-to-donor variability exists, and may affect transfusion efficacy. To understand the limitations of current blood storage technologies and to develop approaches to improve RBC storage and transfusion efficacy, we investigated the global metabolic alterations that occur when RBCs are stored in AS-1 (AS1-RBC). Methods Leukoreduced AS1-RBC units prepared from 9 volunteer research donors (12 total donated units) were serially sampled for metabolomics analysis over 42 days of refrigerated storage. Samples were tested by GC/MS and LC/MS/MS, and specific biochemical compounds were identified by comparison to a library of purified standards. Results Over three experiments, 185–264 defined metabolites were quantified in stored RBC samples. Kinetic changes in these biochemicals confirmed known alterations in glycolysis and other pathways previously identified in RBCs stored in SAGM (SAGM-RBC). Furthermore, we identified additional alterations not previously seen in SAGM-RBCs (e.g., stable pentose phosphate pathway flux, progressive decreases in oxidized glutathione), and we delineated changes occurring in other metabolic pathways not previously studied (e.g., S-adenosyl methionine cycle). These data are presented in the context of a detailed comparison with previous studies of SAGM-RBCs from human donors and murine AS1-RBCs. Conclusion Global metabolic profiling of AS1-RBCs revealed a number of biochemical alterations in stored blood that may affect RBC viability during storage as well as therapeutic effectiveness

  10. Accurate measurement of peripheral blood mononuclear cell concentration using image cytometry to eliminate RBC-induced counting error.

    PubMed

    Chan, Leo Li-Ying; Laverty, Daniel J; Smith, Tim; Nejad, Parham; Hei, Hillary; Gandhi, Roopali; Kuksin, Dmitry; Qiu, Jean

    2013-02-28

    Peripheral blood mononuclear cells (PBMCs) have been widely researched in the fields of immunology, infectious disease, oncology, transplantation, hematological malignancy, and vaccine development. Specifically, in immunology research, PBMCs have been utilized to monitor concentration, viability, proliferation, and cytokine production from immune cells, which are critical for both clinical trials and biomedical research. The viability and concentration of isolated PBMCs are traditionally measured by manual counting with trypan blue (TB) using a hemacytometer. One of the common issues of PBMC isolation is red blood cell (RBC) contamination. The RBC contamination can be dependent on the donor sample and/or technical skill level of the operator. RBC contamination in a PBMC sample can introduce error to the measured concentration, which can pass down to future experimental assays performed on these cells. To resolve this issue, RBC lysing protocol can be used to eliminate potential error caused by RBC contamination. In the recent years, a rapid fluorescence-based image cytometry system has been utilized for bright-field and fluorescence imaging analysis of cellular characteristics (Nexcelom Bioscience LLC, Lawrence, MA). The Cellometer image cytometry system has demonstrated the capability of automated concentration and viability detection in disposable counting chambers of unpurified mouse splenocytes and PBMCs stained with acridine orange (AO) and propidium iodide (PI) under fluorescence detection. In this work, we demonstrate the ability of Cellometer image cytometry system to accurately measure PBMC concentration, despite RBC contamination, by comparison of five different total PBMC counting methods: (1) manual counting of trypan blue-stained PBMCs in hemacytometer, (2) manual counting of PBMCs in bright-field images, (3) manual counting of acetic acid lysing of RBCs with TB-stained PBMCs, (4) automated counting of acetic acid lysing of RBCs with PI-stained PBMCs

  11. Defected red blood cell membranes and direct correlation with the uraemic milieu: the connection with the decreased red blood cell lifespan observed in haemodialysis patients

    NASA Astrophysics Data System (ADS)

    Stamopoulos, D.; Grapsa, E.; Manios, E.; Gogola, V.; Bakirtzi, N.

    2012-12-01

    Together with impaired production of erythropoietin and iron deficiency, the decreased lifespan of red blood cells (RBCs) is a main factor contributing to the chronic anaemia observed in haemodialysis (HD) patients. Atomic force microscopy is employed in this work to thoroughly survey the membrane of intact RBCs (iRBCs) of HD patients in comparison to those of healthy donors, aiming to obtain direct information on the structural status of RBCs that can be related to their decreased lifespan. We observed that the iRBC membrane of the HD patients is overpopulated with extended circular defects, termed ‘orifices’, that have typical dimension ranging between 0.2 and 1.0 μm. The ‘orifice’ index—that is, the mean population of ‘orifices’ per top membrane surface—exhibits a pronounced relative increase of order 54 ± 12% for the HD patients as compared to healthy donors. Interestingly, for the HD patients, the ‘orifice’ index, which relates to the structural status of the RBC membrane, correlates strongly with urea concentration, which is a basic index of the uraemic milieu. Thus, these results indicate that the uraemic milieu downgrades the structural status of the RBC membrane, possibly triggering biochemical processes that result in their premature elimination from the circulation. This process could decrease the lifespan of RBCs, as observed in HD patients.

  12. MPP1 directly interacts with flotillins in erythrocyte membrane - Possible mechanism of raft domain formation.

    PubMed

    Biernatowska, Agnieszka; Augoff, Katarzyna; Podkalicka, Joanna; Tabaczar, Sabina; Gajdzik-Nowak, Weronika; Czogalla, Aleksander; Sikorski, Aleksander F

    2017-11-01

    Flotillins are prominent, oligomeric protein components of erythrocyte (RBC) membrane raft domains and are considered to play an important structural role in lateral organization of the plasma membrane. In our previous work on erythroid membranes and giant plasma membrane vesicles (GPMVs) derived from them we have shown that formation of functional domains (resting state rafts) depends on the presence of membrane palmitoylated protein 1 (MPP1/p55), pointing to its new physiological role. Exploration of the molecular mechanism of MPP1 function in organizing membrane domains described here, through searching for its molecular partners in RBC membrane by using different methods, led to the identification of the raft-marker proteins, flotillin 1 and flotillin 2, as hitherto unreported direct MPP1 binding-partners in the RBC membrane. These proteins are found in high molecular-weight complexes in native RBC membrane and, significantly, their presence was shown to be separate from the well-known protein 4.1-dependent interactions of MPP1 with membrane proteins. Furthermore, FLIM analysis revealed that loss of the endogenous MPP1-flotillins interactions resulted in significant changes in RBC membrane-fluidity, emphasizing the physiological importance of such interactions in vivo. Therefore, our data establish a new perspective on the role of MPP1 in erythroid cells and suggests that direct MPP1-flotillins interactions could be the major driving-force behind the formation of raft domains in RBC. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

  13. Physiologic Impact of Circulating RBC Microparticles upon Blood-Vascular Interactions

    PubMed Central

    Said, Ahmed S.; Rogers, Stephen C.; Doctor, Allan

    2018-01-01

    Here, we review current data elucidating the role of red blood cell derived microparticles (RMPs) in normal vascular physiology and disease progression. Microparticles (MPs) are submicron-size, membrane-encapsulated vesicles derived from various parent cell types. MPs are produced in response to numerous stimuli that promote a sequence of cytoskeletal and membrane phospholipid changes and resulting MP genesis. MPs were originally considered as potential biomarkers for multiple disease processes and more recently are recognized to have pleiotropic biological effects, most notably in: promotion of coagulation, production and handling of reactive oxygen species, immune modulation, angiogenesis, and in initiating apoptosis. RMPs, specifically, form normally during RBC maturation in response to injury during circulation, and are copiously produced during processing and storage for transfusion. Notably, several factors during RBC storage are known to trigger RMP production, including: increased intracellular calcium, increased potassium leakage, and energy failure with ATP depletion. Of note, RMP composition differs markedly from that of intact RBCs and the nature/composition of RMP components are affected by the specific circumstances of RMP genesis. Described RMP bioactivities include: promotion of coagulation, immune modulation, and promotion of endothelial adhesion as well as influence upon vasoregulation via influence upon nitric oxide (NO) bioavailability. Of particular relevance, RMPs scavenge NO more avidly than do intact RBCs; this physiology has been proposed to contribute to the impaired oxygen delivery homeostasis that may be observed following transfusion. In summary, RMPs are submicron particles released from RBCs, with demonstrated vasoactive properties that appear to disturb oxygen delivery homeostasis. The clinical impact of RMPs in normal and patho-physiology and in transfusion recipients is an area of continued investigation. PMID:29379445

  14. Physiologic Impact of Circulating RBC Microparticles upon Blood-Vascular Interactions.

    PubMed

    Said, Ahmed S; Rogers, Stephen C; Doctor, Allan

    2017-01-01

    Here, we review current data elucidating the role of red blood cell derived microparticles (RMPs) in normal vascular physiology and disease progression. Microparticles (MPs) are submicron-size, membrane-encapsulated vesicles derived from various parent cell types. MPs are produced in response to numerous stimuli that promote a sequence of cytoskeletal and membrane phospholipid changes and resulting MP genesis. MPs were originally considered as potential biomarkers for multiple disease processes and more recently are recognized to have pleiotropic biological effects, most notably in: promotion of coagulation, production and handling of reactive oxygen species, immune modulation, angiogenesis, and in initiating apoptosis. RMPs, specifically, form normally during RBC maturation in response to injury during circulation, and are copiously produced during processing and storage for transfusion. Notably, several factors during RBC storage are known to trigger RMP production, including: increased intracellular calcium, increased potassium leakage, and energy failure with ATP depletion. Of note, RMP composition differs markedly from that of intact RBCs and the nature/composition of RMP components are affected by the specific circumstances of RMP genesis. Described RMP bioactivities include: promotion of coagulation, immune modulation, and promotion of endothelial adhesion as well as influence upon vasoregulation via influence upon nitric oxide (NO) bioavailability. Of particular relevance, RMPs scavenge NO more avidly than do intact RBCs; this physiology has been proposed to contribute to the impaired oxygen delivery homeostasis that may be observed following transfusion. In summary, RMPs are submicron particles released from RBCs, with demonstrated vasoactive properties that appear to disturb oxygen delivery homeostasis. The clinical impact of RMPs in normal and patho-physiology and in transfusion recipients is an area of continued investigation.

  15. Synthetic nanoparticles camouflaged with biomimetic erythrocyte membranes for reduced reticuloendothelial system uptake

    NASA Astrophysics Data System (ADS)

    Rao, Lang; Xu, Jun-Hua; Cai, Bo; Liu, Huiqin; Li, Ming; Jia, Yan; Xiao, Liang; Guo, Shi-Shang; Liu, Wei; Zhao, Xing-Zhong

    2016-02-01

    Suppression of the reticuloendothelial system (RES) uptake is one of the most challenging tasks in nanomedicine. Coating stratagems using polymers, such as poly(ethylene glycol) (PEG), have led to great success in this respect. Nevertheless, recent observations of immunological response toward these synthetic polymers have triggered a search for better alternatives. In this work, natural red blood cell (RBC) membranes are camouflaged on the surface of Fe3O4 nanoparticles for reducing the RES uptake. In vitro macrophage uptake, in vivo biodistribution and pharmacokinetic studies demonstrate that the RBC membrane is a superior alternative to the current gold standard PEG for nanoparticle ‘stealth’. Furthermore, we systematically investigate the in vivo potential toxicity of RBC membrane-coated nanoparticles by blood biochemistry, whole blood panel examination and histology analysis based on animal models. The combination of synthetic nanoparticles and natural cell membranes embodies a novel and biomimetic nanomaterial design strategy and presents a compelling property of functional materials for a broad range of biomedical applications.

  16. Concentration dependence of the cell membrane permeability to cryoprotectant and water and implications for design of methods for post-thaw washing of human erythrocytes.

    PubMed

    Lahmann, John M; Benson, James D; Higgins, Adam Z

    2018-02-01

    For more than fifty years the human red blood cell (RBC) has been a widely studied model for transmembrane mass transport. Existing literature spans myriad experimental designs with varying results and physiologic interpretations. In this review, we examine the kinetics and mechanisms of membrane transport in the context of RBC cryopreservation. We include a discussion of the pathways for water and glycerol permeation through the cell membrane and the implications for mathematical modeling of the membrane transport process. In particular, we examine the concentration dependence of water and glycerol transport and provide equations for estimating permeability parameters as a function of concentration based on a synthesis of literature data. This concentration-dependent transport model may allow for design of improved methods for post-thaw removal of glycerol from cryopreserved blood. More broadly, the consideration of the concentration dependence of membrane permeability parameters may be important for other cell types as well, especially for design of methods for equilibration with the highly concentrated solutions used for vitrification. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. A Multiscale Red Blood Cell Model with Accurate Mechanics, Rheology, and Dynamics

    PubMed Central

    Fedosov, Dmitry A.; Caswell, Bruce; Karniadakis, George Em

    2010-01-01

    Abstract Red blood cells (RBCs) have highly deformable viscoelastic membranes exhibiting complex rheological response and rich hydrodynamic behavior governed by special elastic and bending properties and by the external/internal fluid and membrane viscosities. We present a multiscale RBC model that is able to predict RBC mechanics, rheology, and dynamics in agreement with experiments. Based on an analytic theory, the modeled membrane properties can be uniquely related to the experimentally established RBC macroscopic properties without any adjustment of parameters. The RBC linear and nonlinear elastic deformations match those obtained in optical-tweezers experiments. The rheological properties of the membrane are compared with those obtained in optical magnetic twisting cytometry, membrane thermal fluctuations, and creep followed by cell recovery. The dynamics of RBCs in shear and Poiseuille flows is tested against experiments and theoretical predictions, and the applicability of the latter is discussed. Our findings clearly indicate that a purely elastic model for the membrane cannot accurately represent the RBC's rheological properties and its dynamics, and therefore accurate modeling of a viscoelastic membrane is necessary. PMID:20483330

  18. Why do four NICUs using identical RBC transfusion guidelines have different gestational age-adjusted RBC transfusion rates?

    PubMed

    Henry, E; Christensen, R D; Sheffield, M J; Eggert, L D; Carroll, P D; Minton, S D; Lambert, D K; Ilstrup, S J

    2015-02-01

    To compare neonatal red blood cell (RBC) transfusion rates in four large Intermountain Healthcare NICUs, all of which adhere to the same RBC transfusion guidelines. This retrospective analysis was part of a transfusion-management quality-improvement project. De-identified data included RBC transfusions, clinical and laboratory findings, the anemia-prevention strategies in place in each NICU, and specific costs and outcomes. Of 2389 NICU RBC transfusions given during the 4-year period studied, 98.9 ± 2.1% (mean ± S.D.) were compliant with our transfusion guidelines, with no difference in compliance between any of the four NICUs. However, RBC transfusion rates varied widely between the four, with averages ranging from 4.6 transfusions/1000 NICU days to 21.7/1000 NICU days (P < 0.00001). Gestational age-adjusted transfusion rates were correspondingly discordant (P < 0.00001). The lower-transfusing NICUs had written anemia-preventing guidelines, such as umbilical cord milking at very low birth weight delivery, use of cord blood for admission laboratory studies, and darbepoetin dosing for selected neonates. Rates of Bell stage ⩾ 2 necrotizing enterocolitis and grade ⩾ 3 intraventricular hemorrhage were lowest in the two lower-transfusing NICUs (P < 0.0002 and P < 0.0016). Average pharmacy costs for darbepoetin were $84/dose, with an average pharmacy cost of $269 per transfusion averted. With a cost of $900/RBC transfusion, the anemia-preventing strategies resulted in an estimated cost savings to Intermountain Healthcare of about $6970 per 1000 NICU days, or about $282,300 annually. Using transfusion guidelines has been shown previously to reduce practice variability, lower transfusion rates and diminish transfusion costs. Based on our present findings, we maintain that even when transfusion guidelines are in place and adhered to rigorously, RBC transfusion rates are reduced further if anemia-preventing strategies are also in place.

  19. Reincorporated plasma membrane Ca2+-ATPase can mediate B-Type Ca2+ channels observed in native membrane of human red blood cells.

    PubMed

    Pinet, C; Antoine, S; Filoteo, A G; Penniston, J T; Coulombe, A

    2002-06-01

    Recently, we reported indirect evidence that plasma membrane Ca2+-ATPase (PMCA) can mediate B-type Ca2+ channels of cardiac myocytes. In the present study, in order to bring more direct evidence, purified PMCA from human red blood cells (RBC) was reconstituted into giant azolectin liposomes amenable to the patch-clamp technique. Purified RBC PMCA was used because it is available pure in larger quantity than cardiac PMCA. The presence of B-type Ca2+ channels was first investigated in native membranes of human RBC. They were detected and share the characteristics of cardiac myocytes. They spontaneously appeared in scarce short bursts of activity, they were activated by chlorpromazine (CPZ) with an EC50 of 149 mmole/l or 1 mmole/l vanadate, and then switched off by 10 mmole/l eosin or dose-dependently blocked by 1-5 mmole/l ATP. Independent of membrane potential, the channel gating exhibited complex patterns of many conductance levels, with three most often observed conductance levels of 22, 47 and 80 pS. The activation by vanadate suggests that these channels could play a role in the influx of extracellular Ca2+ involved in the vanadate-induced Gardos effect. In PMCA-reconstituted proteoliposomes, nearly half of the ATPase activity was retained and clear "channel-like" openings of Ba2+- or Ca2+-conducting channels were detected. Channel activity could be spontaneously present, lasting the patch lifetime or, when previously quiescent, activity could be induced by application of 50 mmole/l CPZ only in presence of 25 U/ml calmodulin (CaM), or by application of 1 mmole/l vanadate alone. Eosin (10 mmole/l) and ATP (5 mmole/l) significantly reduced spontaneous activity. Channel gating characteristics were similar to those of RBC, with main conductance levels of 21, 40 and 72 pS. The lack of direct activation by CPZ alone might be attributed to a purification-induced modification or absence of unidentified regulatory component(s) of PMCA. Despite a few differences in

  20. A robust mass spectrometry method for rapid profiling of erythrocyte ghost membrane proteomes.

    PubMed

    Fye, Haddy K S; Mrosso, Paul; Bruce, Lesley; Thézénas, Marie-Laëtitia; Davis, Simon; Fischer, Roman; Rwegasira, Gration L; Makani, Julie; Kessler, Benedikt M

    2018-01-01

    Red blood cell (RBC) physiology is directly linked to many human disorders associated with low tissue oxygen levels or anemia including chronic obstructive pulmonary disease, congenital heart disease, sleep apnea and sickle cell anemia. Parasites such as Plasmodium spp. and phylum Apicomplexa directly target RBCs, and surface molecules within the RBC membrane are critical for pathogen interactions. Proteomics of RBC membrane 'ghost' fractions has therefore been of considerable interest, but protocols described to date are either suboptimal or too extensive to be applicable to a larger set of clinical cohorts. Here, we describe an optimised erythrocyte isolation protocol from blood, tested for various storage conditions and explored using different fractionation conditions for isolating ghost RBC membranes. Liquid chromatography mass spectrometry (LC-MS) analysis on a Q-Exactive Orbitrap instrument was used to profile proteins isolated from the comparative conditions. Data analysis was run on the MASCOT and MaxQuant platforms to assess their scope and diversity. The results obtained demonstrate a robust method for membrane enrichment enabling consistent MS based characterisation of > 900 RBC membrane proteins in single LC-MS/MS analyses. Non-detergent based membrane solubilisation methods using the tissue and supernatant fractions of isolated ghost membranes are shown to offer effective haemoglobin removal as well as diverse recovery including erythrocyte membrane proteins of high and low abundance. The methods described in this manuscript propose a medium to high throughput framework for membrane proteome profiling by LC-MS of potential applicability to larger clinical cohorts in a variety of disease contexts.

  1. Agglutination of like-charged red blood cells induced by binding of beta2-glycoprotein I to outer cell surface.

    PubMed

    Lokar, Marusa; Urbanija, Jasna; Frank, Mojca; Hägerstrand, Henry; Rozman, Blaz; Bobrowska-Hägerstrand, Malgorzata; Iglic, Ales; Kralj-Iglic, Veronika

    2008-08-01

    Plasma protein-mediated attractive interaction between membranes of red blood cells (RBCs) and phospholipid vesicles was studied. It is shown that beta(2)-glycoprotein I (beta(2)-GPI) may induce RBC discocyte-echinocyte-spherocyte shape transformation and subsequent agglutination of RBCs. Based on the observed beta(2)-GPI-induced RBC cell shape transformation it is proposed that the hydrophobic portion of beta(2)-GPI molecule protrudes into the outer lipid layer of the RBC membrane and increases the area of this layer. It is also suggested that the observed agglutination of RBCs is at least partially driven by an attractive force which is of electrostatic origin and depends on the specific molecular shape and internal charge distribution of membrane-bound beta(2)-GPI molecules. The suggested beta(2)-GPI-induced attractive electrostatic interaction between like-charged RBC membrane surfaces is qualitatively explained by using a simple mathematical model within the functional density theory of the electric double layer, where the electrostatic attraction between the positively charged part of the first domains of bound beta(2)-GPI molecules and negatively charged glycocalyx of the adjacent RBC membrane is taken into account.

  2. Red blood cell (RBC) suspensions in confined microflows: Pressure-flow relationship.

    PubMed

    Stauber, Hagit; Waisman, Dan; Korin, Netanel; Sznitman, Josué

    2017-10-01

    Microfluidic-based assays have become increasingly popular to explore microcirculation in vitro. In these experiments, blood is resuspended to a desired haematocrit level in a buffer solution, where frequent choices for preparing RBC suspensions comprise notably Dextran and physiological buffer. Yet, the rational for selecting one buffer versus another is often ill-defined and lacks detailed quantification, including ensuing changes in RBC flow characteristics. Here, we revisit RBC suspensions in microflows and attempt to quantify systematically some of the differences emanating between buffers. We measure bulk flow rate (Q) of RBC suspensions, using PBS- and Dextran-40, as a function of the applied pressure drop (ΔP) for two hematocrits (∼0% and 23%). Two distinct microfluidic designs of varying dimensions are employed: a straight channel larger than and a network array similar to the size of individual RBCs. Using the resulting pressure-flow curves, we extract the equivalent hydrodynamic resistances and estimate the relative viscosities. These efforts are a first step in rigorously quantifying the influence of the 'background' buffer on RBC flows within microfluidic devices and thereby underline the importance of purposefully selecting buffer suspensions for microfluidic in vitro assays. Copyright © 2017. Published by Elsevier Ltd.

  3. Proper cytoskeletal architecture beneath the plasma membrane of red blood cells requires Ttll4

    PubMed Central

    Ijaz, Faryal; Hatanaka, Yasue; Hatanaka, Takahiro; Tsutsumi, Koji; Iwaki, Takayuki; Umemura, Kazuo; Ikegami, Koji; Setou, Mitsutoshi

    2017-01-01

    Mammalian red blood cells (RBCs) circulate through blood vessels, including capillaries, for tens of days under high mechanical stress. RBCs tolerate this mechanical stress while maintaining their shape because of their elastic membrane skeleton. This membrane skeleton consists of spectrin-actin lattices arranged as quasi-hexagonal units beneath the plasma membrane. In this study, we found that the organization of the RBC cytoskeleton requires tubulin tyrosine ligase–like 4 (Ttll4). RBCs from Ttll4-knockout mice showed larger average diameters in smear test. Based on the rate of hemolysis, Ttll4-knockout RBCs showed greater vulnerability to phenylhydrazine-induced oxidative stress than did wild-type RBCs. Ultrastructural analyses revealed the macromolecular aggregation of cytoskeletal components in RBCs of Ttll4-knockout mice. Immunoprecipitation using the anti-glutamylation antibody GT335 revealed nucleosome assembly protein 1 (NAP1) to be the sole target of TTLL4 in the RBCs, and NAP1 glutamylation was completely lost in Ttll4-knockout RBCs. In wild-type RBCs, the amount of glutamylated NAP1 in the membrane was nearly double that in the cytosol. Furthermore, the absence of TTLL4-dependent glutamylation of NAP1 weakened the binding of NAP1 to the RBC membrane. Taken together, these data demonstrate that Ttll4 is required for proper cytoskeletal organization in RBCs. PMID:27974641

  4. RBC nuclear scan

    MedlinePlus

    ... page: //medlineplus.gov/ency/article/003835.htm RBC nuclear scan To use the sharing features on this page, please enable JavaScript. An RBC nuclear scan uses small amounts of radioactive material to ...

  5. Detection and characterization of red blood cell (RBC) aggregation with photoacoustics

    NASA Astrophysics Data System (ADS)

    Hysi, Eno; Saha, Ratan K.; Rui, Min; Kolios, Michael C.

    2012-02-01

    Red blood cells (RBCs) aggregate in the presence of increased plasma fibrinogen and low shear forces during blood flow. RBC aggregation has been observed in deep vein thrombosis, sepsis and diabetes. We propose using photoacoustics (PA) as a non-invasive imaging modality to detect RBC aggregation. The theoretical and experimental feasibility of PA for detecting and characterizing aggregation was assessed. A simulation study was performed to generate PA signals from non-aggregated and aggregated RBCs using a frequency domain approach and to study the PA signals' dependence on hematocrit and aggregate size. The effect of the finite bandwidth nature of transducers on the PA power spectra was also investigated. Experimental confirmation of theoretical results was conducted using porcine RBC samples exposed to 1064 nm optical wavelength using the Imagio Small Animal PA imaging system (Seno Medical Instruments, Inc., San Antonio, TX). Aggregation was induced with Dextran-70 (Sigma-Aldrich, St. Louis, MO) and the effect of hematocrit and aggregation level was investigated. The theoretical and experimental PA signal amplitude increased linearly with increasing hematocrit. The theoretical dominant frequency content of PA signals shifted towards lower frequencies (<30 MHz) and 9 dB enhancements in spectral power were observed as the size of aggregates increased compared to non-aggregating RBCs. Calibration of the PA spectra with the transducer response obtained from a 200 nm gold film was performed to remove system dependencies. Analysis of the spectral parameters from the calibrated spectra suggested that PA can assess the degree of aggregation at multiple hematocrit and aggregation levels.

  6. Use of a flow-cell system to investigate virucidal dimethylmethylene blue phototreatment in two RBC additive solutions.

    PubMed

    Wagner, Stephen; Skripchenko, Andrey; Thompson-Montgomery, Dedeene

    2002-09-01

    Limited photoinactivation kinetics, use of low-volume 30 percent Hct RBCs, and hemolysis have restricted the practicality of the use of dimethylmethylene blue (DMMB) and light for RBC decontamination. A flow-cell system was developed to rapidly treat larger volumes of oxygenated 45 percent Hct RBCs with high-intensity red light. CPD-whole blood was WBC reduced, RBCs were diluted in additive solutions (either Adsol or Erythrosol), and suspensions were subsequently oxygenated by gas overlay. Intracellular or extracellular VSV and DMMB were sequentially added. VSV-infected RBC suspensions (45% Hct) were passed through 1-mm-thick flow cells and illuminated. Samples were titered for VSV, stored for up to 42 days, and assayed for Hb, supernatant potassium, ATP, and MCV. The use of oxygenated RBCs resulted in rapid and reproducible photoinactivaton of > or = 6.6 log extracellular and approximately 4.0 log intracellular VSV independent of additive solution. Phototreated Adsol RBCs exhibited more than 10 times greater hemolysis and 30 percent greater MCV during storage than identically treated Erythrosol RBCs. Phototreatment caused RBC potassium leakage from RBCs in both additive solutions. ATP levels were better preserved in Erythrosol than Adsol RBCs. A rapid, reproducible, and robust method for photoinactivating model virus in RBC suspensions was developed. Despite improved hemolysis and ATP levels in Erythrosol-phototreated RBCs, storage properties were not maintained for 42 days.

  7. A multiscale red blood cell model with accurate mechanics, rheology, and dynamics.

    PubMed

    Fedosov, Dmitry A; Caswell, Bruce; Karniadakis, George Em

    2010-05-19

    Red blood cells (RBCs) have highly deformable viscoelastic membranes exhibiting complex rheological response and rich hydrodynamic behavior governed by special elastic and bending properties and by the external/internal fluid and membrane viscosities. We present a multiscale RBC model that is able to predict RBC mechanics, rheology, and dynamics in agreement with experiments. Based on an analytic theory, the modeled membrane properties can be uniquely related to the experimentally established RBC macroscopic properties without any adjustment of parameters. The RBC linear and nonlinear elastic deformations match those obtained in optical-tweezers experiments. The rheological properties of the membrane are compared with those obtained in optical magnetic twisting cytometry, membrane thermal fluctuations, and creep followed by cell recovery. The dynamics of RBCs in shear and Poiseuille flows is tested against experiments and theoretical predictions, and the applicability of the latter is discussed. Our findings clearly indicate that a purely elastic model for the membrane cannot accurately represent the RBC's rheological properties and its dynamics, and therefore accurate modeling of a viscoelastic membrane is necessary. Copyright 2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  8. Numerical analysis of a red blood cell flowing through a thin micropore.

    PubMed

    Omori, Toshihiro; Hosaka, Haruki; Imai, Yohsuke; Yamaguchi, Takami; Ishikawa, Takuji

    2014-01-01

    Red blood cell (RBC) deformability plays a key role in microcirculation, especially in vessels that have diameters even smaller than the nominal cell size. In this study, we numerically investigate the dynamics of an RBC in a thin micropore. The RBC is modeled as a capsule with a thin hyperelastic membrane. In a numerical simulation, we employ a boundary element method for fluid mechanics and a finite element method for membrane mechanics. The resulting RBC deformation towards the flow direction is suppressed considerably by increased cytoplasm viscosity, whereas the gap between the cell membrane and solid wall becomes smaller with higher cytoplasm viscosity. We also measure the transit time of the RBC and find that nondimensional transit time increases nonlinearly with respect to the viscosity ratio, whereas it is invariant to the capillary number. In conclusion, cytoplasmic viscosity plays a key role in the dynamics of an RBC in a thin pore. The results of this study will be useful for designing a microfluidic device to measure cytoplasmic viscosity.

  9. Plasmodium falciparum-Infected Erythrocytes Induce Granzyme B by NK Cells through Expression of Host-Hsp70

    PubMed Central

    Böttger, Evelyn; Multhoff, Gabriele; Kun, Jürgen F. J.; Esen, Meral

    2012-01-01

    In the early immune response to Plasmodium falciparum-infected erythrocytes (iRBC), Natural Killer (NK) cells are activated, which suggests an important role in innate anti-parasitic immunity. However, it is not well understood whether NK cells directly recognize iRBC or whether stimulation of NK cells depends mainly on activating signals from accessory cells through cell-to-cell contact or soluble factors. In the present study, we investigated the influence of membrane-bound host Heat shock protein (Hsp) 70 in triggering cytotoxicity of NK cells from malaria-naïve donors or the cell line NK92 against iRBC. Hsp70 and HLA-E membrane expression on iRBC and potential activatory NK cell receptors (NKG2C, CD94) were assessed by flow cytometry and immunoblot. Upon contact with iRBC, Granzyme B (GzmB) production and release was initiated by unstimulated and Hsp70-peptide (TKD) pre-stimulated NK cells, as determined by Western blot, RT-PCR and ELISPOT analysis. Eryptosis of iRBC was determined by Annexin V-staining. Our results suggest that presence of Hsp70 and absence of HLA-E on the membrane of iRBC prompt the infected host cells to become targets for NK cell-mediated cytotoxicity, as evidenced by impaired parasite development. Contact of iRBC with NK cells induced release of GzmB. We propose that following GzmB uptake, iRBC undergo eryptosis via a perforin-independent, GzmB-mediated mechanism. Since NK activity toward iRBC could be specifically enhanced by TKD peptide and abrogated to baseline levels by blocking Hsp70 exposure, we propose TKD as an innovative immunostimulatory agent to be tested as an adjunct to anti-parasitic treatments in vivo. PMID:22438997

  10. Plasmodium falciparum-infected erythrocytes induce granzyme B by NK cells through expression of host-Hsp70.

    PubMed

    Böttger, Evelyn; Multhoff, Gabriele; Kun, Jürgen F J; Esen, Meral

    2012-01-01

    In the early immune response to Plasmodium falciparum-infected erythrocytes (iRBC), Natural Killer (NK) cells are activated, which suggests an important role in innate anti-parasitic immunity. However, it is not well understood whether NK cells directly recognize iRBC or whether stimulation of NK cells depends mainly on activating signals from accessory cells through cell-to-cell contact or soluble factors. In the present study, we investigated the influence of membrane-bound host Heat shock protein (Hsp) 70 in triggering cytotoxicity of NK cells from malaria-naïve donors or the cell line NK92 against iRBC. Hsp70 and HLA-E membrane expression on iRBC and potential activatory NK cell receptors (NKG2C, CD94) were assessed by flow cytometry and immunoblot. Upon contact with iRBC, Granzyme B (GzmB) production and release was initiated by unstimulated and Hsp70-peptide (TKD) pre-stimulated NK cells, as determined by Western blot, RT-PCR and ELISPOT analysis. Eryptosis of iRBC was determined by Annexin V-staining. Our results suggest that presence of Hsp70 and absence of HLA-E on the membrane of iRBC prompt the infected host cells to become targets for NK cell-mediated cytotoxicity, as evidenced by impaired parasite development. Contact of iRBC with NK cells induced release of GzmB. We propose that following GzmB uptake, iRBC undergo eryptosis via a perforin-independent, GzmB-mediated mechanism. Since NK activity toward iRBC could be specifically enhanced by TKD peptide and abrogated to baseline levels by blocking Hsp70 exposure, we propose TKD as an innovative immunostimulatory agent to be tested as an adjunct to anti-parasitic treatments in vivo.

  11. Prolongation of RBC survival in the hypophysectomized rat.

    NASA Technical Reports Server (NTRS)

    Landaw, S. A.; Bristol, S. K.

    1971-01-01

    Red blood cell (RBC) survival was prolonged in hypophysectomized rats. While the rate of random hemolysis was decreased in some hypophysectomized hosts, in all directly injected and cross-transfused hypophysectomized rat hosts, there was a significant prolongation of the phase of senescent death. In contrast, RBCs from hypophysectomized donors survived normally in normal hosts. These experiments are further evidence of a relationship between RBC aging and metabolic rate, and suggest an intimate involvement with the calorigenic hormones.

  12. A microengineered model of RBC transfusion-induced pulmonary vascular injury.

    PubMed

    Seo, Jeongyun; Conegliano, David; Farrell, Megan; Cho, Minseon; Ding, Xueting; Seykora, Thomas; Qing, Danielle; Mangalmurti, Nilam S; Huh, Dongeun

    2017-06-13

    Red blood cell (RBC) transfusion poses significant risks to critically ill patients by increasing their susceptibility to acute respiratory distress syndrome. While the underlying mechanisms of this life-threatening syndrome remain elusive, studies suggest that RBC-induced microvascular injury in the distal lung plays a central role in the development of lung injury following blood transfusion. Here we present a novel microengineering strategy to model and investigate this key disease process. Specifically, we created a microdevice for culturing primary human lung endothelial cells under physiological flow conditions to recapitulate the morphology and hemodynamic environment of the pulmonary microvascular endothelium in vivo. Perfusion of the microengineered vessel with human RBCs resulted in abnormal cytoskeletal rearrangement and release of intracellular molecules associated with regulated necrotic cell death, replicating the characteristics of acute endothelial injury in transfused lungs in vivo. Our data also revealed the significant effect of hemodynamic shear stress on RBC-induced microvascular injury. Furthermore, we integrated the microfluidic endothelium with a computer-controlled mechanical stretching system to show that breathing-induced physiological deformation of the pulmonary microvasculature may exacerbate vascular injury during RBC transfusion. Our biomimetic microsystem provides an enabling platform to mechanistically study transfusion-associated pulmonary vascular complications in susceptible patient populations.

  13. RBC deformability and amino acid concentrations after hypo-osmotic challenge may reflect chronic cell hydration status in healthy young men

    PubMed Central

    Stookey, Jodi D; Klein, Alexis; Hamer, Janice; Chi, Christine; Higa, Annie; Ng, Vivian; Arieff, Allen; Kuypers, Frans A; Larkin, Sandra; Perrier, Erica; Lang, Florian

    2013-01-01

    Biomarkers of chronic cell hydration status are needed to determine whether chronic hyperosmotic stress increases chronic disease risk in population-representative samples. In vitro, cells adapt to chronic hyperosmotic stress by upregulating protein breakdown to counter the osmotic gradient with higher intracellular amino acid concentrations. If cells are subsequently exposed to hypo-osmotic conditions, the adaptation results in excess cell swelling and/or efflux of free amino acids. This study explored whether increased red blood cell (RBC) swelling and/or plasma or urine amino acid concentrations after hypo-osmotic challenge might be informative about relative chronic hyperosmotic stress in free-living men. Five healthy men (20–25 years) with baseline total water intake below 2 L/day participated in an 8-week clinical study: four 2-week periods in a U-shaped A-B-C-A design. Intake of drinking water was increased by +0.8 ± 0.3 L/day in period 2, and +1.5 ± 0.3 L/day in period 3, and returned to baseline intake (0.4 ± 0.2 L/day) in period 4. Each week, fasting blood and urine were collected after a 750 mL bolus of drinking water, following overnight water restriction. The periods of higher water intake were associated with significant decreases in RBC deformability (index of cell swelling), plasma histidine, urine arginine, and urine glutamic acid. After 4 weeks of higher water intake, four out of five participants had ½ maximal RBC deformability below 400 mmol/kg; plasma histidine below 100 μmol/L; and/or undetectable urine arginine and urine glutamic acid concentrations. Work is warranted to pursue RBC deformability and amino acid concentrations after hypo-osmotic challenge as possible biomarkers of chronic cell hydration. PMID:24303184

  14. Hemoglobin redox reactions and red blood cell aging.

    PubMed

    Rifkind, Joseph M; Nagababu, Enika

    2013-06-10

    The physiological mechanism(s) for recognition and removal of red blood cells (RBCs) from circulation after 120 days of its lifespan is not fully understood. Many of the processes thought to be associated with the removal of RBCs involve oxidative stress. We have focused on hemoglobin (Hb) redox reactions, which is the major source of RBC oxidative stress. The importance of Hb redox reactions have been shown to originate in large parts from the continuous slow autoxidation of Hb producing superoxide and its dramatic increase under hypoxic conditions. In addition, oxidative stress has been shown to be associated with redox reactions that originate from Hb reactions with nitrite and nitric oxide (NO) and the resultant formation of highly toxic peroxynitrite when NO reacts with superoxide released during Hb autoxidation. The interaction of Hb, particularly under hypoxic conditions with band 3 of the RBC membrane is critical for the generating the RBC membrane changes that trigger the removal of cells from circulation. These changes include exposure of antigenic sites, increased calcium leakage into the RBC, and the resultant leakage of potassium out of the RBC causing cell shrinkage and impaired deformability. The need to understand the oxidative damage to specific membrane proteins that result from redox reactions occurring when Hb is bound to the membrane. Proteomic studies that can pinpoint the specific proteins damaged under different conditions will help elucidate the cellular aging processes that result in cells being removed from circulation.

  15. Unexpected similarity in RBC DHA and AA levels between bottlenose dolphins and humans.

    PubMed

    Harris, William S; Schmitt, Todd L

    2014-01-01

    The Omega-3 Index [red blood cell (RBC) content of eicosapentaenoic acid (EPA)+docosahexaenoic acid (DHA)] is inversely related to risk of cardiovascular disease in humans. In the U.S., the average Omega-3 Index is about 4-6% of RBC fatty acids, whereas in Japan it is 9-10%. The range of physiologically-possible levels for the Omega-3 Index in other mammals is unknown. To compare the RBC fatty acid composition of a common piscivorous mammal, the bottlenose dolphin (Tursiops truncatus), with that of (U.S.) humans, and to examine the extent to which dietary fatty acid patterns were reflected in RBCs. RBCs were isolated from routine blood samples collected from 35 healthy dolphins at two display facilities and were analyzed by gas chromatography. For humans, historic, deidentified RBC fatty acid data from our laboratory were used (n=11,329; mean age 58). The mean Omega-3 Index of the dolphins was 19.9% compared with 6.0% for humans. EPA levels were 15.3% vs 1.2%, respectively, but DHA levels were virtually identical (4.6% vs 4.8%). Linoleic acid (LNA) levels were much lower in dolphins vs humans (0.5% vs 12.5%) whereas arachidonic acid (ARA) levels were similar (12.3% vs 14.5%). In a subgroup of humans with an Omega-3 Index in the >99.2 percentile, the mean index was similar to that of the dolphins. Based on an analysis of their food, the dolphins consumed about 60g of EPA+DHA per day as compared to about 0.1g in humans. Dolphins have an Omega-3 Index that is (only) 3-4× higher than that of U.S. adults despite their intake of EPA+DHA being about 165× higher (as a percent of kcal). RBC, EPA and LNA levels are relatively more reflective of dietary intakes than are DHA and ARA levels. The mechanisms by which certain fatty acid levels appear to be fixed and others may vary in RBC membranes are unknown. Copyright © 2013 Elsevier Ltd. All rights reserved.

  16. Microconfined flow behavior of red blood cells.

    PubMed

    Tomaiuolo, Giovanna; Lanotte, Luca; D'Apolito, Rosa; Cassinese, Antonio; Guido, Stefano

    2016-01-01

    Red blood cells (RBCs) perform essential functions in human body, such as gas exchange between blood and tissues, thanks to their ability to deform and flow in the microvascular network. The high RBC deformability is mainly due to the viscoelastic properties of the cell membrane. Since an impaired RBC deformability could be found in some diseases, such as malaria, sickle cell anemia, diabetes and hereditary disorders, there is the need to provide further insight into measurement of RBC deformability in a physiologically relevant flow field. Here, RBCs deformability has been studied in terms of the minimum apparent plasma-layer thickness by using high-speed video microscopy of RBCs flowing in cylindrical glass capillaries. An in vitro systematic microfluidic investigation of RBCs in micro-confined conditions has been performed, resulting in the determination of the RBCs time recovery constant, RBC volume and surface area and RBC membrane shear elastic modulus and surface viscosity. It has been noticed that the deformability of RBCs induces cells aggregation during flow in microcapillaries, allowing the formation of clusters of cells. Overall, our results provide a novel technique to estimate RBC deformability and also RBCs collective behavior, which can be used for the analysis of pathological RBCs, for which reliable quantitative methods are still lacking. Copyright © 2015 IPEM. Published by Elsevier Ltd. All rights reserved.

  17. Compartmental hollow fiber capillary membrane-based bioreactor technology for in vitro studies on red blood cell lineage direction of hematopoietic stem cells.

    PubMed

    Housler, Greggory J; Miki, Toshio; Schmelzer, Eva; Pekor, Christopher; Zhang, Xiaokui; Kang, Lin; Voskinarian-Berse, Vanessa; Abbot, Stewart; Zeilinger, Katrin; Gerlach, Jörg C

    2012-02-01

    Continuous production of red blood cells (RBCs) in an automated closed culture system using hematopoietic stem cell (HSC) progenitor cell populations is of interest for clinical application because of the high demand for blood transfusions. Previously, we introduced a four-compartment bioreactor that consisted of two bundles of hollow fiber microfiltration membranes for transport of culture medium (forming two medium compartments), interwoven with one bundle of hollow fiber membranes for transport of oxygen (O(2)), carbon dioxide (CO(2)), and other gases (forming one gas compartment). Small-scale prototypes were developed of the three-dimensional (3D) perfusion cell culture systems, which enable convection-based mass transfer and integral oxygenation in the cell compartment. CD34(+) HSC were isolated from human cord blood units using a magnetic separation procedure. Cells were inoculated into 2- or 8-mL scaled-down versions of the previously designed 800-mL cell compartment devices and perfused with erythrocyte proliferation and differentiation medium. First, using the small-scale 2-mL analytical scale bioreactor, with an initial seeding density of 800,000 cells/mL, we demonstrated approximately 100-fold cell expansion and differentiation after 7 days of culture. An 8-mL laboratory-scale bioreactor was then used to show pseudocontinuous production by intermediately harvesting cells. Subsequently, we were able to use a model to demonstrate semicontinuous production with up to 14,288-fold expansion using seeding densities of 800,000 cells/mL. The down-scaled culture technology allows for expansion of CD34(+) cells and stimulating these progenitors towards RBC lineage, expressing approximately 40% CD235(+) and enucleation. The 3D perfusion technology provides an innovative tool for studies on RBC production, which is scalable.

  18. Inhibition of membrane Na(+)-K+ Atpase of the brain, liver and RBC in rats administered di(2-ethyl hexyl) phthalate (DEHP) a plasticizer used in polyvinyl chloride (PVC) blood storage bags.

    PubMed

    Dhanya, C R; Indu, A R; Deepadevi, K V; Kurup, P A

    2003-08-01

    Significant amounts of di(2-ethylhexyl) phthalate (DEHP) leach out into blood stored in DEHP plasticized polyvinyl chloride (PVC) bags resulting in the exposure of recipients of blood transfusion to this compound. The aim of this study was to find out whether DEHP at these low levels has any effect on the activity of membrane Na(+)-K+ ATPase, since a decrease in this enzyme activity has been reported to take place in a number of disorders like neurodegenerative and psychiatric disorders, coronary artery disease and stroke, syndrome-X, tumours etc. DEHP was administered (ip) at a low dose of 750 microg/100 g body weight to rats and the activity of membrane Na(+)-K+ ATPase in liver, brain and RBC was estimated. Histopathology of brain, activity of HMG CoA reductase (a major rate limiting enzyme in the isoprenoid pathway of which digoxin, the physiological inhibitor of Na(+)-K+ ATPase is a product), intracellular concentration of Ca2+ and Mg2+ in RBC (which is altered as a result of inhibition of Na(+)-K+ ATPase) were also studied. (In the light of the observation of increase of intracellular Ca2+ load and intracellular depletion of Mg2+ when Na(+)-K+ ATPase is inhibited). Histopathology of brain revealed areas of degeneration in the rats administered DEHP. There was significant inhibition of membrane Na(+)-K+ ATPase in brain, liver and RBC. Intracellular Ca2+ increased in the RBC while intracellular Mg2+ decreased. However activity of hepatic HMG CoA reductase decreased. Activity of Na(+)-K+ ATPase and HMG CoA reductase, however returned to normal levels within 7 days of stopping administration of DEHP. The inhibition of membrane Na(+)-K+ ATPase activity by DEHP may indicate the possibility of predisposing recipients of transfusion of blood or hemodialysis to the various disorders mentioned above. However since this effect is reversed when DEHP administration is stopped, it may not be a serious problem in the case of a few transfusion; but in patients receiving

  19. Red blood cell oxidative stress impairs oxygen delivery and induces red blood cell aging.

    PubMed

    Mohanty, Joy G; Nagababu, Enika; Rifkind, Joseph M

    2014-01-01

    Red Blood Cells (RBCs) need to deform and squeeze through narrow capillaries. Decreased deformability of RBCs is, therefore, one of the factors that can contribute to the elimination of aged or damaged RBCs from the circulation. This process can also cause impaired oxygen delivery, which contributes to the pathology of a number of diseases. Studies from our laboratory have shown that oxidative stress plays a significant role in damaging the RBC membrane and impairing its deformability. RBCs are continuously exposed to both endogenous and exogenous sources of reactive oxygen species (ROS) like superoxide and hydrogen peroxide (H2O2). The bulk of the ROS are neutralized by the RBC antioxidant system consisting of both non-enzymatic and enzymatic antioxidants including catalase, glutathione peroxidase and peroxiredoxin-2. However, the autoxidation of hemoglobin (Hb) bound to the membrane is relatively inaccessible to the predominantly cytosolic RBC antioxidant system. This inaccessibility becomes more pronounced under hypoxic conditions when Hb is partially oxygenated, resulting in an increased rate of autoxidation and increased affinity for the RBC membrane. We have shown that a fraction of peroxyredoxin-2 present on the RBC membrane may play a major role in neutralizing these ROS. H2O2 that is not neutralized by the RBC antioxidant system can react with the heme producing fluorescent heme degradation products (HDPs). We have used the level of these HDP as a measure of RBC oxidative Stress. Increased levels of HDP are detected during cellular aging and various diseases. The negative correlation (p < 0.0001) between the level of HDP and RBC deformability establishes a contribution of RBC oxidative stress to impaired deformability and cellular stiffness. While decreased deformability contributes to the removal of RBCs from the circulation, oxidative stress also contributes to the uptake of RBCs by macrophages, which plays a major role in the removal of RBCs from

  20. Erythrocyte membrane model with explicit description of the lipid bilayer and the spectrin network.

    PubMed

    Li, He; Lykotrafitis, George

    2014-08-05

    The membrane of the red blood cell (RBC) consists of spectrin tetramers connected at actin junctional complexes, forming a two-dimensional (2D) sixfold triangular network anchored to the lipid bilayer. Better understanding of the erythrocyte mechanics in hereditary blood disorders such as spherocytosis, elliptocytosis, and especially, sickle cell disease requires the development of a detailed membrane model. In this study, we introduce a mesoscale implicit-solvent coarse-grained molecular dynamics (CGMD) model of the erythrocyte membrane that explicitly describes the phospholipid bilayer and the cytoskeleton, by extending a previously developed two-component RBC membrane model. We show that the proposed model represents RBC membrane with the appropriate bending stiffness and shear modulus. The timescale and self-consistency of the model are established by comparing our results with experimentally measured viscosity and thermal fluctuations of the RBC membrane. Furthermore, we measure the pressure exerted by the cytoskeleton on the lipid bilayer. We find that defects at the anchoring points of the cytoskeleton to the lipid bilayer (as in spherocytes) cause a reduction in the pressure compared with an intact membrane, whereas defects in the dimer-dimer association of a spectrin filament (as in elliptocytes) cause an even larger decrease in the pressure. We conjecture that this finding may explain why the experimentally measured diffusion coefficients of band-3 proteins are higher in elliptocytes than in spherocytes, and higher than in normal RBCs. Finally, we study the effects that possible attractive forces between the spectrin filaments and the lipid bilayer have on the pressure applied on the lipid bilayer by the filaments. We discover that the attractive forces cause an increase in the pressure as they diminish the effect of membrane protein defects. As this finding contradicts with experimental results, we conclude that the attractive forces are moderate and do

  1. Erythrocyte Membrane Model with Explicit Description of the Lipid Bilayer and the Spectrin Network

    PubMed Central

    Li, He; Lykotrafitis, George

    2014-01-01

    The membrane of the red blood cell (RBC) consists of spectrin tetramers connected at actin junctional complexes, forming a two-dimensional (2D) sixfold triangular network anchored to the lipid bilayer. Better understanding of the erythrocyte mechanics in hereditary blood disorders such as spherocytosis, elliptocytosis, and especially, sickle cell disease requires the development of a detailed membrane model. In this study, we introduce a mesoscale implicit-solvent coarse-grained molecular dynamics (CGMD) model of the erythrocyte membrane that explicitly describes the phospholipid bilayer and the cytoskeleton, by extending a previously developed two-component RBC membrane model. We show that the proposed model represents RBC membrane with the appropriate bending stiffness and shear modulus. The timescale and self-consistency of the model are established by comparing our results with experimentally measured viscosity and thermal fluctuations of the RBC membrane. Furthermore, we measure the pressure exerted by the cytoskeleton on the lipid bilayer. We find that defects at the anchoring points of the cytoskeleton to the lipid bilayer (as in spherocytes) cause a reduction in the pressure compared with an intact membrane, whereas defects in the dimer-dimer association of a spectrin filament (as in elliptocytes) cause an even larger decrease in the pressure. We conjecture that this finding may explain why the experimentally measured diffusion coefficients of band-3 proteins are higher in elliptocytes than in spherocytes, and higher than in normal RBCs. Finally, we study the effects that possible attractive forces between the spectrin filaments and the lipid bilayer have on the pressure applied on the lipid bilayer by the filaments. We discover that the attractive forces cause an increase in the pressure as they diminish the effect of membrane protein defects. As this finding contradicts with experimental results, we conclude that the attractive forces are moderate and do

  2. Drag reducing polymers improve tissue perfusion via modification of the RBC traffic in microvessels.

    PubMed

    Marhefka, J N; Zhao, R; Wu, Z J; Velankar, S S; Antaki, J F; Kameneva, M V

    2009-01-01

    This paper reports a novel, physiologically significant, microfluidic phenomenon generated by nanomolar concentrations of drag-reducing polymers (DRP) dissolved in flowing blood, which may explain previously demonstrated beneficial effects of DRP on tissue perfusion. In microfluidic systems used in this study, DRP additives were found to significantly modify traffic of red blood cells (RBC) into microchannel branches as well as reduce the near-wall cell-free layer, which normally is found in microvessels with a diameter smaller than 0.3 mm. The reduction in plasma layer size led to attenuation of the so-called "plasma skimming" effect at microchannel bifurcations, increasing the number of RBC entering branches. In vivo, these changes in RBC traffic may facilitate gas transport by increasing the near vessel wall concentration of RBC and capillary hematocrit. In addition, an increase in near-wall viscosity due to the redirection of RBC in this region may potentially decrease vascular resistance as a result of increased wall shear stress, which promotes endothelium mediated vasodilation. These microcirculatory phenomena can explain the previously reported beneficial effects of DRP on hemodynamics in vivo observed in many animal studies. We also report here our finding that DRP additives reduce flow separations at microchannel expansions, deflecting RBC closer to the wall and eliminating the plasma recirculation zone. Although the exact mechanism of the DRP effects on RBC traffic in microchannels is yet to be elucidated, these findings may further DRP progress toward clinical use.

  3. Drag reducing polymers improve tissue perfusion via modification of the RBC traffic in microvessels

    PubMed Central

    Marhefka, J.N.; Zhao, R.; Wu, Z.; Velankar, S.S.; Antaki, J.F.; Kameneva, M.V.

    2011-01-01

    This paper reports a novel, physiologically significant, microfluidic phenomenon generated by nanomolar concentrations of drag-reducing polymers (DRP) dissolved in flowing blood, which may explain previously demonstrated beneficial effects of DRP on tissue perfusion. In microfluidic systems used in this study, DRP additives were found to significantly modify traffic of red blood cells (RBC) into microchannel branches as well as reduce the near-wall cell-free layer, which normally is found in microvessels with a diameter smaller than 0.3 mm. The reduction in plasma layer size led to attenuation of the so-called “plasma skimming” effect at microchannel bifurcations, increasing the number of RBC entering branches. In vivo, these changes in RBC traffic may facilitate gas transport by increasing the near vessel wall concentration of RBC and capillary hematocrit. In addition, an increase in near-wall viscosity due to the redirection of RBC in this region may potentially decrease vascular resistance as a result of increased wall shear stress, which promotes endothelium mediated vasodilation. These microcirculatory phenomena may explain the previously reported beneficial effects of DRP on hemodynamics in vivo observed in many animal studies. We also report here our finding that DRP additives reduce flow separations at microchannel expansions, deflecting RBC closer to the wall and eliminating the plasma recirculation zone. Although the exact mechanism of the DRP effects on RBC traffic in microchannels is yet to be elucidated, these findings may further DRP progress toward clinical use. PMID:19721190

  4. DPD simulation on the dynamics of a healthy and infected red blood cell in flow through a constricted channel

    NASA Astrophysics Data System (ADS)

    Hoque, Sazid Zamal; Anand, D. Vijay; Patnaik, B. S. V.

    2017-11-01

    The state of the red blood cell (either healthy or infected RBC) will influence its deformation dynamics. Since the pathological condition related to RBC, primarily originates from a single cell infection, therefore, it is important to relate the deformation dynamics to the mechanical properties (such as, bending rigidity and membrane elasticity). In the present study, numerical simulation of a healthy and malaria infected RBC in a constricted channel is analyzed. The flow simulations are carried out using finite sized dissipative particle dynamics (FDPD) method in conjunction with a discrete model that represents the membrane of the RBC. The numerical equivalent of optical tweezers test is validated against the experimental studies. Two different types of constrictions, viz., a converging-diverging type tapered channel and a stenosed microchannel are considered for the simulation. The effect of degree of constriction and the flow rate effect on the RBC is investigated. It was observed that, as the flow rate decreases, the infected RBC completely blocks the micro vessel. The transit time for infected cell drastically increases compared to healthy RBC. Our simulations indicate that, there is a critical flow rate below which infected RBC cannot pass through the micro capillary.

  5. Red blood cell populations and membrane levels of peroxiredoxin 2 as candidate biomarkers to reveal blood doping.

    PubMed

    Marrocco, Cristina; Pallotta, Valeria; D'alessandro, Angelo; Alves, Gilda; Zolla, Lello

    2012-05-01

    Blood doping represents one main trend in doping strategies. Blood doping refers to the practice of boosting the number of red blood cells (RBCs) in the bloodstream in order to enhance athletic performance, by means of blood transfusions, administration of erythropoiesis-stimulating substances, blood substitutes, natural or artificial altitude facilities, and innovative gene therapies. While detection of recombinant EPO and homologous transfusion is already feasible through electrophoretic, mass spectrometry or flow cytometry-based approaches, no method is currently available to tackle doping strategies relying on autologous transfusions. We exploited an in vitro model of autologous transfusion through a 1:10 dilution of concentrated RBCs after 30 days of storage upon appropriate dilution in freshly withdrawn RBCs from the same donor. Western blot towards membrane Prdx2 and Percoll density gradients were exploited to assess their suitability as biomarkers of transfusion. Membrane Prdx2 was visible in day 30 samples albeit not in day 0, while it was still visible in the 1:10 dilution of day 30 in day 0 RBCs. Cell gradients also highlighted changes in the profile of the RBC subpopulations upon dilution of stored RBCs in the fresh ones. From this preliminary in vitro investigation it emerges that Prdx2 and RBC populations might be further tested as candidate biomarkers of blood doping through autologous transfusion, though it is yet to be assessed whether the kinetics in vivo of Prdx2 exposure in the membrane of transfused RBCs will endow a sufficient time-window to allow reliable anti-doping testing.

  6. Red blood cell populations and membrane levels of peroxiredoxin 2 as candidate biomarkers to reveal blood doping

    PubMed Central

    Marrocco, Cristina; Pallotta, Valeria; D’Alessandro, Angelo; Alves, Gilda; Zolla, Lello

    2012-01-01

    Background Blood doping represents one main trend in doping strategies. Blood doping refers to the practice of boosting the number of red blood cells (RBCs) in the bloodstream in order to enhance athletic performance, by means of blood transfusions, administration of erythropoiesis-stimulating substances, blood substitutes, natural or artificial altitude facilities, and innovative gene therapies. While detection of recombinant EPO and homologous transfusion is already feasible through electrophoretic, mass spectrometry or flow cytometry-based approaches, no method is currently available to tackle doping strategies relying on autologous transfusions. Materials and methods. We exploited an in vitro model of autologous transfusion through a 1:10 dilution of concentrated RBCs after 30 days of storage upon appropriate dilution in freshly withdrawn RBCs from the same donor. Western blot towards membrane Prdx2 and Percoll density gradients were exploited to assess their suitability as biomarkers of transfusion. Results Membrane Prdx2 was visible in day 30 samples albeit not in day 0, while it was still visible in the 1:10 dilution of day 30 in day 0 RBCs. Cell gradients also highlighted changes in the profile of the RBC subpopulations upon dilution of stored RBCs in the fresh ones. Discussion. From this preliminary in vitro investigation it emerges that Prdx2 and RBC populations might be further tested as candidate biomarkers of blood doping through autologous transfusion, though it is yet to be assessed whether the kinetics in vivo of Prdx2 exposure in the membrane of transfused RBCs will endow a sufficient time-window to allow reliable anti-doping testing. PMID:22890272

  7. Measuring electrical and mechanical properties of red blood cells with a double optical tweezers

    NASA Astrophysics Data System (ADS)

    Fontes, Adriana; Fernandes, Heloise P.; Barjas-Castro, Maria L.; de Thomaz, André A.; Pozzo, Liliana d. Y.; Barbosa, Luiz C.; Cesar, Carlos L.

    2006-08-01

    The fluid lipid bilayer viscoelastic membrane of red blood cells (RBC) contains antigen glycolproteins and proteins which can interact with antibodies to cause cell agglutination. This is the basis of most of the immunohematologic tests in blood banks and the identification of the antibodies against the erythrocyte antigens is of fundamental importance for transfusional routines. The negative charges of the RBCs creates a repulsive electric (zeta) potential between the cells and prevents their aggregation in the blood stream. The first counterions cloud strongly binded moving together with the RBC is called the compact layer. This report proposes the use of a double optical tweezers for a new procedure for measuring: (1) the apparent membrane viscosity, (2) the cell adhesion, (3) the zeta potential and (4) the compact layer's size of the charges formed around the cell in the electrolytic solution. To measure the membrane viscosity we trapped silica beads strongly attached to agglutinated RBCs and measured the force to slide one RBC over the other as a function of the relative velocity. The RBC adhesion was measured by slowly displacing two RBCs apart until the disagglutination happens. The compact layer's size was measured using the force on the silica bead attached to a single RBC in response to an applied voltage and the zeta potential was obtained by measuring the terminal velocity after releasing the RBC from the optical trap at the last applied voltage. We believe that the methodology here proposed can improve the methods of diagnosis in blood banks.

  8. Use of mouse models to study the mechanisms and consequences of RBC clearance

    PubMed Central

    Hod, E. A.; Arinsburg, S. A.; Francis, R. O.; Hendrickson, J. E.; Zimring, J. C.; Spitalnik, S. L.

    2013-01-01

    Mice provide tractable animal models for studying the pathophysiology of various human disorders. This review discusses the use of mouse models for understanding red-blood-cell (RBC) clearance. These models provide important insights into the pathophysiology of various clinically relevant entities, such as autoimmune haemolytic anaemia, haemolytic transfusion reactions, other complications of RBC transfusions and immunomodulation by Rh immune globulin therapy. Mouse models of both antibody- and non-antibody-mediated RBC clearance are reviewed. Approaches for exploring unanswered questions in transfusion medicine using these models are also discussed. PMID:20345515

  9. CD59 signaling and membrane pores drive Syk-dependent erythrocyte necroptosis

    PubMed Central

    LaRocca, T J; Stivison, E A; Mal-Sarkar, T; Hooven, T A; Hod, E A; Spitalnik, S L; Ratner, A J

    2015-01-01

    Mature erythrocytes (red blood cells (RBCs)) undergo the programmed cell death (PCD) pathway of necroptosis in response to bacterial pore-forming toxins (PFTs) that target human CD59 (hCD59) but not hCD59-independent PFTs. Here, we investigate the biochemical mechanism of RBC necroptosis with a focus on the mechanism of induction and the minimal requirements for such RBC death. Binding or crosslinking of the hCD59 receptor led to Syk-dependent induction of vesiculated morphology (echinocytes) that was associated with phosphorylation of Band 3 and was required for Fas ligand (FasL) release. FasL-dependent phosphorylation of receptor-interacting protein kinase 1 (RIP1) in combination with plasma membrane pore formation was required for execution of RBC necroptosis. RIP1 phosphorylation led to the phosphorylation of RIP3, which was also critical for RBC necroptosis. Notably, RBC necroptosis was mediated by FasL and not by other candidate inducers, including tumor necrosis factor alpha (TNF-α) and TNF-related apoptosis-inducing ligand (TRAIL). Other types of RBC damage, such as eryptotic damage, failed to induce necroptosis when combined with hCD59 crosslinking. This work sheds light on the requirements for this recently discovered PCD in RBCs and provides a clear picture of the biochemical mechanism of induction of RBC necroptosis. PMID:26018734

  10. CD59 signaling and membrane pores drive Syk-dependent erythrocyte necroptosis.

    PubMed

    LaRocca, T J; Stivison, E A; Mal-Sarkar, T; Hooven, T A; Hod, E A; Spitalnik, S L; Ratner, A J

    2015-05-28

    Mature erythrocytes (red blood cells (RBCs)) undergo the programmed cell death (PCD) pathway of necroptosis in response to bacterial pore-forming toxins (PFTs) that target human CD59 (hCD59) but not hCD59-independent PFTs. Here, we investigate the biochemical mechanism of RBC necroptosis with a focus on the mechanism of induction and the minimal requirements for such RBC death. Binding or crosslinking of the hCD59 receptor led to Syk-dependent induction of vesiculated morphology (echinocytes) that was associated with phosphorylation of Band 3 and was required for Fas ligand (FasL) release. FasL-dependent phosphorylation of receptor-interacting protein kinase 1 (RIP1) in combination with plasma membrane pore formation was required for execution of RBC necroptosis. RIP1 phosphorylation led to the phosphorylation of RIP3, which was also critical for RBC necroptosis. Notably, RBC necroptosis was mediated by FasL and not by other candidate inducers, including tumor necrosis factor alpha (TNF-α) and TNF-related apoptosis-inducing ligand (TRAIL). Other types of RBC damage, such as eryptotic damage, failed to induce necroptosis when combined with hCD59 crosslinking. This work sheds light on the requirements for this recently discovered PCD in RBCs and provides a clear picture of the biochemical mechanism of induction of RBC necroptosis.

  11. Erythrocyte NADPH oxidase activity modulated by Rac GTPases, PKC, and plasma cytokines contributes to oxidative stress in sickle cell disease

    PubMed Central

    Pushkaran, Suvarnamala; Konstantinidis, Diamantis G.; Koochaki, Sebastian; Malik, Punam; Mohandas, Narla; Zheng, Yi; Joiner, Clinton H.; Kalfa, Theodosia A.

    2013-01-01

    Chronic inflammation has emerged as an important pathogenic mechanism in sickle cell disease (SCD). One component of this inflammatory response is oxidant stress mediated by reactive oxygen species (ROS) generated by leukocytes, endothelial cells, plasma enzymes, and sickle red blood cells (RBC). Sickle RBC ROS generation has been attributed to sickle hemoglobin auto-oxidation and Fenton chemistry reactions catalyzed by denatured heme moieties bound to the RBC membrane. In this study, we demonstrate that a significant part of ROS production in sickle cells is mediated enzymatically by NADPH oxidase, which is regulated by protein kinase C, Rac GTPase, and intracellular Ca2+ signaling within the sickle RBC. Moreover, plasma from patients with SCD and isolated cytokines, such as transforming growth factor β1 and endothelin-1, enhance RBC NADPH oxidase activity and increase ROS generation. ROS-mediated damage to RBC membrane components is known to contribute to erythrocyte rigidity and fragility in SCD. Erythrocyte ROS generation, hemolysis, vaso-occlusion, and the inflammatory response to tissue damage may therefore act in a positive-feedback loop to drive the pathophysiology of sickle cell disease. These findings suggest a novel pathogenic mechanism in SCD and may offer new therapeutic targets to counteract inflammation and RBC rigidity and fragility in SCD. PMID:23349388

  12. Effects of phloretin on lipid organization in the erythrocyte membrane as measured by EPR

    NASA Astrophysics Data System (ADS)

    Abumrad, Nada A.; Perkins, Ray C.; Dalton, Larry R.; Park, Charles R.; Park, Jane H.

    Phloretin is a lipophilic compound which has been widely studied as a broad spectrum effector of metabolite transport in red blood cells (RBC). Phloretin effects on the organization of lipids in the RBC membrane are investigated using the spin-labeled fatty acids, 5 and 16-nitroxyl stearate (5-NS and 16-NS, respectively). Phloretin at different concentrations produced biphasic effects on the lineshape of the EPR response from 16-NS-labeled RBC. The dependence of these changes on the flat cell orientation with respect to the magnetic field suggested that phloretin promoted lipid order at low concentrations (5 to 40 μ M) and disorder at high concentrations (40 to 250 μ M). The biphasic effects of phloretin occurred at concentrations which parallel its dual actions on metabolite transfer. Phloretin generally inhibits transport (protein-mediated) and stimulates diffusion (lipid-mediated) processes. The spectroscopic effects were best characterized through second-harmonic, in-phase detection. The possible contribution of other factors to the spectroscopic changes is discussed. When RBC were spin labeled with 5-NS, higher concentrations of the probe were required for adequate detection and only monophasic effects of phoretin were observed. The results suggest that membrane lipids are important in phloretin effects on transport and diffusion processes.

  13. On the shape memory of red blood cells

    NASA Astrophysics Data System (ADS)

    Cordasco, Daniel; Bagchi, Prosenjit

    2017-04-01

    Red blood cells (RBCs) undergo remarkably large deformations when subjected to external forces but return to their biconcave discoid resting shape as the forces are withdrawn. In many experiments, such as when RBCs are subjected to a shear flow and undergo the tank-treading motion, the membrane elements are also displaced from their original (resting) locations along the cell surface with respect to the cell axis, in addition to the cell being deformed. A shape memory is said to exist if after the flow is stopped the RBC regains its biconcave shape and the membrane elements also return to their original locations. The shape memory of RBCs was demonstrated by Fischer ["Shape memory of human red blood cells," Biophys. J. 86, 3304-3313 (2004)] using shear flow go-and-stop experiments. Optical tweezer and micropipette based stretch-relaxation experiments do not reveal the complete shape memory because while the RBC may be deformed, the membrane elements are not significantly displaced from their original locations with respect to the cell axis. Here we present the first three-dimensional computational study predicting the complete shape memory of RBCs using shear flow go-and-stop simulations. The influence of different parameters, namely, membrane shear elasticity and bending rigidity, membrane viscosity, cytoplasmic and suspending fluid viscosity, as well as different stress-free states of the RBC is studied. For all cases, the RBCs always exhibit shape memory. The complete recovery of the RBC in shear flow go-and-stop simulations occurs over a time that is orders of magnitude longer than that for optical tweezer and micropipette based relaxations. The response is also observed to be more complex and composed of widely disparate time scales as opposed to only one time scale that characterizes the optical tweezer and micropipette based relaxations. We observe that the recovery occurs in three phases: a rapid compression of the RBC immediately after the flow is stopped

  14. Three-dimensional motion and deformation of a red blood cell in bifurcated microvessels

    NASA Astrophysics Data System (ADS)

    Ye, Ting; Peng, Lina; Li, Yu

    2018-02-01

    Microvessels are generally not simple straight tubes, but rather they continually bifurcate (namely, diverging bifurcation) and merge with other microvessels (namely, converging bifurcation). This paper presents a simulation study on the three-dimensional motion and deformation of a red blood cell (RBC) in a bifurcated microvessel with both diverging and converging bifurcations. The motion of the fluids inside and outside of the RBC is modeled by smooth dissipative particle dynamics. The RBC membrane is modeled as a triangular network, having the ability to not only resist the stretching and bending deformations, but also to conserve the RBC volume and surface area. The bifurcation configurations have been studied, including the bifurcated angle and the branch diameter, as well as the RBC properties, including the initial shape, shear modulus, and bending modulus. The simulation results show that the RBC deformation can be divided into five stages, when the RBC flows through a diverging-converging bifurcated microvessel. In these five stages, the RBCs have similar deformation trends but different deformation indices, subject to different bifurcation configurations or different RBC properties. If the shear modulus is large enough, the RBC membrane presents several folds; if the bending modulus is large enough, the RBC loses the symmetry completely with the long shape. These results are helpful in understanding the motion and deformation of healthy or unhealthy cells in blood microcirculation.

  15. Reversal of anemia with allogenic RBC transfusion prevents post-cardiopulmonary bypass acute kidney injury in swine

    PubMed Central

    Patel, Nishith N.; Lin, Hua; Toth, Tibor; Welsh, Gavin I.; Jones, Ceri; Ray, Paramita; Satchell, Simon C.; Sleeman, Philippa; Angelini, Gianni D.

    2011-01-01

    Anemia during cardiopulmonary bypass (CPB) is strongly associated with acute kidney injury in clinical studies; however, reversal of anemia with red blood cell (RBC) transfusions is associated with further renal injury. To understand this paradox, we evaluated the effects of reversal of anemia during CPB with allogenic RBC transfusion in a novel large-animal model of post-cardiac surgery acute kidney injury with significant homology to that observed in cardiac surgery patients. Adult pigs undergoing general anesthesia were allocated to a Sham procedure, CPB alone, Sham+RBC transfusion, or CPB+RBC transfusion, with recovery and reassessment at 24 h. CPB was associated with dilutional anemia and caused acute kidney injury in swine characterized by renal endothelial dysfunction, loss of nitric oxide (NO) bioavailability, vasoconstriction, medullary hypoxia, cortical ATP depletion, glomerular sequestration of activated platelets and inflammatory cells, and proximal tubule epithelial cell stress. RBC transfusion in the absence of CPB also resulted in renal injury. This was characterized by endothelial injury, microvascular endothelial dysfunction, platelet activation, and equivalent cortical tubular epithelial phenotypic changes to those observed in CPB pigs, but occurred in the absence of severe intrarenal vasoconstriction, ATP depletion, or reductions in creatinine clearance. In contrast, reversal of anemia during CPB with RBC transfusion prevented the reductions in creatinine clearance, loss of NO bioavailability, platelet activation, inflammation, and epithelial cell injury attributable to CPB although it did not prevent the development of significant intrarenal vasoconstriction and endothelial dysfunction. In conclusion, contrary to the findings of observational studies in cardiac surgery, RBC transfusion during CPB protects pigs against acute kidney injury. Our study underlines the need for translational research into indications for transfusion and prevention

  16. Measurement of red blood cell mechanics during morphological changes

    PubMed Central

    Park, YongKeun; Best, Catherine A.; Badizadegan, Kamran; Dasari, Ramachandra R.; Feld, Michael S.; Kuriabova, Tatiana; Henle, Mark L.; Levine, Alex J.; Popescu, Gabriel

    2010-01-01

    The human red blood cell (RBC) membrane, a fluid lipid bilayer tethered to an elastic 2D spectrin network, provides the principal control of the cell’s morphology and mechanics. These properties, in turn, influence the ability of RBCs to transport oxygen in circulation. Current mechanical measurements of RBCs rely on external loads. Here we apply a noncontact optical interferometric technique to quantify the thermal fluctuations of RBC membranes with 3 nm accuracy over a broad range of spatial and temporal frequencies. Combining this technique with a new mathematical model describing RBC membrane undulations, we measure the mechanical changes of RBCs as they undergo a transition from the normal discoid shape to the abnormal echinocyte and spherical shapes. These measurements indicate that, coincident with this morphological transition, there is a significant increase in the membrane’s shear, area, and bending moduli. This mechanical transition can alter cell circulation and impede oxygen delivery. PMID:20351261

  17. Vitamin E-coated polysulfone membrane improved red blood cell antioxidant status in hemodialysis patients.

    PubMed

    Bargnoux, Anne-Sophie; Cristol, Jean-Paul; Jaussent, Isabelle; Chalabi, Lotfi; Bories, Pierre; Dion, Jean-Jacques; Henri, Patrick; Delage, Martine; Dupuy, Anne-Marie; Badiou, Stéphanie; Canaud, Bernard; Morena, Marion

    2013-01-01

    Oxidative stress has emerged as a strong pathogenic cofactor implicated in the development of long-term complications in hemodialysis (HD) patients, such as anemia, and as a major component of the malnutrition inflammation complex syndrome. This prospective multicenter study aimed at evaluating the short-term effects of the new vitamin E (vitE)-coated polysulfone (PS) membrane (VitabranE) on biocompatibility performances and anemia in HD patients. After a 3-month washout period with a high-flux synthetic dialyzer, 43 HD patients were switched to a vitE-PS dialyzer. Sampling was performed at baseline (corresponding to the end of the washout period) and after 1, 2 and 3 months of treatment. Oxidative stress status, as well as inflammatory parameters, was investigated at the end of each study period. Hemoglobin levels and administered doses of recombinant human erythropoietin or epoetin (EPO) were available in each center. The use of vitE-coated membranes for 3 months was not associated with any change in inflammatory parameters. By contrast, vitE-PS dialyzer resulted in a progressive increase in red blood cell (RBC) vitE concentration and in RBC superoxide dismutase activity. A concomitant progressive significant decrease in advanced oxidation protein product concentration at 2 months was observed, suggesting a preventive effect on oxidative stress. Finally, a significant decrease of the erythropoietin resistance index was obtained after 3 months of treatment. Use of the vitE-PS membrane during a short period improves erythrocyte antioxidant defense mechanisms and seems to lead to a reduction in EPO requirements in HD patients.

  18. In vitro measures of membrane changes reveal differences between red blood cells stored in SAGM and AS-1 additive solutions: a paired study

    PubMed Central

    Sparrow, Rosemary L; Sran, Amrita; Healey, Geraldine; Veale, Margaret F; Norris, Philip J

    2014-01-01

    Background Saline-Adenine-Glucose-Mannitol (SAGM) and a variant solution, AS-1 have been used for over 30 years to preserve red blood cells (RBCs). Reputedly these RBC components have similar quality, although no paired study has been reported. To determine whether differences exist, a paired study of SAGM-RBCs and AS-1-RBCs was conducted to identify membrane changes, including microparticle (MP) quantitation and in vitro RBC-endothelial cell (EC) interaction. Study Design and Methods Two whole blood packs were pooled-and-split and RBCs prepared (n=6 pairs). One pack was suspended in SAGM and one in AS-1. Samples were collected during 42 days of refrigerated storage. RBC shape/size, glycophorin A (GPA)+ and phosphatidylserine (PS)+ MPs were measured by flow cytometry. RBC adhesion to ECs was determined by an in vitro flow perfusion assay. Routine parameters (pH, hemolysis) were also measured. Results Compared to SAGM-RBCs, AS-1-RBCs had lower hemolysis (p<0.04), lower GPA+ MPs (p<0.03) and lower PS+ MPs (p<0.03) from day 14 onwards. AS-1-RBCs had higher (p<0.02) side scatter from day 28 onwards, compared to SAGM-RBCs. SAGM-RBCs were more adherent to ECs at day 28 of storage compared to AS-1 RBCs (p=0.04), but reversed at day 42 (p=0.02). No significant differences in forward scatter or pH were found. Conclusion SAGM-RBCs lose more membrane during storage. SAGM-RBCs had increased adherence to ECs at day 28 of storage, while AS-1-RBCs were more adherent at day 42. The effect of these differences on the function and survival of SAGM-RBCs and AS-1-RBCs following transfusion remains to be determined. PMID:23869602

  19. Residual blood loss in single use dialyzers: effect of different membranes and flux.

    PubMed

    Kalocheretis, P; Vlamis, I; Belesi, C; Makriniotou, I; Zerbala, S; Savidou, E; Zorbas, S; Arvanitis, N; Iatrou, C

    2006-03-01

    The aim of this study was to evaluate the residual blood loss in new type single use dialyzers under the usually prevailing conditions during hemodialysis and to investigate whether or not this loss is dependent on dialyzer membrane composition or flux characteristics. In 158 hemodialysis (HD) patients, 158 single used dialyzers were studied in corresponding HD sessions. 52/158 dialyzers were made from modified cellulose (acetate, CA or triacetate, CTA) membrane and 106 from synthetic ones (58 with ethyl-vinyl-alcohol (EVAL), 48 with polyacrilonitrile (AN69)). Of those dialyzers 85/158 (58 EVAL+27CA) were low flux (LF) while the other 73 were high flux (HF). Patients underwent 4 hour HD sessions and at the end of the session blood was drawn for the measurement of hematocrit (Ht) and hemoglobin (Hb). Additionally, after the end of dialysis the used dialyzers were rinsed with 1000 mL of 0.05% NH(3) solution in distilled water. The wash was collected and subsequently Hb was measured using the benzidine method. From the volume of the solution and its concentration of Hb, total Hb of the solution was measured and blood loss in terms of red blood cell (RBC) volume was estimated by the use of the formula: RBC (mL) = Total Hb (g) in the solution x patient's Ht (ml/dL) / Patient's Hb (g/dL). For results to be comparable between dialyzers, RBC volume/m(2) of dialyzer membrane was expressed. In 5/158 patients blood loss was also estimated in 6 consecutive HD sessions using the same type of dialyzer. For the sum of the dialyzers, blood loss / dialyzer in terms of RBC volume, expressed as median (range), was 0.978 mL (0.01-23.9). There was statistically significant (p<0.001 or p<0.05) higher blood loss with the use of AN69 dialyzer than with the other three. RBC HF >RBC LF (p<0.001) constrained the first group of patients to use a 6% higher dosage of ferrum and 3.5% higher dosage of erythropoietin than the other group to achieve the optimal hemoglobulin values. No difference

  20. Evaluation of overnight hold of whole blood at room temperature before component processing: effect of red blood cell (RBC) additive solutions on in vitro RBC measures.

    PubMed

    van der Meer, Pieter F; Cancelas, Jose A; Cardigan, Rebecca; Devine, Dana V; Gulliksson, Hans; Sparrow, Rosemary L; Vassallo, Ralph R; de Wildt-Eggen, Janny; Baumann-Baretti, Bärbel; Hess, John R

    2011-01-01

    Whole blood (WB) can be held at room temperature (18-25°C) up to 8 hours after collection; thereafter the unit must be refrigerated, rendering it unsuitable for platelet (PLT) production. Overnight hold at room temperature before processing has logistic advantages, and we evaluated this process in an international multicenter study for both buffy coat (BC)- and PLT-rich plasma (PRP)-based blood components and compared three red blood cell (RBC) additive solutions (ASs) for their ability to offset effects of overnight hold. Nine centers participated; seven used the BC method, and two used the PRP method. Four WB units were pooled and split; 1 unit was processed less than 8 hours from collection (Group A), and the other three (Groups B, C, and D) were held at room temperature and processed after 24 to 26 hours. RBCs in Groups A and B were resuspended in saline-adenine-glucose-mannitol, Group C in phosphate-adenine-guanosine-glucose-saline-mannitol, and Group D in ErythroSol-4 RBCs were stored at 2 to 6°C for 49 days. RBCs from overnight-held WB had lower 2,3-diphosphoglycerate (2,3-DPG) and higher adenosine triphosphate (ATP). At the end of storage there were no differences between groups, apart from a slightly higher hemolysis in Group B. ErythroSol-4 showed a slightly higher initial ATP and 2,3-DPG content, but at the end of storage no differences were found. Overnight hold of WB before processing has no lasting deleterious effects on in vitro quality of subsequently prepared components. The use of different RBC ASs did not appear to offer significant advantages in terms of RBC quality at the end, regardless of the processing method. © 2010 American Association of Blood Banks.

  1. Effects of Poloxamer 188 on red blood cell membrane properties in sickle cell anaemia.

    PubMed

    Sandor, Barbara; Marin, Mickaël; Lapoumeroulie, Claudine; Rabaï, Miklos; Lefevre, Sophie D; Lemonne, Nathalie; El Nemer, Wassim; Mozar, Anaïs; Français, Olivier; Le Pioufle, Bruno; Connes, Philippe; Le Van Kim, Caroline

    2016-04-01

    Vaso-occlusive crisis (VOC) is the main acute complication in sickle cell anaemia (SS) and several clinical trials are investigating different drugs to improve the clinical severity of SS patients. A phase III study is currently exploring the profit of Velopoloxamer in SS during VOCs. We analysed, in-vitro, the effect of poloxamer (P188) on red blood cell (RBC) properties by investigating haemorheology, mechanical and adhesion functions using ektacytometry, microfluidics and dynamic adhesion approaches, respectively. We show that poloxamer significantly reduces blood viscosity, RBC aggregation and adhesion to endothelial cells, supporting the beneficial use of this molecule in SS therapy. © 2016 John Wiley & Sons Ltd.

  2. Red blood cell dynamics: from cell deformation to ATP release.

    PubMed

    Wan, Jiandi; Forsyth, Alison M; Stone, Howard A

    2011-10-01

    The mechanisms of red blood cell (RBC) deformation under both static and dynamic, i.e., flow, conditions have been studied extensively since the mid 1960s. Deformation-induced biochemical reactions and possible signaling in RBCs, however, were proposed only fifteen years ago. Therefore, the fundamental relationship between RBC deformation and cellular signaling dynamics i.e., mechanotransduction, remains incompletely understood. Quantitative understanding of the mechanotransductive pathways in RBCs requires integrative studies of physical models of RBC deformation and cellular biochemical reactions. In this article we review the physical models of RBC deformation, spanning from continuum membrane mechanics to cellular skeleton dynamics under both static and flow conditions, and elaborate the mechanistic links involved in deformation-induced ATP release. This journal is © The Royal Society of Chemistry 2011

  3. Cell Membrane Softening in Cancer Cells

    NASA Astrophysics Data System (ADS)

    Schmidt, Sebastian; Händel, Chris; Käs, Josef

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

  4. Hydroxycarbamide-Induced Changes in E/beta Thalassemia Red Blood Cells

    PubMed Central

    Sylvia, Singer T.; Elliott, Vichinsky; Sandra, Larkin; Nancy, Olivieri; Nancy, Sweeters; Frans, Kuypers A.

    2010-01-01

    In thalassemia, fetal hemoglobin (HbF) augmentation with hydroxycarbamide (also known as hydroxyurea) is not always successful. The expected parallel effects on RBC membrane deformability, cell hydration and membrane phospholipid organization, all important for extending RBC life span and increasing Hb, have been infrequently examined. We analyzed these characteristics in 15 non-transfused E/β 0 thalassemia patients treated with HU (mean 10.2 months). Membrane deformability and cell hydration mildly improved in association with increased HbF levels approaching statistical significance (r = 0.51, p=0.06). All measures improved considerably splenectomized patients. These findings underscore the disappointing results of hydroxyurea treatment in clinical trials; and the importance of examining the effect on red cell characteristics for the development and understanding of HbF-enhancing agents. PMID:18821710

  5. Red blood cells serve as intravascular carriers of myeloperoxidase.

    PubMed

    Adam, Matti; Gajdova, Silvie; Kolarova, Hana; Kubala, Lukas; Lau, Denise; Geisler, Anne; Ravekes, Thorben; Rudolph, Volker; Tsao, Philip S; Blankenberg, Stefan; Baldus, Stephan; Klinke, Anna

    2014-09-01

    Myeloperoxidase (MPO) is a heme enzyme abundantly expressed in polymorphonuclear neutrophils. MPO is enzymatically capable of catalyzing the generation of reactive oxygen species (ROS) and the consumption of nitric oxide (NO). Thus MPO has both potent microbicidal and, upon binding to the vessel wall, pro-inflammatory properties. Interestingly, MPO - a highly cationic protein - has been shown to bind to both endothelial cells and leukocyte membranes. Given the anionic surface charge of red blood cells, we investigated binding of MPO to erythrocytes. Red blood cells (RBCs) derived from patients with elevated MPO plasma levels showed significantly higher amounts of MPO by flow cytometry and ELISA than healthy controls. Heparin-induced MPO-release from patient-derived RBCs was significantly increased compared to controls. Ex vivo experiments revealed dose and time dependency for MPO-RBC binding, and immunofluorescence staining as well as confocal microscopy localized MPO-RBC interaction to the erythrocyte plasma membrane. NO-consumption by RBC-membrane fragments (erythrocyte "ghosts") increased with incrementally greater concentrations of MPO during incubation, indicating preserved catalytic MPO activity. In vivo infusion of MPO-loaded RBCs into C57BL/6J mice increased local MPO tissue concentrations in liver, spleen, lung, and heart tissue as well as within the cardiac vasculature. Further, NO-dependent relaxation of aortic rings was altered by RBC bound-MPO and systemic vascular resistance significantly increased after infusion of MPO-loaded RBCs into mice. In summary, we find that MPO binds to RBC membranes in vitro and in vivo, is transported by RBCs to remote sites in mice, and affects endothelial function as well as systemic vascular resistance. RBCs may avidly bind circulating MPO, and act as carriers of this leukocyte-derived enzyme. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Pathogen inactivation by riboflavin and ultraviolet light illumination accelerates the red blood cell storage lesion and promotes eryptosis.

    PubMed

    Qadri, Syed M; Chen, Deborah; Schubert, Peter; Perruzza, Darian L; Bhakta, Varsha; Devine, Dana V; Sheffield, William P

    2017-03-01

    Pathogen reduction treatment using riboflavin and ultraviolet light illumination (Mirasol) effectively reduces the risk of transfusion-transmitted infections. This treatment is currently licensed for only platelets and plasma products, while its application to whole blood (WB) to generate pathogen-inactivated red blood cells (RBCs) is under development. RBC storage lesion, constituting numerous morphologic and biochemical changes, influences RBC quality and limits shelf life. Stored RBCs further show enhanced susceptibility to RBC programmed cell death (eryptosis) characterized by increased cytosolic Ca 2+ -provoked membrane phosphatidylserine (PS) externalization. Using a "pool-and-split" approach, we examined multiple variables of RBC storage lesion and eryptosis in RBC units, derived from Mirasol-treated or untreated WB, after 4 to 42 days of storage, under blood bank conditions. In comparison to untreated RBC units, Mirasol treatment significantly altered membrane microvesiculation, supernatant hemoglobin, osmotic fragility, and intracellular adenosine triphosphate levels but did not influence membrane CD47 expression and 2,3-diphosphoglycerate levels. Mirasol-treated RBCs showed significantly higher PS exposure after 42, but not after not more than 21, days of storage, which was accompanied by enhanced cytosolic Ca 2+ activity, ceramide abundance, and oxidative stress, but not p38 kinase activation. Mirasol treatment significantly augmented PS exposure, Ca 2+ entry, and protein kinase C activation after energy depletion, a pathophysiologic cell stressor. Mirasol-treated RBCs were, however, more resistant to cell shrinkage. Prolonged storage of Mirasol-treated RBCs significantly increases the proportion of eryptotic RBCs, while even short-term storage enhances the susceptibility of RBCs to stress-induced eryptosis, which could reduce posttransfusion RBC recovery in patients. © 2016 AABB.

  7. A novel RNA binding protein affects rbcL gene expression and is specific to bundle sheath chloroplasts in C4 plants

    PubMed Central

    2013-01-01

    Background Plants that utilize the highly efficient C4 pathway of photosynthesis typically possess kranz-type leaf anatomy that consists of two morphologically and functionally distinct photosynthetic cell types, the bundle sheath (BS) and mesophyll (M) cells. These two cell types differentially express many genes that are required for C4 capability and function. In mature C4 leaves, the plastidic rbcL gene, encoding the large subunit of the primary CO2 fixation enzyme Rubisco, is expressed specifically within BS cells. Numerous studies have demonstrated that BS-specific rbcL gene expression is regulated predominantly at post-transcriptional levels, through the control of translation and mRNA stability. The identification of regulatory factors associated with C4 patterns of rbcL gene expression has been an elusive goal for many years. Results RLSB, encoded by the nuclear RLSB gene, is an S1-domain RNA binding protein purified from C4 chloroplasts based on its specific binding to plastid-encoded rbcL mRNA in vitro. Co-localized with LSU to chloroplasts, RLSB is highly conserved across many plant species. Most significantly, RLSB localizes specifically to leaf bundle sheath (BS) cells in C4 plants. Comparative analysis using maize (C4) and Arabidopsis (C3) reveals its tight association with rbcL gene expression in both plants. Reduced RLSB expression (through insertion mutation or RNA silencing, respectively) led to reductions in rbcL mRNA accumulation and LSU production. Additional developmental effects, such as virescent/yellow leaves, were likely associated with decreased photosynthetic function and disruption of associated signaling networks. Conclusions Reductions in RLSB expression, due to insertion mutation or gene silencing, are strictly correlated with reductions in rbcL gene expression in both maize and Arabidopsis. In both plants, accumulation of rbcL mRNA as well as synthesis of LSU protein were affected. These findings suggest that specific accumulation

  8. From artificial red blood cells, oxygen carriers, and oxygen therapeutics to artificial cells, nanomedicine, and beyond

    PubMed Central

    Chang, Thomas M. S.

    2013-01-01

    The first experimental artificial red blood cells have all three major functions of red blood cells (rbc). However, the first practical one is a simple polyhemoglobin (PolyHb) that only has an oxygen-carrying function. This is now in routine clinical use in South Africa and Russia. An oxygen carrier with antioxidant functions, PolyHb-catalase-superoxide dismutase, can fulfill two of the three functions of rbc. Even more complete is one with all three functions of rbc in the form of PolyHb-catalase-superoxide dismutase-carbonic anhydrase. The most advanced ones are nanodimension artificial rbc with either PEG-lipid membrane or PEG-PLA polymermembrane. Extensions in to oxygen therapeutics include a PolyHb-tyrosinase that suppresses the growth of melanoma in a mice model. Another is a PolyHb-fibrinogen that is an oxygen carrier with platelet-like function. Research has now extended well beyond the original research on artificial rbc into many areas of artificial cells. These include nanoparticles, nanotubules, lipid vesicles, liposomes, polymer-tethered lipid vesicles, polymersomes, microcapsules, bioencapsulation, nanocapules, macroencapsulation, synthetic cells, and others. These are being used in nanotechnology, nanomedicine, regenerative medicine, enzyme/gene therapy, cell/stem cell therapy, biotechnology, drug delivery, hemoperfusion, nanosensers, and even by some groups in agriculture, industry, aquatic culture, nanocomputers, and nanorobotics. PMID:22409281

  9. Mechanical clearance of red blood cells by the human spleen: Potential therapeutic applications of a biomimetic RBC filtration method.

    PubMed

    Duez, J; Holleran, J P; Ndour, P A; Pionneau, C; Diakité, S; Roussel, C; Dussiot, M; Amireault, P; Avery, V M; Buffet, P A

    2015-08-01

    During their lifespan, circulating RBC are frequently checked for their deformability. This mechanical quality control operates essentially in the human spleen. RBC unable to squeeze though narrow splenic slits are retained and cleared from the blood circulation. Under physiological conditions this prevents microvessels from being clogged by senescent, rigid RBC. Retention of poorly deformable RBC is an important determinant of pathogenesis in malaria and may also impact the clinical benefit of transfusion. Modulating the splenic retention of RBC has already been proposed to support therapeutic approaches in these research fields. To this aim, the development of microplates for high throughput filtration of RBC through microsphere layers (microplate-based microsphiltration) has been undertaken. This review focuses on potential therapeutic applications provided by this technology in malaria chemotherapy and transfusion. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  10. Membrane lipidomics in schizophrenia patients: a correlational study with clinical and cognitive manifestations.

    PubMed

    Tessier, C; Sweers, K; Frajerman, A; Bergaoui, H; Ferreri, F; Delva, C; Lapidus, N; Lamaziere, A; Roiser, J P; De Hert, M; Nuss, P

    2016-10-04

    Schizophrenia is a severe mental condition in which several lipid abnormalities-either structural or metabolic-have been described. We tested the hypothesis that an abnormality in membrane lipid composition may contribute to aberrant dopamine signaling, and thereby symptoms and cognitive impairment, in schizophrenia (SCZ) patients. Antipsychotic-medicated and clinically stable SCZ outpatients (n=74) were compared with matched healthy subjects (HC, n=40). A lipidomic analysis was performed in red blood cell (RBC) membranes examining the major phospholipid (PL) classes and their associated fatty acids (FAs). Clinical manifestations were examined using the positive and negative syndrome scale (PANSS). Cognitive function was assessed using the Continuous Performance Test, Salience Attribution Test and Wisconsin Card Sorting Test. Sphingomyelin (SM) percentage was the lipid abnormality most robustly associated with a schizophrenia diagnosis. Two groups of patients were defined. The first group (SCZ c/SM-) is characterized by a low SM membrane content. In this group, all other PL classes, plasmalogen and key polyunsaturated FAs known to be involved in brain function, were significantly modified, identifying a very specific membrane lipid cluster. The second patient group (SCZ c/SM+) was similar to HCs in terms of RBC membrane SM composition. Compared with SCZ c/SM+, SCZ c/SM- patients were characterized by significantly more severe PANSS total, positive, disorganized/cognitive and excited psychopathology. Cognitive performance was also significantly poorer in this subgroup. These data show that a specific RBC membrane lipid cluster is associated with clinical and cognitive manifestations of dopamine dysfunction in schizophrenia patients. We speculate that this membrane lipid abnormality influences presynaptic dopamine signaling.

  11. Modeling malaria infected cells in microcirculation

    NASA Astrophysics Data System (ADS)

    Raffiee, Amir Hossein; Dabiri, Sadegh; Motavalizadeh Ardekani, Arezoo

    2016-11-01

    Plasmodim (P.) falciparum is one of the deadliest types of malaria species that invades healthy red blood cells (RBC) in human blood flow. This parasite develops through 48-hour intra-RBC process leading to significant morphological and mechanical (e.g., stiffening) changes in RBC membrane. These changes have remarkable effects on blood circulation such as increase in flow resistance and obstruction in microcirculation. In this work a computational framework is developed to model RBC suspension in blood flow using front-tracking technique. The present study focuses on blood flow behavior under normal and infected circumstances and predicts changes in blood rheology for different levels of parasitemia and hematocrit. This model allows better understanding of blood flow circulation up to a single cell level and provides us with realistic and deep insight into hematologic diseases such as malaria.

  12. The proportion of total C18:1 trans-fatty acids in red blood cell membranes relates to carotid plaque prevalence.

    PubMed

    Herreras, Zoe; Cofán, Montserrat; Catalan, Marta; Calvo, Carlos; Pinyol, Montserrat; Amor, Antonio J; Gilabert, Rosa; Ros, Emilio; Sala-Vila, Aleix; Ortega, Emilio

    2016-12-01

    Consistent evidence supports the pro-atherogenic properties of dietary trans-fatty acids (TFAs). However, there are no clinical data on TFA intake and atheroma plaque. We cross sectionally investigated whether the proportion of total C18:1 TFA in red blood cells (RBCs), which mirrors dietary TFA intake, independently relates to carotid plaque prevalence in subjects with new-onset type 2 diabetes mellitus without prior cardiovascular disease (n=101, 56% men, mean age 61 years) and age- and sex-matched controls (n=96). RBC fatty acid composition was determined by gas chromatography. Plaque (defined as carotid intima-media thickness ≥1.5 mm) was sonographically assessed at three bilateral carotid segments. In multivariate models adjusting for group (diabetes or control) and classical cardiovascular risk factors, for each 0.1% increase in RBC total C18:1 TFA isomers, plaque prevalence increased by 53% (P=.002). In contrast, for each 0.1% increase in RBC alpha-linolenic acid, the vegetable omega-3 fatty acid, plaque prevalence decreased by 43% (P<.001). We conclude that the RBC membrane proportion of total C18:1 TFA, considered a proxy of intake, directly relates to the ultrasound feature that best predicts future cardiovascular events. Our findings support current recommendations to limit TFA intake for cardiovascular health promotion. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Multiscale modelling of Flow-Induced Blood Cell Damage

    NASA Astrophysics Data System (ADS)

    Liu, Yaling; Sohrabi, Salman

    2017-11-01

    We study red blood cell (RBC) damage and hemolysis at cellular level. Under high shear rates, pores form on RBC membranes through which hemoglobin (Hb) leaks out and increases free Hb content of plasma leading to hemolysis. By coupling lattice Boltzmann and spring connected network models through immersed boundary method, we estimate hemolysis of a single RBC under various shear rates. The developed cellular damage model can be used as a predictive tool for hydrodynamic and hematologic design optimization of blood-wetting medical devices.

  14. Identification of Rubisco rbcL and rbcS in Camellia oleifera and their potential as molecular markers for selection of high tea oil cultivars.

    PubMed

    Chen, Yongzhong; Wang, Baoming; Chen, Jianjun; Wang, Xiangnan; Wang, Rui; Peng, Shaofeng; Chen, Longsheng; Ma, Li; Luo, Jian

    2015-01-01

    Tea oil derived from seeds of Camellia oleifera Abel. is high-quality edible oil in China. This study isolated full-length cDNAs of Rubisco subunits rbcL and rbcS from C. oleifera. The rbcL has 1,522 bp with a 1,425 bp coding region, encoding 475 amino acids; and the rbcS has 615 bp containing a 528 bp coding region, encoding 176 amino acids. The expression level of the two genes, designated as Co-rbcL and Co-rbcS, was determined in three C. oleifera cultivars: Hengchong 89, Xianglin 1, and Xianglin 14 whose annual oil yields were 546.9, 591.4, and 657.7 kg ha(-1), respectively. The Co-rbcL expression in 'Xianglin 14' was significantly higher than 'Xianglin 1', and 'Xianglin 1' was greater than 'Hengchong 89'. The expression levels of Co-rbcS in 'Xianglin 1' and 'Xianglin 14' were similar but were significantly greater than in 'Hengchong 89'. The net photosynthetic rate of 'Xianglin 14' was significantly higher than 'Xianglin 1', and 'Xianglin 1' was higher than 'Hengchong 89'. Pearson's correlation analysis showed that seed yields and oil yields were highly correlated with the expression level of Co-rbcL at P < 0.001 level; and the expression of Co-rbcS was correlated with oil yield at P < 0.01 level. Net photosynthetic rate was also correlated with oil yields and seed yields at P < 0.001 and P < 0.01 levels, respectively. Our results suggest that Co-rbcS and Co-rbcL in particular could potentially be molecular markers for early selection of high oil yield cultivars. In combination with the measurement of net photosynthetic rates, the early identification of potential high oil production cultivars would significantly shorten plant breeding time and increase breeding efficiency.

  15. Vesiculation of healthy and defective red blood cells

    NASA Astrophysics Data System (ADS)

    Li, He; Lykotrafitis, George

    2015-07-01

    Vesiculation of mature red blood cells (RBCs) contributes to removal of defective patches of the erythrocyte membrane. In blood disorders, which are related to defects in proteins of the RBC membrane, vesiculation of the plasma membrane is intensified. Several hypotheses have been proposed to explain RBC vesiculation but the exact underlying mechanisms and what determines the sizes of the vesicles are still not completely understood. In this work, we apply a two-component coarse-grained molecular dynamics RBC membrane model to study how RBC vesiculation is controlled by the membrane spontaneous curvature and by lateral compression of the membrane. Our simulation results show that the formation of small homogeneous vesicles with a diameter less than 40 nm can be attributed to a large spontaneous curvature of membrane domains. On the other hand, compression on the membrane can cause the formation of vesicles with heterogeneous composition and with sizes comparable with the size of the cytoskeleton corral. When spontaneous curvature and lateral compression are simultaneously considered, the compression on the membrane tends to facilitate formation of vesicles originating from curved membrane domains. We also simulate vesiculation of RBCs with membrane defects connected to hereditary elliptocytosis (HE) and to hereditary spherocytosis (HS). When the vertical connectivity between the lipid bilayer and the membrane skeleton is elevated, as in normal RBCs, multiple vesicles are shed from the compressed membrane with diameters similar to the cytoskeleton corral size. In HS RBCs, where the connectivity between the lipid bilayer and the cytoskeleton is reduced, larger-size vesicles are released under the same compression ratio as in normal RBCs. Lastly, we find that vesicles released from HE RBCs can contain cytoskeletal filaments due to fragmentation of the membrane skeleton while vesicles released from the HS RBCs are depleted of cytoskeletal filaments.

  16. Experimental prestorage filtration removes antibodies and decreases lipids in RBC supernatants mitigating TRALI in vivo

    PubMed Central

    Kelher, Marguerite R.; Khan, Samina Y.; LaSarre, Monica; West, F. Bernadette; Land, Kevin J.; Mish, Barbara; Ceriano, Linda; Sowemimo-Coker, Samuel

    2014-01-01

    Transfusion-related acute lung injury (TRALI) remains a significant cause of transfusion-related mortality with red cell transfusion. We hypothesize that prestorage filtration may reduce proinflammatory activity in the red blood cell (RBC) supernatant and prevent TRALI. Filters were manufactured for both small volumes and RBC units. Plasma containing antibodies to human lymphocyte antigen (HLA)-A2 or human neutrophil antigen (HNA)-3a was filtered, and immunoglobulins and specific HNA-3a and HLA-2a neutrophil (PMN) priming activity were measured. Antibodies to OX27 were added to plasma, and filtration was evaluated in a 2-event animal model of TRALI. RBC units from 31 donors known to have antibodies against HLA antigens and from 16 antibody-negative controls were filtered. Furthermore, 4 RBC units were drawn and underwent standard leukoreduction. Immunoglobulins, HLA antibodies, PMN priming activity, and the ability to induce TRALI in an animal model were measured. Small-volume filtration of plasma removed >96% of IgG, antibodies to HLA-A2 and HNA-3a, and their respective priming activity, as well as mitigating antibody-mediated in vivo TRALI. In RBC units, experimental filtration removed antibodies to HLA antigens and inhibited the accumulation of lipid priming activity and lipid-mediated TRALI. We conclude that filtration removes proinflammatory activity and the ability to induce TRALI from RBCs and may represent a TRALI mitigation step. PMID:24747436

  17. Nanodefects of membranes cause destruction of packed red blood cells during long-term storage

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kozlova, Elena, E-mail: waterlake@mail.ru; I.M. Sechenov First Moscow State Medical University, Moscow; Chernysh, Aleksandr

    Packed red blood cells (PRBC) are used for blood transfusion. PRBC were stored for 30 days under 4 °C in hermetic blood bags with CPD anticoagulant-preservative solution. Hematocrit was 50–55%. The distortions of PRBC membranes nanostructure and cells morphology during storage were studied by atomic force microscopy. Basic measurements were performed at the day 2, 6, 9, 16, 23 and 30 of storage and additionally 2–3 days after it. Topological defects occurred on RBC membranes by day 9. They appeared as domains with grain-like structures (“grains”) sized up to 200 nm. These domains were appeared in almost all cells. Latermore » these domains merged and formed large defects on cells. It was the formation of domains with the “grains” which was onset process leading eventually to destruction of PRBC. Possible mechanisms of transformation of PRBC and their membrane are related to the alterations of spectrin cytoskeleton. During this storage period potassium ions and lactat concentrations increased, pH decreased, intracellular concentration of reduced glutathione diminished in the preservative solution. Changes of PRBC morphology were detected within the entire period of PRBC storage. Discocytes predominated at the days 1 and 2. By day 30 PRBC transformed into irreversible echinocytes and spheroechinocytes. Study of defects of membranes nanostructure may form the basis of assessing the quality of the stored PRBC. This method may allow to work out the best recommendations for blood transfusion. - Highlights: • Domains with “grains” are formed on membranes surface on 9–16 days of PRBC storage. • The development of domains is the reason of irreversible changes of PRBC structure. • The origin of domains is the consequence of alterations of spectrin cytoskeleton. • Study of nanostructure may form basis of assessing the quality of the stored PRBC.« less

  18. Variability of cholesterol accessibility in human red blood cells measured using a bacterial cholesterol-binding toxin

    PubMed Central

    Chakrabarti, Rima S; Ingham, Sally A; Kozlitina, Julia; Gay, Austin; Cohen, Jonathan C; Radhakrishnan, Arun; Hobbs, Helen H

    2017-01-01

    Cholesterol partitions into accessible and sequestered pools in cell membranes. Here, we describe a new assay using fluorescently-tagged anthrolysin O, a cholesterol-binding bacterial toxin, to measure accessible cholesterol in human red blood cells (RBCs). Accessible cholesterol levels were stable within individuals, but varied >10 fold among individuals. Significant variation was observed among ethnic groups (Blacks>Hispanics>Whites). Variation in accessibility of RBC cholesterol was unrelated to the cholesterol content of RBCs or plasma, but was associated with the phospholipid composition of the RBC membranes and with plasma triglyceride levels. Pronase treatment of RBCs only modestly altered cholesterol accessibility. Individuals on hemodialysis, who have an unexplained increase in atherosclerotic risk, had significantly higher RBC cholesterol accessibility. Our data indicate that RBC accessible cholesterol is a stable phenotype with significant inter-individual variability. Factors both intrinsic and extrinsic to the RBC contribute to variation in its accessibility. This assay provides a new tool to assess cholesterol homeostasis among tissues in humans. DOI: http://dx.doi.org/10.7554/eLife.23355.001 PMID:28169829

  19. Erythrocyte membrane-coated nanogel for combinatorial antivirulence and responsive antimicrobial delivery against Staphylococcus aureus infection.

    PubMed

    Zhang, Yue; Zhang, Jianhua; Chen, Wansong; Angsantikul, Pavimol; Spiekermann, Kevin A; Fang, Ronnie H; Gao, Weiwei; Zhang, Liangfang

    2017-10-10

    We reported an erythrocyte membrane-coated nanogel (RBC-nanogel) system with combinatorial antivirulence and responsive antibiotic delivery for the treatment of methicillin-resistant Staphylococcus aureus (MRSA) infection. RBC membrane was coated onto the nanogel via a membrane vesicle templated in situ gelation process, whereas the redox-responsiveness was achieved by using a disulfide bond-based crosslinker. We demonstrated that the RBC-nanogels effectively neutralized MRSA-associated toxins in extracellular environment and the toxin neutralization in turn promoted bacterial uptake by macrophages. In intracellular reducing environment, the RBC-nanogels showed an accelerated drug release profile, which resulted in more effective bacterial inhibition. When added to the macrophages infected with intracellular MRSA bacteria, the RBC-nanogels significantly inhibited bacterial growth compared to free antibiotics and non-responsive nanogel counterparts. These results indicate the great potential of the RBC-nanogel system as a new and effective antimicrobial agent against MRSA infection. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Brownian dynamics simulations of lipid bilayer membrane with hydrodynamic interactions in LAMMPS

    NASA Astrophysics Data System (ADS)

    Fu, Szu-Pei; Young, Yuan-Nan; Peng, Zhangli; Yuan, Hongyan

    2016-11-01

    Lipid bilayer membranes have been extensively studied by coarse-grained molecular dynamics simulations. Numerical efficiencies have been reported in the cases of aggressive coarse-graining, where several lipids are coarse-grained into a particle of size 4 6 nm so that there is only one particle in the thickness direction. Yuan et al. proposed a pair-potential between these one-particle-thick coarse-grained lipid particles to capture the mechanical properties of a lipid bilayer membrane (such as gel-fluid-gas phase transitions of lipids, diffusion, and bending rigidity). In this work we implement such interaction potential in LAMMPS to simulate large-scale lipid systems such as vesicles and red blood cells (RBCs). We also consider the effect of cytoskeleton on the lipid membrane dynamics as a model for red blood cell (RBC) dynamics, and incorporate coarse-grained water molecules to account for hydrodynamic interactions. The interaction between the coarse-grained water molecules (explicit solvent molecules) is modeled as a Lennard-Jones (L-J) potential. We focus on two sets of LAMMPS simulations: 1. Vesicle shape transitions with varying enclosed volume; 2. RBC shape transitions with different enclosed volume. This work is funded by NSF under Grant DMS-1222550.

  1. Brownian dynamics simulations of lipid bilayer membrane with hydrodynamic interactions in LAMMPS

    NASA Astrophysics Data System (ADS)

    Fu, Szu-Pei; Young, Yuan-Nan; Peng, Zhangli; Yuan, Hongyan

    Lipid bilayer membranes have been extensively studied by coarse-grained molecular dynamics simulations. Numerical efficiency has been reported in the cases of aggressive coarse-graining, where several lipids are coarse-grained into a particle of size 4 6 nm so that there is only one particle in the thickness direction. Yuan et al. proposed a pair-potential between these one-particle-thick coarse-grained lipid particles to capture the mechanical properties of a lipid bilayer membrane (such as gel-fluid-gas phase transitions of lipids, diffusion, and bending rigidity). In this work we implement such interaction potential in LAMMPS to simulate large-scale lipid systems such as vesicles and red blood cells (RBCs). We also consider the effect of cytoskeleton on the lipid membrane dynamics as a model for red blood cell (RBC) dynamics, and incorporate coarse-grained water molecules to account for hydrodynamic interactions. The interaction between the coarse-grained water molecules (explicit solvent molecules) is modeled as a Lennard-Jones (L-J) potential. We focus on two sets of LAMMPS simulations: 1. Vesicle shape transitions with varying enclosed volume; 2. RBC shape transitions with different enclosed volume.

  2. Biomechanics and dynamics of red blood cells probed by optical tweezers and digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Cardenas, Nelson; Thomas, Pattrick; Yu, Lingfeng; Mohanty, Samarendra

    2011-03-01

    Red blood cells (RBC), with their unique viscoelastic properties, can undergo large deformations during interaction with fluid flow and migration through narrow capillaries. Both local and overall viscoelastic property is important for cellular function and change in these properties indicate diseased condition. Though biomechanics of the cells have been studied using variety of physical techniques (AFM, optically-trapped anchoring beads and microcapilary aspiration) in force regime 10pN, little is studied at low force regime <1pN. Such perturbations are not only hard to exercise on the cell membrane, but quantification of such deformations becomes extremely difficult. By application of low power optical tweezers directly on cell membrane, we could locally perturb discotic RBC along the axial direction, which was monitored dynamically by digital holographic microscopy-a real time, wide-field imaging method having nm axial resolution. The viscoelastic property of the RBC at low force regime was found to be significantly different from that of high-force regime. The results were found to be in good agreement with the simulation results obtained using finite element model of the axially-stretched RBC. The simulations and results of viscoelestic measurements will be presented.

  3. Identification of Rubisco rbcL and rbcS in Camellia oleifera and their potential as molecular markers for selection of high tea oil cultivars

    PubMed Central

    Chen, Yongzhong; Wang, Baoming; Chen, Jianjun; Wang, Xiangnan; Wang, Rui; Peng, Shaofeng; Chen, Longsheng; Ma, Li; Luo, Jian

    2015-01-01

    Tea oil derived from seeds of Camellia oleifera Abel. is high-quality edible oil in China. This study isolated full-length cDNAs of Rubisco subunits rbcL and rbcS from C. oleifera. The rbcL has 1,522 bp with a 1,425 bp coding region, encoding 475 amino acids; and the rbcS has 615 bp containing a 528 bp coding region, encoding 176 amino acids. The expression level of the two genes, designated as Co-rbcL and Co-rbcS, was determined in three C. oleifera cultivars: Hengchong 89, Xianglin 1, and Xianglin 14 whose annual oil yields were 546.9, 591.4, and 657.7 kg ha-1, respectively. The Co-rbcL expression in ‘Xianglin 14’ was significantly higher than ‘Xianglin 1’, and ‘Xianglin 1’ was greater than ‘Hengchong 89’. The expression levels of Co-rbcS in ‘Xianglin 1’ and ‘Xianglin 14’ were similar but were significantly greater than in ‘Hengchong 89’. The net photosynthetic rate of ‘Xianglin 14’ was significantly higher than ‘Xianglin 1’, and ‘Xianglin 1’ was higher than ‘Hengchong 89’. Pearson’s correlation analysis showed that seed yields and oil yields were highly correlated with the expression level of Co-rbcL at P < 0.001 level; and the expression of Co-rbcS was correlated with oil yield at P < 0.01 level. Net photosynthetic rate was also correlated with oil yields and seed yields at P < 0.001 and P < 0.01 levels, respectively. Our results suggest that Co-rbcS and Co-rbcL in particular could potentially be molecular markers for early selection of high oil yield cultivars. In combination with the measurement of net photosynthetic rates, the early identification of potential high oil production cultivars would significantly shorten plant breeding time and increase breeding efficiency. PMID:25873921

  4. Molecular cloning and expression of a chloride channel-associated protein pICln in human young red blood cells: association with actin.

    PubMed

    Schwartz, R S; Rybicki, A C; Nagel, R L

    1997-10-15

    We report the cloning and sequencing from human reticulocytes of cDNA coding for the Cl- channel-associated protein, pICln. Human reticulocyte pICln (HRpICln) cDNA encodes a protein (predicted molecular mass 26293Da) identical with human non-pigmented ciliary epithelial cell pICln. By using full-length HRpICln cDNA (approx. 1.2 kb) to probe human lymphocyte metaphase-chromosome spreads, the location of the human ICln gene was mapped to 11q13 by fluorescence in situ hybridization analysis. Polyclonal antibodies to recombinant HRpICln detected bands at approx. 43 kDa and approx. 37 kDa in both normal (AA) and sickle (SS) red blood cell (RBC) ghost membranes. In SS ghosts, and in ghosts from a patient with autoimmune haemolytic anaemia with 9.8% reticulocytes, the amount of HRpICln was increased compared with AA ghosts, suggesting that the expression or membrane assembly of HRpICln is cell age-dependent. Laser scanning confocal fluorescent microscopy immunolocalized HRpICln largely to the RBC membrane. The increased staining intensity of HRpICln in a reticulocyte-enriched AA RBC density-separated fraction is consistent with a dependence of HRpICln membrane content on cell age. HRpICln and beta-actin form stable complexes in vivo, demonstrated with the yeast two-hybrid system. Low-ionic-strength extraction of ghost membranes, which results in the extraction of the spectrin-actin cytoskeleton, also results in the extraction of HRpICln, consistent with the possibility for the association of these proteins in RBCs in vivo. The results presented here establish the presence of the Cl- channel-associated protein, pICln, in human RBCs, and raises the possibility that this protein has a role in RBC Cl- transport and volume regulation in young RBCs. Moreover the association of RBC pICln with actin offers a model in which to test interactions between RBC ion channels and the cytoskeleton.

  5. Static and dynamic light scattering by red blood cells: A numerical study.

    PubMed

    Mauer, Johannes; Peltomäki, Matti; Poblete, Simón; Gompper, Gerhard; Fedosov, Dmitry A

    2017-01-01

    Light scattering is a well-established experimental technique, which gains more and more popularity in the biological field because it offers the means for non-invasive imaging and detection. However, the interpretation of light-scattering signals remains challenging due to the complexity of most biological systems. Here, we investigate static and dynamic scattering properties of red blood cells (RBCs) using two mesoscopic hydrodynamics simulation methods-multi-particle collision dynamics and dissipative particle dynamics. Light scattering is studied for various membrane shear elasticities, bending rigidities, and RBC shapes (e.g., biconcave and stomatocyte). Simulation results from the two simulation methods show good agreement, and demonstrate that the static light scattering of a diffusing RBC is not very sensitive to the changes in membrane properties and moderate alterations in cell shapes. We also compute dynamic light scattering of a diffusing RBC, from which dynamic properties of RBCs such as diffusion coefficients can be accessed. In contrast to static light scattering, the dynamic measurements can be employed to differentiate between the biconcave and stomatocytic RBC shapes and generally allow the differentiation based on the membrane properties. Our simulation results can be used for better understanding of light scattering by RBCs and the development of new non-invasive methods for blood-flow monitoring.

  6. Static and dynamic light scattering by red blood cells: A numerical study

    PubMed Central

    Mauer, Johannes; Peltomäki, Matti; Poblete, Simón; Gompper, Gerhard

    2017-01-01

    Light scattering is a well-established experimental technique, which gains more and more popularity in the biological field because it offers the means for non-invasive imaging and detection. However, the interpretation of light-scattering signals remains challenging due to the complexity of most biological systems. Here, we investigate static and dynamic scattering properties of red blood cells (RBCs) using two mesoscopic hydrodynamics simulation methods—multi-particle collision dynamics and dissipative particle dynamics. Light scattering is studied for various membrane shear elasticities, bending rigidities, and RBC shapes (e.g., biconcave and stomatocyte). Simulation results from the two simulation methods show good agreement, and demonstrate that the static light scattering of a diffusing RBC is not very sensitive to the changes in membrane properties and moderate alterations in cell shapes. We also compute dynamic light scattering of a diffusing RBC, from which dynamic properties of RBCs such as diffusion coefficients can be accessed. In contrast to static light scattering, the dynamic measurements can be employed to differentiate between the biconcave and stomatocytic RBC shapes and generally allow the differentiation based on the membrane properties. Our simulation results can be used for better understanding of light scattering by RBCs and the development of new non-invasive methods for blood-flow monitoring. PMID:28472125

  7. Predicting dynamics and rheology of blood flow: A comparative study of multiscale and low-dimensional models of red blood cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Pan, Wenxiao; Fedosov, Dmitry A.; Caswell, Bruce

    In this work we compare the predictive capability of two mathematical models for red blood cells (RBCs) focusing on blood flow in capillaries and arterioles. Both RBC models as well as their corresponding blood flows are based on the dissipative particle dynamics (DPD) method, a coarse-grained molecular dynamics approach. The first model employs a multiscale description of the RBC (MS-RBC), with its membrane represented by hundreds or even thousands of DPD-particles connected by springs into a triangular network in combination with out-of-plane elastic bending resistance. Extra dissipation within the network accounts for membrane viscosity, while the characteristic biconcave RBC shapemore » is achieved by imposition of constraints for constant membrane area and constant cell volume. The second model is based on a low-dimensional description (LD-RBC) constructed as a closed torus-like ring of only 10 large DPD colloidal particles. They are connected into a ring by worm-like chain (WLC) springs combined with bending resistance. The LD-RBC model can be fitted to represent the entire range of nonlinear elastic deformations as measured by optical-tweezers for healthy and for infected RBCs in malaria. MS-RBCs suspensions model the dynamics and rheology of blood flow accurately for any size vessel but this approach is computationally expensive above 100 microns. Surprisingly, the much more economical suspensions of LD-RBCs also capture the blood flow dynamics and rheology accurately except for vessels with sizes comparable to RBC diameter. In particular, the LD-RBC suspensions are shown to properly capture the experimental data for the apparent viscosity of blood and its cell-free layer (CFL) in tube flow. Taken together, these findings suggest a hierarchical approach in modeling blood flow in the arterial tree, whereby the MS-RBC model should be employed for capillaries and arterioles below 100 microns, the LD-RBC model for arterioles, and the continuum description for

  8. Estimation of adult and neonatal RBC lifespans in anemic neonates using RBCs labeled at several discrete biotin densities.

    PubMed

    Kuruvilla, Denison J; Widness, John A; Nalbant, Demet; Schmidt, Robert L; Mock, Donald M; An, Guohua; Veng-Pedersen, Peter

    2017-06-01

    Prior conclusions that autologous neonatal red blood cells (RBC) have substantially shorter lifespans than allogeneic adult RBCs were not based on direct comparison of autologous neonatal vs. allogeneic adult RBCs performed concurrently in the same infant. Biotin labeling of autologous neonatal RBCs and allogeneic adult donor RBCs permits concurrent direct comparison of autologous vs. allogeneic RBC lifespan. RBCs from 15 allogeneic adult donors and from 15 very-low-birth-weight (VLBW) neonates were labeled at separate biotin densities and transfused simultaneously into the 15 neonates. Two mathematical models that account for the RBC differences were employed to estimate lifespans for the two RBC populations. Mean ± SD lifespan for adult allogeneic RBC was 70.1 ± 19.1 d, which is substantially shorter than the 120 d lifespan of both autologous and adult allogeneic RBC in healthy adults. Mean ± SD lifespan for neonatal RBC was 54.2 ± 11.3 d, which is only about 30% shorter than that of the adult allogeneic RBCs. This study provides evidence that extrinsic environmental factors primarily determine RBC survival (e.g., small bore of the capillaries of neonates, rate of oxygenation/deoxygenation cycles) rather than factors intrinsic to RBC.

  9. Cell Membrane Electropulsation: Chemical Analysis of Cell Membrane Modifications and Associated Transport Mechanisms.

    PubMed

    Azan, Antoine; Gailliègue, Florian; Mir, Lluis M; Breton, Marie

    2017-01-01

    The transport of substances across the cell membrane is complex because the main physiological role of the membrane is the control of the substances that would enter or exit the cells. Life would not be possible without this control. Cell electropulsation corresponds to the delivery of electric pulses to the cells and comprises cell electroporation and cell electropermeabilization. Cell electropulsation allows for the transport of non-permeant molecules across the membrane, bypassing the physiological limitations. In this chapter we discuss the changes occurring in the cell membrane during electroporation or electropermeabilization as they allow to understand which molecules can be transported as well as when and how their transport can occur. Electrophoretic or diffusive transports across the cell membrane can be distinguished. This understanding has a clear impact on the choice of the electrical parameters to be applied to the cells as well as on other aspects of the experimental protocols that have to be set to load the cells with non-permeant molecules.

  10. The Effects of Ethanol on the Morphological and Biochemical Properties of Individual Human Red Blood Cells.

    PubMed

    Lee, Sang Yun; Park, Hyun Joo; Best-Popescu, Catherine; Jang, Seongsoo; Park, Yong Keun

    2015-01-01

    Here, we report the results of a study on the effects of ethanol exposure on human red blood cells (RBCs) using quantitative phase imaging techniques at the level of individual cells. Three-dimensional refractive index tomograms and dynamic membrane fluctuations of RBCs were measured using common-path diffraction optical tomography, from which morphological (volume, surface area, and sphericity); biochemical (hemoglobin (Hb) concentration and Hb content); and biomechanical (membrane fluctuation) parameters were retrieved at various concentrations of ethanol. RBCs exposed to the ethanol concentration of 0.1 and 0.3% v/v exhibited cell sphericities higher than those of normal cells. However, mean surface area and sphericity of RBCs in a lethal alcoholic condition (0.5% v/v) are not statistically different with those of healthy RBCs. Meanwhile, significant decreases of Hb content and concentration in RBC cytoplasm at the lethal condition were observed. Furthermore, dynamic fluctuation of RBC membranes increased significantly upon ethanol treatments, indicating ethanol-induced membrane fluidization.

  11. Use of RBC-O and S-MCV parameters of SYSMEX XE-2100 in a patient with RBC cold agglutination.

    PubMed

    Wang, Hong; Lu, Lin; Zhou, Yun; Liu, Jian; Qian, Min; Tang, Weiming; Jie, Zhang; Pan, Shiyang

    2013-01-01

    Sometimes EDTA blood of erythrocyte agglutination cannot be well resolved by incubation at 37 degrees C. In this case report, however, such a specimen was detected from a lymphoma patient at room temperature by using RBC-O and S-MCV parameters of the SYSMEX XE-2100 hematology analyzer. The specimen was diluted with 0.9% NaCL solution at 1:1 before measurement. HCT, MCV, and MCHC, corrected by RBC-O, HGB and S-MCV, were all in their normal ranges. This case indicates that RBC-O and S-MCV parameters of XE-2100 can be used in the routine blood examination of erythrocyte agglutination specimen at room temperature.

  12. Cell membrane disruption stimulates cAMP and Ca2+ signaling to potentiate cell membrane resealing in neighboring cells.

    PubMed

    Togo, Tatsuru

    2017-12-15

    Disruption of cellular plasma membranes is a common event in many animal tissues, and the membranes are usually rapidly resealed. Moreover, repeated membrane disruptions within a single cell reseal faster than the initial wound in a protein kinase A (PKA)- and protein kinase C (PKC)-dependent manner. In addition to wounded cells, recent studies have demonstrated that wounding of Madin-Darby canine kidney (MDCK) cells potentiates membrane resealing in neighboring cells in the short-term by purinergic signaling, and in the long-term by nitric oxide/protein kinase G signaling. In the present study, real-time imaging showed that cell membrane disruption stimulated cAMP synthesis and Ca 2+ mobilization from intracellular stores by purinergic signaling in neighboring MDCK cells. Furthermore, inhibition of PKA and PKC suppressed the ATP-mediated short-term potentiation of membrane resealing in neighboring cells. These results suggest that cell membrane disruption stimulates PKA and PKC via purinergic signaling to potentiate cell membrane resealing in neighboring MDCK cells. © 2017. Published by The Company of Biologists Ltd.

  13. Erythrocyte plasma membrane-bound ERK1/2 activation promotes ICAM-4-mediated sickle red cell adhesion to endothelium.

    PubMed

    Zennadi, Rahima; Whalen, Erin J; Soderblom, Erik J; Alexander, Susan C; Thompson, J Will; Dubois, Laura G; Moseley, M Arthur; Telen, Marilyn J

    2012-02-02

    The core pathology of sickle cell disease (SCD) starts with the erythrocyte (RBC). Aberration in MAPK/ERK1/2 signaling, which can regulate cell adhesion, occurs in diverse pathologies. Because RBCs contain abundant ERK1/2, we predicted that ERK1/2 is functional in sickle (SS) RBCs and promotes adherence, a hallmark of SCD. ERK1/2 remained active in SS but not normal RBCs. β(2)-adrenergic receptor stimulation by epinephrine can enhance ERK1/2 activity only in SS RBCs via PKA- and tyrosine kinase p72(syk)-dependent pathways. ERK signaling is implicated in RBC ICAM-4 phosphorylation, promoting SS RBC adhesion to the endothelium. SS RBC adhesion and phosphorylation of both ERK and ICAM-4 all decreased with continued cell exposure to epinephrine, implying that activation of ICAM-4-mediated SS RBC adhesion is temporally associated with ERK1/2 activation. Furthermore, recombinant ERK2 phosphorylated α- and β-adducins and dematin at the ERK consensus motif. Cytoskeletal protein 4.1 also showed dynamic phosphorylation but not at the ERK consensus motif. These results demonstrate that ERK activation induces phosphorylation of cytoskeletal proteins and the adhesion molecule ICAM-4, promoting SS RBC adhesion to the endothelium. Thus, blocking RBC ERK1/2 activation, such as that promoted by catecholamine stress hormones, could ameliorate SCD pathophysiology.

  14. n-3 and n-6 Fatty Acid Changes in the Erythrocyte Membranes of Patients with 658240251 Clostridium difficile Infection.

    PubMed

    Czepiel, Jacek; Gdula-Argasińska, Joanna; Garlicki, Aleksander

    2016-01-01

    The implications of circulating essential fatty acids (FA) on the inflammatory risk profile and clinical outcome are still unclear. In order to gain a deeper understanding of the role of polyunsaturated fatty acids (PUFA) in the pathogenesis of acute infection, we analyzed the FA content in red blood cell (RBC) membranes of patients with Clostridium difficile infection (CDI) and controls. We prospectively studied 60 patients including 30 patients with CDI and 30 controls to assess lipid concentrations in erythrocyte membranes using gas chromatography. We observed a higher level of saturated fatty acids (SFA) in RBC membranes from patients with CDI. In patients with CDI, we also noticed a higher level of 20:4 n-6 FA and only a small amounts of C20:2n-6, C20:3n-6 FAs, arachidonic acid (AA) precursors, which suggest an intense inflammatory reaction in the organism during infection. We also noticed low levels of n-3 FA in the RBC membranes of patients infected with CDI. There is a deficit of n-3 FA in patients with CDI. n-3 FA are probably used during CDI as precursors of pro-resolving mediators that may indicate a therapeutic role of n-3 PUFAs in CDI. The changes in fatty acids in erythrocyte membranes during CDI alter their functions which may have an impact on the clinical outcome.

  15. Influence of Pre-Storage Irradiation on the Oxidative Stress Markers, Membrane Integrity, Size and Shape of the Cold Stored Red Blood Cells.

    PubMed

    Antosik, Adam; Czubak, Kamila; Gajek, Arkadiusz; Marczak, Agnieszka; Glowacki, Rafal; Borowczyk, Kamila; Zbikowska, Halina Malgorzata

    2015-05-01

    To investigate the extent of oxidative damage and changes in morphology of manually isolated red blood cells (RBCs) from whole blood, cold stored (up to 20 days) in polystyrene tubes and subjected to pre-storage irradiation (50 Gy) and to compare the properties of SAGM-preserved RBCs stored under experimental conditions (polystyrene tubes) with RBCs from standard blood bag storage. The percentage of hemolysis as well as the extracellular activity of LDH, thiobarbituric acid-reactive substances, reduced glutathione (GSH), and total antioxidant capacity (TAC) were measured. Changes in the topology of RBC membrane, shape, and size were evaluated by flow cytometry and judged against microscopy images. Irradiation caused significant LDH release as well as increased hemolysis and lipid peroxidation, GSH depletion, and reduction of TAC. Prolonged storage of irradiated RBCs resulted in phosphatidylserine exposure on the cell surface. By day 20, approximately 60% of RBCs displayed non-discoid shape. We did not notice significant differences in percentage of altered cells and cell volume between RBCs exposed to irradiation and those not exposed. Irradiation of RBC transfusion units with a dose of 50 Gy should be avoided. For research purposes such as studying the role of antioxidants, storage of small volumes of RBCs derived from the same donor would be more useful, cheaper, and blood-saving.

  16. Stretching of red blood cells by optical tweezers quantified by digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Cardenas, Nelson; Yu, Lingfeng; Mohanty, Samarendra K.

    2011-03-01

    Red blood cells (RBC) possess unique viscoelastic characteristics which allow them to pass through capillaries narrower than their size. Measurement of viscoelastic property of cells (e.g. RBC) in low-force regime is of high significance as it represents conditions of membrane fluctuation in response to physiological conditions. Estimation of visco-elastic properties of RBC requires measurement of extent of deformation in RBC subjected to known force. Optical tweezers, being gentle and absolutely sterile, are emerging as the tool of choice for application of localized force on cells. However, stretching of RBC in very low force regime has not been quantified. Further, though deformations in transverse directions have been measured, vertical deformations due to stretching of cells cannot be quantified by classical microscopic images. Here, we report realization of offaxis digital holographic microscopy (DHM) for highly sensitive axial changes in RBC shape due to stretching by optical tweezers without attaching microscopic beads. The RBC was stretched in axial direction with nanometer precision by change of divergence of the trapping beam. The obtained deformation patterns were compared with the axial position of the tweezers focus. Since the pathophysiology of progression of diseases like malaria and cancer is reflected in the biophysical (both mechanical and material) properties of the cells, it is possible to identify the changes by simultaneous measurement of refractive index and elasticity using this approach.

  17. Rapid rather than gradual weight reduction impairs hemorheological parameters of Taekwondo athletes through reduction in RBC-NOS activation.

    PubMed

    Yang, Woo Hwi; Heine, Oliver; Pauly, Sebastian; Kim, Pilsang; Bloch, Wilhelm; Mester, Joachim; Grau, Marijke

    2015-01-01

    Rapid weight reduction is part of the pre-competition routine and has been shown to negatively affect psychological and physiological performance of Taekwondo (TKD) athletes. This is caused by a reduction of the body water and an electrolyte imbalance. So far, it is unknown whether weight reduction also affects hemorheological properties and hemorheology-influencing nitric oxide (NO) signaling, important for oxygen supply to the muscles and organs. For this purpose, ten male TKD athletes reduced their body weight by 5% within four days (rapid weight reduction, RWR). After a recovery phase, athletes reduced body weight by 5% within four weeks (gradual weight reduction, GWR). Each intervention was preceded by two baseline measurements and followed by a simulated competition. Basal blood parameters (red blood cell (RBC) count, hemoglobin concentration, hematocrit, mean corpuscular volume, mean cellular hemoglobin and mean cellular hemoglobin concentration), RBC-NO synthase activation, RBC nitrite as marker for NO synthesis, RBC deformability and aggregation parameters were determined on a total of eight investigation days. Basal blood parameters were not affected by the two interventions. In contrast to GWR, RWR decreased activation of RBC-NO synthase, RBC nitrite, respective NO concentration and RBC deformability. Additionally, RWR increased RBC aggregation and disaggregation threshold. The results point out that a rapid weight reduction negatively affects hemorheological parameters and NO signaling in RBC which might limit performance capacity. Thus, GWR should be preferred to achieve the desired weight prior to a competition to avoid these negative effects.

  18. Kinetic parameters of rubidium transport pathways are normal in cystic fibrosis red cells.

    PubMed

    Joiner, C H

    1988-10-01

    The abnormalities in ion transport in cystic fibrosis (CF) respiratory and sweat duct epithelia have prompted studies of ion permeability in CF red blood cells (RBC) although previous reports have been contradictory. In this study, the kinetic characteristics of the three major cation transport systems in RBC were evaluated by measuring rubidium (Rb) uptake at various external Rb concentrations. The maximal velocity and affinity for external Rb (K1/2) of the NaK pump were normal in CF RBC, as were the maximal velocity and Km for Rb of the NaK cotransport system. Residual (ouabain and bumetanide insensitive) Rb uptake, and steady state RBC Na and K contents were also normal. These data indicate the NaK pump and cotransport system do not exhibit primary or secondary perturbations in CF RBC, and suggest that the noncarrier-mediated membrane permeability to cations is also normal in these cells.

  19. Biological Fuel Cells and Membranes.

    PubMed

    Ghassemi, Zahra; Slaughter, Gymama

    2017-01-17

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

  20. Biological Fuel Cells and Membranes

    PubMed Central

    Ghassemi, Zahra; Slaughter, Gymama

    2017-01-01

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

  1. A simple biofuel cell cathode with human red blood cells as electrocatalysts for oxygen reduction reaction.

    PubMed

    Ayato, Yusuke; Sakurai, Kenichiro; Fukunaga, Saori; Suganuma, Takuya; Yamagiwa, Kiyofumi; Shiroishi, Hidenobu; Kuwano, Jun

    2014-05-15

    A red blood cell (RBC) from human exhibited direct electron transfer (DET) activity on a bare indium tin oxide (ITO) electrode. A formal potential of -0.152 V vs. a silver-silver chloride saturated potassium chloride (Ag|AgCl|KCl(satd.)) was estimated for the human RBC (type AB) from a pair of redox peaks at around 0.089 and -0.215 V (vs. Ag|AgCl|KCl(satd.)) on cyclic voltammetric (CV) measurements in a phosphate buffered saline (PBS; 39 mM; pH 7.4) solution. The results agreed well with those of a redox couple for iron-bearing heme groups in hemoglobin molecules (HbFe(II)/HbFe(III)) on the bare ITO electrodes, indicated that DET active species were hemoglobin (Hb) molecules encapsulated by a phospholipid bilayer membrane of the human RBC. The quantity of electrochemically active Hb in the human RBC was estimated to be 30 pmol cm(-2). In addition, the human RBC exhibited oxygen reduction reaction (ORR) activity in the dioxygen (O2) saturated PBS solution at the negative potential from ca. -0.15 V (vs. Ag|AgCl|KCl(satd.)). A single cell test proved that a biofuel cell (BFC) with an O2|RBC|ITO cathode showed the open-circuit voltage (OCV) of ca. 0.43 V and the maximum power density of ca. 0.68 μW cm(-2). © 2013 Published by Elsevier B.V.

  2. Cell Membrane Transport Mechanisms: Ion Channels and Electrical Properties of Cell Membranes.

    PubMed

    Kulbacka, Julita; Choromańska, Anna; Rossowska, Joanna; Weżgowiec, Joanna; Saczko, Jolanta; Rols, Marie-Pierre

    2017-01-01

    Cellular life strongly depends on the membrane ability to precisely control exchange of solutes between the internal and external (environmental) compartments. This barrier regulates which types of solutes can enter and leave the cell. Transmembrane transport involves complex mechanisms responsible for passive and active carriage of ions and small- and medium-size molecules. Transport mechanisms existing in the biological membranes highly determine proper cellular functions and contribute to drug transport. The present chapter deals with features and electrical properties of the cell membrane and addresses the questions how the cell membrane accomplishes transport functions and how transmembrane transport can be affected. Since dysfunctions of plasma membrane transporters very often are the cause of human diseases, we also report how specific transport mechanisms can be modulated or inhibited in order to enhance the therapeutic effect.

  3. The Molecules of the Cell Membrane.

    ERIC Educational Resources Information Center

    Bretscher, Mark S.

    1985-01-01

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

  4. Transfusion of cell saver salvaged blood in neonates and infants undergoing open heart surgery significantly reduces RBC and coagulant product transfusions and donor exposures: results of a prospective, randomized, clinical trial.

    PubMed

    Cholette, Jill M; Powers, Karen S; Alfieris, George M; Angona, Ronald; Henrichs, Kelly F; Masel, Debra; Swartz, Michael F; Daugherty, L Eugene; Belmont, Kevin; Blumberg, Neil

    2013-02-01

    To evaluate whether transfusion of cell saver salvaged, stored at the bedside for up to 24 hrs, would decrease the number of postoperative allogeneic RBC transfusions and donor exposures, and possibly improve clinical outcomes. Prospective, randomized, controlled, clinical trial. Pediatric cardiac intensive care unit. Infants weighing less than 20 kg (n = 106) presenting for cardiac surgery with cardiopulmonary bypass. Subjects were randomized to a cell saver transfusion group where cell saver blood was available for transfusion up to 24 hrs after collection, or to a control group. Cell saver subjects received cell saver blood for volume replacement and/or RBC transfusions. Control subjects received crystalloid or albumin for volume replacement and RBCs for anemia. Blood product transfusions, donor exposures, and clinical outcomes were compared between groups. Children randomized to the cell saver group had significantly fewer RBC transfusions (cell saver: 0.19 ± 0.44 vs. control: 0.75 ± 1.2; p = 0.003) and coagulant product transfusions in the first 48 hrs post-op (cell saver: 0.09 ± 0.45 vs. control: 0.62 ± 1.4; p = 0.013), and significantly fewer donor exposures (cell saver: 0.60 ± 1.4 vs. control: 2.3 ± 4.8; p = 0.019). This difference persisted over the first week post-op, but did not reach statistical significance (cell saver: 0.64 ± 1.24 vs. control: 1.1 ± 1.4; p = 0.07). There were no significant clinical outcome differences. Cell saver blood can be safely stored at the bedside for immediate transfusion for 24 hrs after collection. Administration of cell saver blood significantly reduces the number of RBC and coagulant product transfusions and donor exposures in the immediate postoperative period. Reduction of blood product transfusions has the potential to reduce transfusion-associated complications and decrease postoperative morbidity. Larger studies are needed to determine whether this transfusion strategy will improve clinical outcomes.

  5. Composite fuel cell membranes

    DOEpatents

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

    1997-08-05

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

  6. Composite fuel cell membranes

    DOEpatents

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

    1997-01-01

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

  7. Studying red blood cell agglutination by measuring electrical and mechanical properties with a double optical tweezers

    NASA Astrophysics Data System (ADS)

    Fontes, Adriana; Fernandes, Heloise P.; de Thomaz, André A.; Barbosa, Luiz C.; Barjas-Castro, Maria L.; Cesar, Carlos L.

    2007-07-01

    The red blood cell (RBC) viscoelastic membrane contains proteins and glycolproteins embedded in, or attached, to a fluid lipid bilayer and are negatively charged, which creates a repulsive electric (zeta) potential between the cells and prevents their aggregation in the blood stream. The basis of the immunohematologic tests is the interaction between antigens and antibodies that causes hemagglutination. The identification of antibodies and antigens is of fundamental importance for the transfusional routine. This agglutination is induced by decreasing the zeta-potential through the introduction of artificial potential substances. This report proposes the use of the optical tweezers to measure the membrane viscosity, the cell adhesion, the zeta-potential and the size of the double layer of charges (CLC) formed around the cell in an electrolytic solution. The adhesion was quantified by slowly displacing two RBCs apart until the disagglutination. The CLC was measured using the force on the bead attached to a single RBC in response to an applied voltage. The zeta-potential was obtained by measuring the terminal velocity after releasing the RBC from the optical trap at the last applied voltage. For the membrane viscosity experiment, we trapped a bead attached to RBCs and measured the force to slide one RBC over the other as a function of the relative velocity. After we tested the methodology, we performed measurements using antibody and potential substances. We observed that this experiment can provide information about cell agglutination that helps to improve the tests usually performed in blood banks. We also believe that this methodology can be applied for measurements of zeta-potentials in other kind of samples.

  8. OpenRBC: Redefining the Frontier of Red Blood Cell Simulations at Protein Resolution

    NASA Astrophysics Data System (ADS)

    Tang, Yu-Hang; Lu, Lu; Li, He; Grinberg, Leopold; Sachdeva, Vipin; Evangelinos, Constantinos; Karniadakis, George

    We present a from-scratch development of OpenRBC, a coarse-grained molecular dynamics code, which is capable of performing an unprecedented in silico experiment - simulating an entire mammal red blood cell lipid bilayer and cytoskeleton modeled by 4 million mesoscopic particles - on a single shared memory node. To achieve this, we invented an adaptive spatial searching algorithm to accelerate the computation of short-range pairwise interactions in an extremely sparse 3D space. The algorithm is based on a Voronoi partitioning of the point cloud of coarse-grained particles, and is continuously updated over the course of the simulation. The algorithm enables the construction of a lattice-free cell list, i.e. the key spatial searching data structure in our code, in O (N) time and space space with cells whose position and shape adapts automatically to the local density and curvature. The code implements NUMA/NUCA-aware OpenMP parallelization and achieves perfect scaling with up to hundreds of hardware threads. The code outperforms a legacy solver by more than 8 times in time-to-solution and more than 20 times in problem size, thus providing a new venue for probing the cytomechanics of red blood cells. This work was supported by the Department of Energy (DOE) Collaboratory on Mathematics for Mesoscopic Model- ing of Materials (CM4). YHT acknowledges partial financial support from an IBM Ph.D. Scholarship Award.

  9. Membrane Cells for Brine Electrolysis.

    ERIC Educational Resources Information Center

    Tingle, M.

    1982-01-01

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

  10. Influence of Pre-Storage Irradiation on the Oxidative Stress Markers, Membrane Integrity, Size and Shape of the Cold Stored Red Blood Cells

    PubMed Central

    Antosik, Adam; Czubak, Kamila; Gajek, Arkadiusz; Marczak, Agnieszka; Glowacki, Rafal; Borowczyk, Kamila; Zbikowska, Halina Malgorzata

    2015-01-01

    Background To investigate the extent of oxidative damage and changes in morphology of manually isolated red blood cells (RBCs) from whole blood, cold stored (up to 20 days) in polystyrene tubes and subjected to pre-storage irradiation (50 Gy) and to compare the properties of SAGM-preserved RBCs stored under experimental conditions (polystyrene tubes) with RBCs from standard blood bag storage. Methods The percentage of hemolysis as well as the extracellular activity of LDH, thiobarbituric acid-reactive substances, reduced glutathione (GSH), and total antioxidant capacity (TAC) were measured. Changes in the topology of RBC membrane, shape, and size were evaluated by flow cytometry and judged against microscopy images. Results Irradiation caused significant LDH release as well as increased hemolysis and lipid peroxidation, GSH depletion, and reduction of TAC. Prolonged storage of irradiated RBCs resulted in phosphatidylserine exposure on the cell surface. By day 20, approximately 60% of RBCs displayed non-discoid shape. We did not notice significant differences in percentage of altered cells and cell volume between RBCs exposed to irradiation and those not exposed. Conclusion Irradiation of RBC transfusion units with a dose of 50 Gy should be avoided. For research purposes such as studying the role of antioxidants, storage of small volumes of RBCs derived from the same donor would be more useful, cheaper, and blood-saving. PMID:26195927

  11. The protective effect of aqueous extracts of roselle (Hibiscus sabdariffa L. UKMR-2) against red blood cell membrane oxidative stress in rats with streptozotocin-induced diabetes.

    PubMed

    Mohamed, Jamaludin; Shing, Saw Wuan; Idris, Muhd Hanis Md; Budin, Siti Balkis; Zainalabidin, Satirah

    2013-10-01

    The aim of this study was to investigate the protective effects of aqueous extracts of roselle (Hibiscus sabdariffa L. UKMR-2) against red blood cell (RBC) membrane oxidative stress in rats with streptozotocin-induced diabetes. Forty male Sprague-Dawley rats weighing 230-250 g were randomly divided into four groups (n = 10 rats each): control group (N), roselle-treated control group, diabetic group, and roselle-treated diabetic group. Roselle was administered by force-feeding with aqueous extracts of roselle (100 mg/kg body weight) for 28 days. The results demonstrated that the malondialdehyde levels of the red blood cell membranes in the diabetic group were significantly higher than the levels in the roselle-treated control and roselle-treated diabetic groups. The protein carbonyl level was significantly higher in the roselle-treated diabetic group than in the roselle-treated control group but lower than that in the diabetic group. A significant increase in the red blood cell membrane superoxide dismutase enzyme was found in roselle-treated diabetic rats compared with roselle-treated control rats and diabetic rats. The total protein level of the red blood cell membrane, osmotic fragility, and red blood cell morphology were maintained. The present study demonstrates that aqueous extracts of roselle possess a protective effect against red blood cell membrane oxidative stress in rats with streptozotocin-induced diabetes. These data suggest that roselle can be used as a natural antioxidative supplement in the prevention of oxidative damage in diabetic patients.

  12. Mechanical properties of stored red blood cells using optical tweezers

    NASA Astrophysics Data System (ADS)

    Fontes, Adriana; Alexandre de Thomaz, Andre; de Ysasa Pozzo, Liliana; de Lourdes Barjas-Castro, Maria; Brandao, Marcelo M.; Saad, Sara T. O.; Barbosa, Luiz Carlos; Cesar, Carlos Lenz

    2005-08-01

    We have developed a method for measuring the red blood cell (RBC) membrane overall elasticity μ by measuring the deformation of the cells when dragged at a constant velocity through a plasma fluid by an optical tweezers. The deformability of erythrocytes is a critical determinant of blood flow in the microcirculation. We tested our method and hydrodynamic models, which included the presence of two walls, by measuring the RBC deformation as a function of drag velocity and of the distance to the walls. The capability and sensitivity of this method can be evaluated by its application to a variety of studies, such as, the measurement of RBC elasticity of sickle cell anemia patients comparing homozygous (HbSS), including patients taking hydroxyrea (HU) and heterozygous (HbAS) with normal donors and the RBC elasticity measurement of gamma irradiated stored blood for transfusion to immunosupressed patients as a function of time and dose. These studies show that the technique has the sensitivity to discriminate heterozygous and homozygous sickle cell anemia patients from normal donors and even follow the course of HU treatment of Homozygous patients. The gamma irradiation studies show that there is no significant change in RBC elasticity over time for up to 14 days of storage, regardless of whether the unit was irradiated or not, but there was a huge change in the measured elasticity for the RBC units stored for more than 21 days after irradiation. These finds are important for the assessment of stored irradiated RBC viability for transfusion purposes because the present protocol consider 28 storage days after irradiation as the limit for the RBC usage.

  13. The endomembrane requirement for cell surface repair

    NASA Technical Reports Server (NTRS)

    McNeil, Paul L.; Miyake, Katsuya; Vogel, Steven S.

    2003-01-01

    The capacity to reseal a plasma membrane disruption rapidly is required for cell survival in many physiological environments. Intracellular membrane (endomembrane) is thought to play a central role in the rapid resealing response. We here directly compare the resealing response of a cell that lacks endomembrane, the red blood cell, with that of several nucleated cells possessing an abundant endomembrane compartment. RBC membrane disruptions inflicted by a mode-locked Ti:sapphire laser, even those initially smaller than hemoglobin, failed to reseal rapidly. By contrast, much larger laser-induced disruptions made in sea urchin eggs, fibroblasts, and neurons exhibited rapid, Ca(2+)-dependent resealing. We conclude that rapid resealing is not mediated by simple physiochemical mechanisms; endomembrane is required.

  14. Simulations of molecular diffusion in lattices of cells: insights for NMR of red blood cells.

    PubMed

    Regan, David G; Kuchel, Philip W

    2002-07-01

    The pulsed field-gradient spin-echo (PGSE) nuclear magnetic resonance (NMR) experiment, conducted on a suspension of red blood cells (RBC) in a strong magnetic field yields a q-space plot consisting of a series of maxima and minima. This is mathematically analogous to a classical optical diffraction pattern. The method provides a noninvasive and novel means of characterizing cell suspensions that is sensitive to changes in cell shape and packing density. The positions of the features in a q-space plot characterize the rate of exchange across the membrane, cell dimensions, and packing density. A diffusion tensor, containing information regarding the diffusion anisotropy of the system, can also be derived from the PGSE NMR data. In this study, we carried out Monte Carlo simulations of diffusion in suspensions of "virtual" cells that had either biconcave disc (as in RBC) or oblate spheroid geometry. The simulations were performed in a PGSE NMR context thus enabling predictions of q-space and diffusion tensor data. The simulated data were compared with those from real PGSE NMR diffusion experiments on RBC suspensions that had a range of hematocrit values. Methods that facilitate the processing of q-space data were also developed.

  15. Role of enzyme-treated cells in RBC antibody screening using the gel test: a study of anti-RH1, -RH2, and -RH3 antibodies.

    PubMed

    Conne, Jocelyne; Schneider, Philippe; Tissot, Jean-Daniel

    2007-01-01

    The role of enzyme-treated cells (ETCs) in red blood cell (RBC) antibody screening has been the subject of controversy, and its place in the clinical routine remains to be determined. In this work, plasma samples containing anti-RH1 (anti-D; N = 10), anti-RH2 (anti-C; N = 10), or anti-RH3 (anti-E; N = 10) antibodies were studied. The samples were diluted in nonbuffered or buffered normal saline, as well as in a pool of AB plasma samples. Titers and scores were determined by means of the gel test, using the indirect antiglobulin test (IAT) as well as ETCs, with R(0)r, r'r, or r''r test cells. Our results showed that compared to the IAT, ETCs allowed a clearer detection of anti-RH2 and anti-RH3, but not of anti-RH1 antibodies. Based on our study, it is not clear whether the ETC phase of the gel test should be maintained for RBC antibody screening. 2007 Wiley-Liss, Inc.

  16. Effects of o-vanillin on K+ transport of red blood cells from patients with sickle cell disease

    PubMed Central

    Hannemann, A.; Cytlak, U.M.C.; Gbotosho, O.T.; Rees, D.C.; Tewari, S.; Gibson, J.S.

    2014-01-01

    Aromatic aldehydes like o-vanillin were designed to reduce the complications of sickle cell disease (SCD) by interaction with HbS, to reduce polymerisation and RBC sickling. Present results show that o-vanillin also directly affects RBC membrane permeability. Both the K+–Cl− cotransporter (KCC) and the Ca2 +-activated K+ channel (or Gardos channel) were inhibited with IC50 of about 0.3 and 1 mM, respectively, with activities almost completely abolished by 5 mM. Similar effects were observed in RBCs treated with the thiol reacting reagent N-ethylmaleimide or with the Ca2 + ionophore A23187, to circumvent any action via HbS polymerisation. The deoxygenation-induced cation conductance (sometimes termed Psickle) was partially inhibited, whilst deoxygenation-induced exposure of phosphatidylserine was completely abrogated. Na+/K+ pump activity was also reduced. Notwithstanding, o-vanillin stimulated K+ efflux through an unidentified pathway and resulted in reduction in cell volume (as measured by wet weight − dry weight). These actions are relevant to understanding how aromatic aldehydes may affect RBC membrane permeability per se as well as HbS polymerisation and thereby inform design of compounds most efficacious in ameliorating the complications of SCD. PMID:24594314

  17. The dielectric spectroscopy of human red blood cells: the differentiation of old from fresh cells.

    PubMed

    David, Marcelo; Levy, Evgeniya; Feldman, Yuri; Ben Ishai, Paul; Zelig, Orly; Yedgar, Saul; Barshtein, Gregory

    2017-06-22

    The objective of the study was to gauge the effect of storage lesions on the dielectric response of red blood cells (RBC), in particular those processes linked to deformations of the cellular membrane known as the β-dispersion. The dielectric response of RBC suspensions, exposed to blood-bank cold storage, was studied using time-domain dielectric spectroscopy (TDDS) in the frequency range of 500 kHz up to 1 GHz. The measured dielectric processes are characterized by their dielectric strength (Δε) and relaxation time (τ). Changes in the dielectric properties of the RBC suspensions due to storage-related lesions were evaluated. For a quantitative characterization of RBC lesions, we measured the deformability of fresh and stored RBC as expressed by their elongation ratio (ER), which was achieved under a shear stress of 3.0 Pa. The results show that the storage of RBC induced a statistically significant decrease of dielectric relaxation times. In addition, a sound correlation between the mean values of ER and the relaxation times was observed (Spearman's correlation coefficient ρ  =  0.847). We draw the conclusion that those alterations in the relaxation time are induced by changes in the shape of the RBC that happen during cold-storage. The evolution of the β-dispersion of RBC opens new possibilities in the blood bank inventory management.

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

    NASA Astrophysics Data System (ADS)

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

    2015-08-01

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

  19. In-membrane micro fuel cell

    DOEpatents

    Omosebi, Ayokunle; Besser, Ronald

    2016-09-06

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

  20. Effects of o-vanillin on K⁺ transport of red blood cells from patients with sickle cell disease.

    PubMed

    Hannemann, A; Cytlak, U M C; Gbotosho, O T; Rees, D C; Tewari, S; Gibson, J S

    2014-01-01

    Aromatic aldehydes like o-vanillin were designed to reduce the complications of sickle cell disease (SCD) by interaction with HbS, to reduce polymerisation and RBC sickling. Present results show that o-vanillin also directly affects RBC membrane permeability. Both the K(+)-Cl(-) cotransporter (KCC) and the Ca(2+)-activated K(+) channel (or Gardos channel) were inhibited with IC50 of about 0.3 and 1 mM, respectively, with activities almost completely abolished by 5 mM. Similar effects were observed in RBCs treated with the thiol reacting reagent N-ethylmaleimide or with the Ca(2+) ionophore A23187, to circumvent any action via HbS polymerisation. The deoxygenation-induced cation conductance (sometimes termed P(sickle)) was partially inhibited, whilst deoxygenation-induced exposure of phosphatidylserine was completely abrogated. Na(+)/K(+) pump activity was also reduced. Notwithstanding, o-vanillin stimulated K(+) efflux through an unidentified pathway and resulted in reduction in cell volume (as measured by wet weight-dry weight). These actions are relevant to understanding how aromatic aldehydes may affect RBC membrane permeability per se as well as HbS polymerisation and thereby inform design of compounds most efficacious in ameliorating the complications of SCD. Copyright © 2014. Published by Elsevier Inc.

  1. Single-cell force spectroscopy as a technique to quantify human red blood cell adhesion to subendothelial laminin.

    PubMed

    Maciaszek, Jamie L; Partola, Kostyantyn; Zhang, Jing; Andemariam, Biree; Lykotrafitis, George

    2014-12-18

    Single-cell force spectroscopy (SCFS), an atomic force microscopy (AFM)-based assay, enables quantitative study of cell adhesion while maintaining the native state of surface receptors in physiological conditions. Human healthy and pathological red blood cells (RBCs) express a large number of surface proteins which mediate cell-cell interactions, or cell adhesion to the extracellular matrix. In particular, RBCs adhere with high affinity to subendothelial matrix laminin via the basal cell adhesion molecule and Lutheran protein (BCAM/Lu). Here, we established SCFS as an in vitro technique to study human RBC adhesion at baseline and following biochemical treatment. Using blood obtained from healthy human subjects, we recorded adhesion forces from single RBCs attached to AFM cantilevers as the cell was pulled-off of substrates coated with laminin protein. We found that an increase in the overall cell adhesion measured via SCFS is correlated with an increase in the resultant total force measured on 1 µm(2) areas of the RBC membrane. Further, we showed that SCFS can detect significant changes in the adhesive response of RBCs to modulation of the cyclic adenosine monophosphate (cAMP) and protein kinase A (PKA) pathway. Lastly, we identified variability in the RBC adhesion force to laminin amongst the human subjects, suggesting that RBCs maintain diverse levels of active BCAM/Lu adhesion receptors. By using single-cell measurements, we established a powerful new method for the quantitative measurement of single RBC adhesion with specific receptor-mediated binding. Copyright © 2014 Elsevier Ltd. All rights reserved.

  2. Erythrocyte swelling and membrane hole formation in hypotonic media as studied by conductometry.

    PubMed

    Pribush, A; Meyerstein, D; Hatskelzon, L; Kozlov, V; Levi, I; Meyerstein, N

    2013-02-01

    Hypoosmotic swelling of erythrocytes and the formation of membrane holes were studied by measuring the dc conductance (G). In accordance with the theoretical predictions, these processes are manifested by a decrease in G followed by its increase. Thus, unlike the conventional osmotic fragility test, the proposed methodological approach allows investigations of both the kinetics of swelling and the erythrocyte fragility. It is shown that the initial rate of swelling and the equilibrium size of the cells are affected by the tonicity of a hypotonic solution and the membrane rheological properties. Because the rupture of biological membranes is a stochastic process, a time-dependent increase in the conductance follows an integral distribution function of the membrane lifetime. The main conclusion which stems from reported results is that information about rheological properties of red blood cell (RBC) membranes and the resistivity of RBCs to a certain osmotic shock may be extracted from conductance signals.

  3. Potential electron mediators to extract electron energies of RBC glycolysis for prolonged in vivo functional lifetime of hemoglobin vesicles.

    PubMed

    Kettisen, Karin; Bülow, Leif; Sakai, Hiromi

    2015-04-15

    Developing a functional blood substitute as an alternative to donated blood for clinical use is believed to relieve present and future blood shortages, and to reduce the risks of infection and blood type mismatching. Hemoglobin vesicle (HbV) encapsulates a purified and concentrated human-derived Hb solution in a phospholipid vesicle (liposome). The in vivo safety and efficacy of HbV as a transfusion alternative have been clarified. Auto-oxidation of ferrous Hb in HbV gradually increases the level of ferric methemoglobin (metHb) and impairs the oxygen transport capabilities. The extension of the functional half-life of HbV has recently been proposed using an electron mediator, methylene blue (MB), which acts as a shuttle between red blood cells (RBC) and HbV. MB transfers electron energies of NAD(P)H, produced by RBC glycolysis, to metHb in HbV. Work presented here focuses on screening of 15 potential electron mediators, with appropriate redox potential and water solubility, for electron transfer from RBC to HbV. The results are assessed with regard to the chemical properties of the candidates. The compounds examined in this study were dimethyl methylene blue (DMB), methylene green, azure A, azure B, azure C, toluidine blue (TDB), thionin acetate, phenazine methosulfate, brilliant cresyl blue, cresyl violet, gallocyanine, toluylene blue, indigo carmine, indigotetrasulfonate, and MB. Six candidates were found to be unsuitable because of their insufficient diffusion across membranes, or overly high or nonexistent reactivity with relevant biomolecules. However, 9 displayed favorable metHb reduction. Among the suitable candidates, phenothiazines DMB and TDB exhibited effectiveness like MB did. In comparison to MB, they showed faster reduction by electron-donating NAD(P)H, coupled with showing a lower rate of reoxidation in the presence of molecular oxygen. Ascertaining the best electron mediator can provide a pathway for extending the lifetime and efficiency of

  4. 77 FR 50487 - Application To Export Electric Energy; RBC Energy Services LP

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-08-21

    ... DEPARTMENT OF ENERGY [OE Docket No. EA-328-A] Application To Export Electric Energy; RBC Energy Services LP AGENCY: Office of Electricity Delivery and Energy Reliability, DOE. ACTION: Notice of application. SUMMARY: RBC Energy Services LP (RBC Energy) has applied to renew its authority to transmit...

  5. [Germ cell membrane lipids in spermatogenesis].

    PubMed

    Wang, Ting; Shi, Xiao; Quan, Song

    2016-05-01

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

  6. Increased adherence of sickled and phosphatidylserine-enriched human erythrocytes to cultured human peripheral blood monocytes.

    PubMed

    Schwartz, R S; Tanaka, Y; Fidler, I J; Chiu, D T; Lubin, B; Schroit, A J

    1985-06-01

    The precise mechanism by which sickle erythrocytes (RBC) are removed from the circulation is controversial, although it is possible that enhanced recognition of these cells by circulating mononuclear phagocytes could contribute to this process. We investigated this possibility by interacting sickle cells with cultured human peripheral blood monocytes. Our results show that both irreversibly sickled cells (ISC) and deoxygenated reversibly sickled cells (RSC) had a higher avidity for adherence to monocytes than did oxygenated sickle and normal RBC. ISC were the most adherent cell type. Adherence of RSC to monocytes was found to be reversible; reoxygenation of deoxygenated RSC resulted in a significant decrease in RSC--monocyte adherence. Concomitant with alterations in sickle RBC adherence were alterations in the organization and bilayer distribution of membrane phospholipids in these cells. Specifically, enhanced adherence was associated with increased exposure of RBC membrane outer leaflet phosphatidylserine (PS) and phosphatidylethanolamine, whereas lack of adherence was associated with normal patterns of membrane phospholipid distribution. To investigate the possibility of whether the exposure of PS in the outer membrane leaflet of these cells might be responsible for their recognition by monocytes, the membranes of normal RBC were enriched with the fluorescent PS analogue 1-acyl-2[(N-4-nitro-benzo-2-oxa-1,3-diazole)aminocaproyl]-phosphatidy lse rine (NBD-PS) via transfer of the exogenous lipid from a population of donor phospholipid vesicles (liposomes). RBC enriched with NBD-PS exhibited enhanced adherence to monocytes, whereas adherence of RBC enriched with similar amounts of NBD-phosphatidylcholine (NBD-PC) was not increased. Furthermore, preincubation of monocytes with PS liposomes resulted in a approximately 60% inhibition of ISC adherence to monocytes, whereas no inhibition occurred when monocytes were preincubated with PC liposomes. These findings

  7. The protective effect of aqueous extracts of roselle (Hibiscus sabdariffa L. UKMR-2) against red blood cell membrane oxidative stress in rats with streptozotocin-induced diabetes

    PubMed Central

    Mohamed, Jamaludin; Shing, Saw Wuan; Md Idris, Muhd Hanis; Budin, Siti Balkis; Zainalabidin, Satirah

    2013-01-01

    OBJECTIVES: The aim of this study was to investigate the protective effects of aqueous extracts of roselle (Hibiscus sabdariffa L. UKMR-2) against red blood cell (RBC) membrane oxidative stress in rats with streptozotocin-induced diabetes. METHODS: Forty male Sprague-Dawley rats weighing 230-250 g were randomly divided into four groups (n = 10 rats each): control group (N), roselle-treated control group, diabetic group, and roselle-treated diabetic group. Roselle was administered by force-feeding with aqueous extracts of roselle (100 mg/kg body weight) for 28 days. RESULTS: The results demonstrated that the malondialdehyde levels of the red blood cell membranes in the diabetic group were significantly higher than the levels in the roselle-treated control and roselle-treated diabetic groups. The protein carbonyl level was significantly higher in the roselle-treated diabetic group than in the roselle-treated control group but lower than that in the diabetic group. A significant increase in the red blood cell membrane superoxide dismutase enzyme was found in roselle-treated diabetic rats compared with roselle-treated control rats and diabetic rats. The total protein level of the red blood cell membrane, osmotic fragility, and red blood cell morphology were maintained. CONCLUSION: The present study demonstrates that aqueous extracts of roselle possess a protective effect against red blood cell membrane oxidative stress in rats with streptozotocin-induced diabetes. These data suggest that roselle can be used as a natural antioxidative supplement in the prevention of oxidative damage in diabetic patients. PMID:24212844

  8. RBC lithium transport in the psychoses.

    PubMed

    Hitzemann, R; Mark, C; Hirschowitz, J; Garver, D

    1989-02-01

    In vitro and in vivo red blood cell (RBC) lithium (Li+) intracellular/extracellular ratios were determined in 93 DSM-III schizophrenics (SCZ) in 47 DSM-III schizophreniform disorder patients (SF), in 22 DSM-III bipolar manics (M), in 15 affective disorders patients with mood-incongruent psychotic features (AD-MIP), and in 40 normal controls. There were no significant differences among groups in the in vitro Li+ ratio. Similarly, there was no significant difference among patient groups in the in vivo Li+ ratio. Furthermore, there was no significant difference in the mean Li+ ratios between the Li+-responsive and nonresponsive schizophrenic-like subjects. However, the distribution of Li+ ratios (both in vitro and in vivo) in the Li+-responsive group was significantly abnormal, showing more ratios in the extremes of the distribution (a platykurtic distribution).

  9. Effects of Storage-Aged RBC Transfusions on Endothelial Function in Hospitalized Patients

    PubMed Central

    Neuman, Robert; Hayek, Salim; Rahman, Ayaz; Poole, Joseph C.; Menon, Vivek; Sher, Salman; Newman, James L.; Karatela, Sulaiman; Polhemus, David; Lefer, David J.; De Staercke, Christine; Hooper, Craig; Quyyumi, Arshed A.; Roback, John D.

    2014-01-01

    Background Clinical and animal studies indicate that transfusions of older stored RBCs impair clinical outcomes as compared to fresh RBC transfusions. It has been suggested that this effect is due to inhibition of NO-mediated vasodilation following transfusion of older RBC units. However, to date this effect has not been identified in human transfusion recipients. Study Design and Methods Forty-three hospitalized patients with transfusion orders were randomized to receive either fresh (< 14 days) or older stored (> 21 days) RBC units. Prior to transfusion, and at selected time points after the start of transfusion, endothelial function was assessed using non-invasive flow-mediated dilation assays. Results Following transfusion of older RBC units, there was a significant reduction in NO-mediated vasodilation at 24 hours after transfusion (p=0.045), while fresh RBC transfusions had no effect (p=0.231). Conclusions The present study suggests for the first time a significant inhibitory effect of transfused RBC units stored > 21 days on NO-mediated vasodilation in anemic hospitalized patients. This finding lends further support to the hypothesis that deranged NO signaling mediates adverse clinical effects of older RBC transfusions. Future investigations will be necessary to address possible confounding factors and confirm these results. PMID:25393772

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

    NASA Technical Reports Server (NTRS)

    Kinder, James D.

    2005-01-01

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

  11. A cell culture technique for human epiretinal membranes to describe cell behavior and membrane contraction in vitro.

    PubMed

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

    2017-11-01

    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.

  12. Membrane oxidation in cell delivery and cell killing applications

    PubMed Central

    Wang, Ting-Yi; Libardo, M. Daben J.; Angeles-Boza, Alfredo M.; Pellois, Jean-Philippe

    2018-01-01

    Cell delivery or cell killing processes often involve the crossing or disruption of cellular membranes. We review how, by modifying the composition and properties of membranes, membrane oxidation can be exploited to enhance the delivery of macromolecular cargos into live human cells. We also describe how membrane oxidation can be utilized to achieve efficient killing of bacteria by antimicrobial peptides. Finally, we present recent evidence highlighting how membrane oxidation is intimately engaged in natural biological processes such as antigen delivery in dendritic cells and in the killing of bacteria by human macrophages. Overall, the insights that have been recently gained in this area should facilitate the development of more effective delivery technologies and antimicrobial therapeutic approaches. PMID:28355059

  13. Effect of Oral Docosahexaenoic Acid (DHA) Supplementation on DHA Levels and Omega-3 Index in Red Blood Cell Membranes of Breast Cancer Patients.

    PubMed

    Molfino, Alessio; Amabile, Maria I; Mazzucco, Sara; Biolo, Gianni; Farcomeni, Alessio; Ramaccini, Cesarina; Antonaroli, Simonetta; Monti, Massimo; Muscaritoli, Maurizio

    2017-01-01

    Rationale: Docosahexaenoic acid (DHA) in cell membrane may influence breast cancer (BC) patients' prognosis, affecting tumor cells sensitivity to chemo- and radio-therapy and likely modulating inflammation. The possibility of identifying BC patients presenting with low DHA levels and/or low ability of DHA incorporation into cell membrane might help to treat this condition. Methods: We enrolled BC patients and healthy controls, recording their seafood dietary intake. DHA in form of algal oil was administered for 10 consecutive days (2 g/day). Blood samples were collected at baseline (T0) and after 10 days of supplementation (T1) to assess DHA, omega-3 index, as the sum of DHA + eicosapentaenoic acid (EPA), in red blood cells (RBC) membranes and plasma tumor necrosis factor-alpha and interleukin-6 levels. Pre- and post-treatment fatty acid profiles were obtained by gas-chromatography. Parametric and non-parametric tests were performed, as appropriate, and P -value < 0.05 was considered statistically significant. Results: Forty-three women were studied, divided into 4 groups: 11 patients with BRCA1/2 gene mutation (M group), 12 patients with familiar positive history for BC (F group), 10 patients with sporadic BC (S group), and 10 healthy controls (C group). DHA and omega-3 index increased from T0 to T1 in the 3 groups of BC patients and in controls ( P < 0.001). No difference was found in DHA incorporation between each group of BC patients and between patients and controls, except for M group, which incorporated higher DHA levels with respect to controls (β = 0.42; P = 0.03). No association was documented between cytokines levels and DHA and omega-3 index at baseline and after DHA supplementation. Independent of the presence of BC, women considered as "good seafood consumers" showed at baseline DHA and omega-3 index higher with respect to "low seafood consumers" ( P = 0.04; P = 0.007, respectively). After supplementation, the increase in DHA levels was greater in

  14. 2-D Model for Normal and Sickle Cell Blood Microcirculation

    NASA Astrophysics Data System (ADS)

    Tekleab, Yonatan; Harris, Wesley

    2011-11-01

    Sickle cell disease (SCD) is a genetic disorder that alters the red blood cell (RBC) structure and function such that hemoglobin (Hb) cannot effectively bind and release oxygen. Previous computational models have been designed to study the microcirculation for insight into blood disorders such as SCD. Our novel 2-D computational model represents a fast, time efficient method developed to analyze flow dynamics, O2 diffusion, and cell deformation in the microcirculation. The model uses a finite difference, Crank-Nicholson scheme to compute the flow and O2 concentration, and the level set computational method to advect the RBC membrane on a staggered grid. Several sets of initial and boundary conditions were tested. Simulation data indicate a few parameters to be significant in the perturbation of the blood flow and O2 concentration profiles. Specifically, the Hill coefficient, arterial O2 partial pressure, O2 partial pressure at 50% Hb saturation, and cell membrane stiffness are significant factors. Results were found to be consistent with those of Le Floch [2010] and Secomb [2006].

  15. Sputter-deposited fuel cell membranes and electrodes

    NASA Technical Reports Server (NTRS)

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

    2001-01-01

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

  16. Simulations of molecular diffusion in lattices of cells: insights for NMR of red blood cells.

    PubMed Central

    Regan, David G; Kuchel, Philip W

    2002-01-01

    The pulsed field-gradient spin-echo (PGSE) nuclear magnetic resonance (NMR) experiment, conducted on a suspension of red blood cells (RBC) in a strong magnetic field yields a q-space plot consisting of a series of maxima and minima. This is mathematically analogous to a classical optical diffraction pattern. The method provides a noninvasive and novel means of characterizing cell suspensions that is sensitive to changes in cell shape and packing density. The positions of the features in a q-space plot characterize the rate of exchange across the membrane, cell dimensions, and packing density. A diffusion tensor, containing information regarding the diffusion anisotropy of the system, can also be derived from the PGSE NMR data. In this study, we carried out Monte Carlo simulations of diffusion in suspensions of "virtual" cells that had either biconcave disc (as in RBC) or oblate spheroid geometry. The simulations were performed in a PGSE NMR context thus enabling predictions of q-space and diffusion tensor data. The simulated data were compared with those from real PGSE NMR diffusion experiments on RBC suspensions that had a range of hematocrit values. Methods that facilitate the processing of q-space data were also developed. PMID:12080109

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

    PubMed

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

    2013-01-01

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

  18. Durability of PEM Fuel Cell Membranes

    NASA Astrophysics Data System (ADS)

    Huang, Xinyu; Reifsnider, Ken

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

  19. Discriminating plants using the DNA barcode rbcLb: an appraisal based on a large data set.

    PubMed

    Dong, Wenpan; Cheng, Tao; Li, Changhao; Xu, Chao; Long, Ping; Chen, Chumming; Zhou, Shiliang

    2014-03-01

    The ideal DNA barcode for plants remains to be discovered, and the candidate barcode rbcL has been met with considerable skepticism since its proposal. In fact, the variability within this gene has never been fully explored across all plant groups from algae to flowering plants, and its performance as a barcode has not been adequately tested. By analysing all of the rbcL sequences currently available in GenBank, we attempted to determine how well a region of rbcL performs as a barcode in species discrimination. We found that the rbcLb region was more variable than the frequently used rbcLa region. Both universal and plant group-specific primers were designed to amplify rbcLb, and the performance of rbcLa and rbcLb was tested in several ways. Using blast, both regions successfully identified all families and nearly all genera; however, the successful species identification rates varied significantly among plant groups, ranging from 24.58% to 85.50% for rbcLa and from 36.67% to 90.89% for rbcLb. Successful species discrimination ranged from 5.19% to 96.33% for rbcLa and from 22.09% to 98.43% for rbcLb in species-rich families, and from 0 to 88.73% for rbcLa and from 2.04% to 100% for rbcLb in species-rich genera. Both regions performed better for lower plants than for higher plants, although rbcLb performed significantly better than rbcLa overall, particularly for angiosperms. Considering the applicability across plants, easy and unambiguous alignment, high primer universality, high sequence quality and high species discrimination power for lower plants, we suggest rbcLb as a universal plant barcode. © 2013 John Wiley & Sons Ltd.

  20. Velocity measurements of heterogeneous RBC flow in capillary vessels using dynamic laser speckle signal.

    PubMed

    Li, Chenxi; Wang, Ruikang

    2017-04-01

    We propose an approach to measure heterogeneous velocities of red blood cells (RBCs) in capillary vessels using full-field time-varying dynamic speckle signals. The approach utilizes a low coherent laser speckle imaging system to record the instantaneous speckle pattern, followed by an eigen-decomposition-based filtering algorithm to extract dynamic speckle signal due to the moving RBCs. The velocity of heterogeneous RBC flows is determined by cross-correlating the temporal dynamic speckle signals obtained at adjacent locations. We verify the approach by imaging mouse pinna in vivo, demonstrating its capability for full-field RBC flow mapping and quantifying flow pattern with high resolution. It is expected to investigate the dynamic action of RBCs flow in capillaries under physiological changes.

  1. Velocity measurements of heterogeneous RBC flow in capillary vessels using dynamic laser speckle signal

    PubMed Central

    Li, Chenxi; Wang, Ruikang

    2017-01-01

    Abstract. We propose an approach to measure heterogeneous velocities of red blood cells (RBCs) in capillary vessels using full-field time-varying dynamic speckle signals. The approach utilizes a low coherent laser speckle imaging system to record the instantaneous speckle pattern, followed by an eigen-decomposition-based filtering algorithm to extract dynamic speckle signal due to the moving RBCs. The velocity of heterogeneous RBC flows is determined by cross-correlating the temporal dynamic speckle signals obtained at adjacent locations. We verify the approach by imaging mouse pinna in vivo, demonstrating its capability for full-field RBC flow mapping and quantifying flow pattern with high resolution. It is expected to investigate the dynamic action of RBCs flow in capillaries under physiological changes. PMID:28384709

  2. Membrane Electromechanics at Hair-Cell Synapses

    NASA Astrophysics Data System (ADS)

    Brownell, W. E.; Farrell, B.; Raphael, R. M.

    2003-02-01

    Both outer hair cell electromotility and neurotransmission at the inner hair cell synapse are rapid mechanical events that are synchronized to the hair-cell receptor potential. We analyze whether the forces and potentials resulting from membrane flexoelectricity could affect synaptic vesicle fusion. The results suggest that the coupling of membrane curvature with membrane potential is of sufficient magnitude to influence neurotransmitter release.

  3. Rotating Biological Contactors (RBC's). Student Manual. Biological Treatment Process Control.

    ERIC Educational Resources Information Center

    Zickefoose, Charles S.

    This student manual provides the textual material for a unit on rotating biological contactors (RBC's). Topic areas considered include: (1) flow patterns of water through RBC installations; (2) basic concepts (shaft and stage); (3) characteristics of biomass; (4) mechanical features (bearings, mechanical drive systems, and air drive systems); (5)…

  4. RBC Storage Effect on Coagulation, Microparticles and Microchimerism in Critically Ill Patients

    DTIC Science & Technology

    2015-03-01

    Award Number: W81XWH-11-2-0028 TITLE: “RBC Storage Effect on Coagulation, Microparticles and Microchimerism in Critically Ill Patients...27 DEC 2010 - 26 DEC 2015 – 4. TITLE AND SUBTITLE "“RBC Storage Effect on Coagulation, Microparticles and 5a. CONTRACT NUMBER Microchimerism in...15. SUBJECT TERMS RBC storage age; microchimerism; critically ill patients; coagulation; microparticles 16. SECURITY CLASSIFICATION OF: U 17

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

    PubMed

    Martinière, Alexandre; Runions, John

    2013-01-01

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

  6. The influence of saponins on cell membrane cholesterol.

    PubMed

    Böttger, Stefan; Melzig, Matthias F

    2013-11-15

    We studied the influence of structurally different saponins on the cholesterol content of cellular membranes. Therefore a cell culture model using ECV-304 urinary bladder carcinoma cells was developed. To measure the cholesterol content we used radiolabeled (3)H-cholesterol which is chemically and physiologically identical to natural cholesterol. The cells were pre-incubated with (3)H-cholesterol and after a medium change, they were treated with saponins to assess a saponin-induced cholesterol liberation from the cell membrane. In another experiment the cells were pre-incubated with saponins and after a medium change, they were treated with (3)H-cholesterol to assess a saponin-induced inhibition of cholesterol uptake into the cell membrane. Furthermore, the membrane toxicity of all applied saponins was analyzed using extracellular LDH quantification and the general cytotoxicity was analyzed using a colorimetric MTT-assay and DNA quantification. Our results revealed a correlation between membrane toxicity and general cytotoxicity. We also compared the results from the experiments on the saponin-induced cholesterol liberation as well as the saponin-induced inhibition of cholesterol uptake with the membrane toxicity. A significant reduction in the cell membrane cholesterol content was noted for those saponins who showed membrane toxicity (IC50 <60 μM). These potent membrane toxic saponins either liberated (3)H-cholesterol from intact cell membranes or blocked the integration of supplemented (3)H-cholesterol into the cell membrane. Saponins with little influence on the cell membrane (IC50 >100 μM) insignificantly altered the cell membrane cholesterol content. The results suggested that the general cytotoxicity of saponins is mainly dependent on their membrane toxicity and that the membrane toxicity might be caused by the loss of cholesterol from the cell membrane. We also analyzed the influence of a significantly membrane toxic saponin on the cholesterol content of

  7. Pharmacological inhibition of calpain-1 prevents red cell dehydration and reduces Gardos channel activity in a mouse model of sickle cell disease.

    PubMed

    De Franceschi, Lucia; Franco, Robert S; Bertoldi, Mariarita; Brugnara, Carlo; Matté, Alessandro; Siciliano, Angela; Wieschhaus, Adam J; Chishti, Athar H; Joiner, Clinton H

    2013-02-01

    Sickle cell disease (SCD) is a globally distributed hereditary red blood cell (RBC) disorder. One of the hallmarks of SCD is the presence of circulating dense RBCs, which are important in SCD-related clinical manifestations. In human dense sickle cells, we found reduced calpastatin activity and protein expression compared to either healthy RBCs or unfractionated sickle cells, suggesting an imbalance between activator and inhibitor of calpain-1 in favor of activator in dense sickle cells. Calpain-1 is a nonlysosomal cysteine proteinase that modulates multiple cell functions through the selective cleavage of proteins. To investigate the relevance of this observation in vivo, we evaluated the effects of the orally active inhibitor of calpain-1, BDA-410 (30 mg/kg/d), on RBCs from SAD mice, a mouse model for SCD. In SAD mice, BDA-410 improved RBC morphology, reduced RBC density (D(20); from 1106 ± 0.001 to 1100 ± 0.001 g/ml; P<0.05) and increased RBC-K(+) content (from 364 ± 10 to 429 ± 12.3 mmol/kg Hb; P<0.05), markedly reduced the activity of the Ca(2+)-activated K(+)channel (Gardos channel), and decreased membrane association of peroxiredoxin-2. The inhibitory effect of calphostin C, a specific inhibitor of protein kinase C (PKC), on the Gardos channel was eliminated after BDA-410 treatment, which suggests that calpain-1 inhibition affects the PKC-dependent fraction of the Gardos channel. BDA-410 prevented hypoxia-induced RBC dehydration and K(+) loss in SAD mice. These data suggest a potential role of BDA-410 as a novel therapeutic agent for treatment of SCD.

  8. Pf155/RESA protein influences the dynamic microcirculatory behavior of ring-stage Plasmodium falciparum infected red blood cells

    PubMed Central

    Diez-Silva, Monica; Park, YongKeun; Huang, Sha; Bow, Hansen; Mercereau-Puijalon, Odile; Deplaine, Guillaume; Lavazec, Catherine; Perrot, Sylvie; Bonnefoy, Serge; Feld, Michael S.; Han, Jongyoon; Dao, Ming; Suresh, Subra

    2012-01-01

    Proteins exported by Plasmodium falciparum to the red blood cell (RBC) membrane modify the structural properties of the parasitized RBC (Pf-RBC). Although quasi-static single cell assays show reduced ring-stage Pf-RBCs deformability, the parameters influencing their microcirculatory behavior remain unexplored. Here, we study the dynamic properties of ring-stage Pf-RBCs and the role of the parasite protein Pf155/Ring-Infected Erythrocyte Surface Antigen (RESA). Diffraction phase microscopy revealed RESA-driven decreased Pf-RBCs membrane fluctuations. Microfluidic experiments showed a RESA-dependent reduction in the Pf-RBCs transit velocity, which was potentiated at febrile temperature. In a microspheres filtration system, incubation at febrile temperature impaired traversal of RESA-expressing Pf-RBCs. These results show that RESA influences ring-stage Pf-RBCs microcirculation, an effect that is fever-enhanced. This is the first identification of a parasite factor influencing the dynamic circulation of young asexual Pf-RBCs in physiologically relevant conditions, offering novel possibilities for interventions to reduce parasite survival and pathogenesis in its human host. PMID:22937223

  9. Pf155/RESA protein influences the dynamic microcirculatory behavior of ring-stage Plasmodium falciparum infected red blood cells

    NASA Astrophysics Data System (ADS)

    Diez-Silva, Monica; Park, Yongkeun; Huang, Sha; Bow, Hansen; Mercereau-Puijalon, Odile; Deplaine, Guillaume; Lavazec, Catherine; Perrot, Sylvie; Bonnefoy, Serge; Feld, Michael S.; Han, Jongyoon; Dao, Ming; Suresh, Subra

    2012-08-01

    Proteins exported by Plasmodium falciparum to the red blood cell (RBC) membrane modify the structural properties of the parasitized RBC (Pf-RBC). Although quasi-static single cell assays show reduced ring-stage Pf-RBCs deformability, the parameters influencing their microcirculatory behavior remain unexplored. Here, we study the dynamic properties of ring-stage Pf-RBCs and the role of the parasite protein Pf155/Ring-Infected Erythrocyte Surface Antigen (RESA). Diffraction phase microscopy revealed RESA-driven decreased Pf-RBCs membrane fluctuations. Microfluidic experiments showed a RESA-dependent reduction in the Pf-RBCs transit velocity, which was potentiated at febrile temperature. In a microspheres filtration system, incubation at febrile temperature impaired traversal of RESA-expressing Pf-RBCs. These results show that RESA influences ring-stage Pf-RBCs microcirculation, an effect that is fever-enhanced. This is the first identification of a parasite factor influencing the dynamic circulation of young asexual Pf-RBCs in physiologically relevant conditions, offering novel possibilities for interventions to reduce parasite survival and pathogenesis in its human host.

  10. Nanobiotechnology: Cell Membrane-Based Delivery Systems.

    PubMed

    Zhang, Pengfei; Liu, Gang; Chen, Xiaoyuan

    2017-04-01

    The increasingly rapid pace of research in the field of bioinspired drug delivery systems is revealing the promise of cell membrane-based nanovesicles for biomedical applications. Those cell membrane-based nanoparticles combine the natural functionalities of cell plasma membranes and the bioengineering flexibility of synthetic nanomaterials, and such versatility provides a means of designing exciting new drug formulations for personalized treatment in future nanomedicine.

  11. Nitric oxide in red blood cell adaptation to hypoxia.

    PubMed

    Zhao, Yajin; Wang, Xiang; Noviana, Milody; Hou, Man

    2018-06-01

    Nitric oxide (NO) appears to be involved in virtually every aspect of cardiovascular biology. Most attention has been focused on the role of endothelial-derived NO in basal blood flow regulation by relaxing vascular smooth muscle; however, it is now known that NO derived from red blood cells (RBCs) plays a fundamental role in vascular homeostasis by enhancing oxygen (O2) release at the cellular and physiological level. Hypoxia is an often seen problem in diverse conditions; systemic adaptations to hypoxia permit people to adjust to the hypoxic environment at high altitudes and to disease processes. In addition to the cardiopulmonary and hematologic adaptations that support systemic O2 delivery in hypoxia, RBCs assist through newly described NO-based mechanisms, in line with their vital role in O2 transport and delivery. Furthermore, to increase the local blood flow in proportion to metabolic demand, NO regulates membrane mechanical properties thereby modulating RBC deformability and O2 carrying-releasing function. In this review article, we focus on the effect of NO bioactivity on RBC-based mechanisms that regulate blood flow and RBC deformability. RBC adaptations to hypoxia are summarized, with particular attention to NO-dependent S-nitrosylation of membrane proteins and hemoglobin (S-nitrosohemoglobin). The NO/S-nitrosylation/RBC vasoregulatory cascade contributes fundamentally to the molecular understanding of the role of NO in human adaptation to hypoxia and may inform novel therapeutic strategies.

  12. Red blood cell-deformability measurement: review of techniques.

    PubMed

    Musielak, M

    2009-01-01

    Cell-deformability characterization involves general measurement of highly complex relationships between cell biology and physical forces to which the cell is subjected. The review takes account of the modern technical solutions simulating the action of the force applied to the red blood cell in macro- and microcirculation. Diffraction ektacytometers and rheoscopes measure the mean deformability value for the total red blood cell population investigated and the deformation distribution index of individual cells, respectively. Deformation assays of a whole single cell are possible by means of optical tweezers. The single cell-measuring setups for micropipette aspiration and atomic force microscopy allow conducting a selective investigation of deformation parameters (e.g., cytoplasm viscosity, viscoelastic membrane properties). The distinction between instrument sensitivity to various RBC-rheological features as well as the influence of temperature on measurement are discussed. The reports quoted confront fascinating possibilities of the techniques with their medical applications since the RBC-deformability has the key position in the etiology of a wide range of conditions.

  13. Quantifying the deformation of the red blood cell skeleton in shear flow

    NASA Astrophysics Data System (ADS)

    Peng, Zhangli; Zhu, Qiang

    2012-02-01

    To quantitatively predict the response of red blood cell (RBC) membrane in shear flow, we carried out multiphysics simulations by coupling a three-level multiscale approach of RBC membranes with a Boundary Element Method (BEM) for surrounding flows. Our multiscale approach includes a model of spectrins with the domain unfolding feature, a molecular-based model of the junctional complex with detailed protein connectivity and a whole cell Finite Element Method (FEM) model with the bilayer-skeleton friction derived from measured transmembrane protein diffusivity based on the Einstein-Stokes relation. Applying this approach, we investigated the bilayer-skeleton slip and skeleton deformation of healthy RBCs and RBCs with hereditary spherocytosis anemia during tank-treading motion. Compared with healthy cells, cells with hereditary spherocytosis anemia sustain much larger skeleton-bilayer slip and area deformation of the skeleton due to deficiency of transmembrane proteins. This leads to extremely low skeleton density and large bilayer-skeleton interaction force, both of which may cause bilayer loss. This finding suggests a possible mechanism of the development of hereditary spherocytosis anemia.

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

    NASA Astrophysics Data System (ADS)

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

    2015-01-01

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

  15. Polymer electrolyte membrane assembly for fuel cells

    NASA Technical Reports Server (NTRS)

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

    2002-01-01

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

  16. Polymer electrolyte membrane assembly for fuel cells

    NASA Technical Reports Server (NTRS)

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

    2000-01-01

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

  17. Temperature-Sensitive Indicators for Monitoring RBC Concentrates Out of Controlled Temperature Storage.

    PubMed

    Sigle, Joerg P; Holbro, Andreas; Lehmann, Thomas; Infanti, Laura; Gerull, Sabine; Stern, Martin; Tichelli, Andre; Passweg, Jakob; Buser, Andreas

    2015-07-01

    The 30-minute rule for RBC concentrates out of controlled temperature storage does not take into account multiple parameters that influence warming of RBC concentrates. This study evaluated two temperature-sensitive indicators (TIs) for monitoring RBC concentrates during transport. TI labels (Check-Spot [Harald H. Temmel KG, Gleisdorf, Austria] and Thermoindikator V4 [BASF, Basel, Switzerland]) were attached to RBC concentrates prior to delivery. Duration of transport, ambient temperatures, and label results (valid vs expired) were recorded. We evaluated the proportion of labels discrepant to the 30-minute rule overall and among deliveries 30 minutes or less and more than 30 minutes and compared the rates of valid and expired readings between both TIs. In total, 201 RBC concentrate deliveries (86.6%) lasted 30 minutes or less, and 31 (13.4%) were more than 30 minutes. Forty-six (19.8%) Check-Spot and 37 (15.9%) Thermoindikator V4 results were discrepant to the 30-minute rule. Sixteen (51.6%) and 27 (87.1%) RBC concentrate deliveries more than 30 minutes displayed valid label readings with Check-Spot and Thermoindikator V4, respectively. Rates of expired labels among deliveries 30 minutes or less and valid labels among deliveries more than 30 minutes differed significantly between TIs (P < .01). TIs identified a considerable number of RBC concentrates whose temperatures may not be adequately reflected by the 30-minute rule. Variability of readings between TIs stresses the necessity of validation prior to implementation. Copyright© by the American Society for Clinical Pathology.

  18. Effect of Processing and Storage on RBC function in vivo

    PubMed Central

    Doctor, Allan; Spinella, Phil

    2012-01-01

    Red Blood Cell (RBC) transfusion is indicated to improve oxygen delivery to tissue, and for no other purpose. We have come to appreciate that donor RBCs are fundamentally altered during processing and storage, in a fashion that both impairs oxygen transport efficacy and introduces additional risk by perturbing both immune and coagulation systems. The protean biophysical and physiologic changes in RBC function arising from storage are termed the ‘storage lesion’; many have been understood for some time; for example, we know that the oxygen affinity of stored blood rises during the storage period1 and that intracellular allosteric regulators, notably 2,3-bisphosphoglyceric acid (DPG) and ATP, are depleted during storage. Our appreciation of other storage lesion features has emerged with improved understanding of coagulation, immune and vascular signaling systems. Herein we review key features of the ‘storage lesion’. Additionally, we call particular attention to the newly appreciated role of RBCs in regulating linkage between regional blood flow and regional O2 consumption by regulating the bioavailability of key vasoactive mediators in plasma, as well as discuss how processing and storage disturbs this key signaling function and impairs transfusion efficacy. PMID:22818545

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

    PubMed

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

    2012-05-01

    We are investigating a new class of penetration enhancers that enable poorly membrane-permeable molecules physically mixed with them to effectively penetrate cell membranes without their concomitant cellular uptake. Since we previously revealed that poly(N-vinylacetamide-co-acrylic acid) modified with d-octaarginine, which is a typical cell-penetrating peptide, significantly enhanced the nasal absorption of insulin, we examined the performance of the polymers on cell membranes. When Caco-2 cells were incubated with 5(6)-carboxyfluorescein (CF) for 30 min, approximately 0.1% of applied CF was internalized into the cells. This poor membrane permeability was dramatically enhanced by d-octaarginine-linked polymers; a 25-fold increase in the cellular uptake of CF was observed when the polymer concentration was adjusted to 0.2mg/mL. None of the individual components, for example, d-octaarginine, had any influence on CF uptake, demonstrating that only d-octaarginine anchored chemically to the polymeric platform enhanced the membrane permeation of CF. The polymer-induced CF uptake was consistently high even when the incubation time was extended to 120 min. Confocal laser scanning microphotographs of cells incubated with d-octaarginine-linked polymers bearing rhodamine red demonstrated that the cell outline was stained with red fluorescence. The polymer-induced CF uptake was significantly suppressed by 5-(N-ethyl-N-isopropyl)amiloride, which is an inhibitor of macropinocytosis. Results indicated that d-octaarginine-linked polymers remained on the cell membrane and poorly membrane-permeable CF was continuously internalized into cells mainly via macropinocytosis repeated for the individual peptidyl branches in the polymer backbone. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. Cell Membrane Coating Nanotechnology.

    PubMed

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

    2018-06-01

    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.

  1. Modification of host erythrocyte membranes by trypsin and chymotrypsin treatments and effects on the in vitro growth of bovine and equine Babesia parasites.

    PubMed

    Okamura, Masashi; Yokoyama, Naoaki; Takabatake, Noriyuki; Okubo, Kazuhiro; Ikehara, Yuzuru; Igarashi, Ikuo

    2007-02-01

    In the present study, we investigated the effects of protease pretreatments of host erythrocytes (RBC) on the in vitro growth of bovine Babesia parasites (Babesia bovis and B. bigemina) and equine Babesia parasites (B. equi and B. caballi). The selected proteases, trypsin and chymotrypsin, clearly modified several membrane proteins of both bovine and equine RBC, as demonstrated by SDS-PAGE analysis; however, the protease treatments also modified the sialic acid content exclusively in bovine RBC, as demonstrated by lectin blot analysis. An in vitro growth assay using the protease-treated RBC showed that the trypsin-treated bovine RBC, but not the chymotrypsin-treated ones, significantly reduced the growth of B. bovis and B. bigemina as compared to the control. In contrast, the growth of B. equi and B. caballi was not affected by any of these proteases. Thus, the bovine, but not the equine, Babesia parasites require the trypsin-sensitive membrane (sialoglyco) proteins to infect the RBC.

  2. The effect of syndecan-4 and glypican-1 knockdown on the proliferation and differentiation of turkey satellite cells differing in age and growth rates.

    PubMed

    Velleman, Sandra G; Clark, Daniel L; Tonniges, Jeffrey R

    2018-09-01

    Posthatch skeletal muscle growth requires myogenic satellite cells and the dynamic expression of cell membrane-associated proteins. The membrane associated heparan sulfate proteoglycans, syndecan-4 and glypican-1, link the satellite cell niche to the intracellular environment. Sydnecan-4 and glypican-1 are differentially expressed with age in turkey satellite cells and their over-expression impacts both satellite cell proliferation and differentiation, but their effect on satellite cells from lines with different growth potentials is not known. The objective of the current study was to determine if syndecan-4 and glypican-1 regulation of satellite cell proliferation and differentiation is affected by age and growth selection. Pectoralis major satellite cells isolated at 1 d, 7 and 16-wk of age from a Randombred Control 2 (RBC2) line and a 16-wk body weight (F) line selected from the RBC2 line turkeys were studied. Syndecan-4 and glypican-1 expression was knocked down in both lines. The F-line cells proliferated faster than RBC2 line cells regardless of age, while differentiation tended to be greater in RBC2 line cells than F-line cells at each age. Syndecan-4 knockdown decreased proliferation at 7- and 16-wk but not 1 d cells, and increased differentiation at 1 d and 7 wk but not 16 wk cells. Glypican-1 knockdown differentially affected proliferation depending on cell age, whereas differentiation was decreased for 7- and 16-wk but not 1 d cells. These data suggest syndecan-4 and glypican-1 differentially affected satellite cell function in an age-dependent manner, but had little impact on differences in proliferation and differentiation due to growth selection. Copyright © 2018. Published by Elsevier Inc.

  3. Membrane tension and cytoskeleton organization in cell motility.

    PubMed

    Sens, Pierre; Plastino, Julie

    2015-07-15

    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.

  4. Fixed charge in the cell membrane

    PubMed Central

    Elul, R.

    1967-01-01

    1. Focal electric field was generated by passing a current of 5 × 10-7 to 1 × 10-5 A from a micropipette into the culture medium. Movement of cells at a distance of 5-50 μ from the electrode tip was observed. In case of cells embedded in the culture only local deformation of the membrane was observed. 2. The cell species explored included neurones, glia, muscle fibres, connective cells, malignant cells and erythrocytes. All cells responded in a similar manner to the electric field, and the current required was in the same range. 3. Cells were attracted to a positive micropipette and repelled from a negative one: the only exception was observed in certain malignant cells which moved in the opposite direction. 4. Movement and membrane deformation could be obtained with electrodes filled with various concentrated and isotonic solutions. The composition of the culture medium also had no qualitative influence on these effects. 5. Metabolic poisons or rupture of the cell membrane had no effect on the movement. Isolated membrane fragments showed movement similar to that of intact cells. 6. The possibility of artifacts due to proximity of the focal electrode is considered. It is shown that electro-osmosis cannot account for the present observations. Some other artifacts are also excluded. 7. It is proposed that the most satisfactory way to account for the present observations is by a membrane carrying negative fixed charge of the order of 2·5 × 103 e.s.u./cm2. Some physiological consequences of presence of negative charge in the membrane are briefly discussed. ImagesFig. 1Fig. 2Fig. 3 PMID:6040152

  5. Investigating catalyst coated membrane equilibration time for polymer electrolyte membrane fuel cell manufacturing

    NASA Astrophysics Data System (ADS)

    Cote, Philippe

    Mercedes-Benz Canada Inc., Fuel Cell Division, manufactures polymer electrolyte membrane fuel cell stacks for use in vehicles. The manufacturing line is being optimized for efficiency and quality control, in order to uphold the high standards of Mercedes-Benz Inc. vehicles. In an operating polymer electrolyte membrane fuel cell, the catalyst coated membrane facilitates the electrochemical reaction that generates electricity. This research examines the equilibration of catalyst coated membrane rolls to controlled temperature and humidity conditions, before they are used in the manufacturing of polymer electrolyte membrane fuel cells. Equilibration involves allowing the water content in the catalyst coated membrane to stabilize at the controlled conditions, in order to reduce mechanical stress in the material for better manufacturability. Initial equilibration measurements were conducted on discrete catalyst coated membrane samples using novel electronic conductivity measurements of the catalyst layer, and compared to ionic conductivity measurements of the membrane. Electronic conductivity measurements are easier to implement in the manufacturing environment than the more complex ionic conductivity measurements. When testing discrete catalyst coated membrane samples in an environmental chamber, the equilibration trends for the measured ionic and electronic conductivity signals were similar enough to permit us to adapt the electronic conductivity measurements for catalyst coated membrane in roll form. Equilibration measurements of catalyst coated membrane rolls were optimized to achieve a robust and repeatable procedure which could be used in the manufacturing environment at Mercedes-Benz Canada Inc., Fuel Cell Division.

  6. Biocompatible water soluble quantum dots as new biophotonic tools for hematologic cells: applications for flow cell cytometry

    NASA Astrophysics Data System (ADS)

    Lira, Rafael B.; de Sales Neto, Antonio T.; Carvalho, Kilmara K. H. G.; Leite, Elisa S.; Brasil, Aluizio G., Jr.; Azevedo, Denise P. L.; Cabral Filho, Paulo E.; Cavalcanti, Mariana B.; Amaral, Ademir J.; Farias, Patricía M. A.; Santos, Beate S.; Fontes, Adriana

    2010-02-01

    Quantum dots (QDs) are a promising class of fluorescent probes that can be conjugated to a variety of specific cell antibodies. For this reason, simple, cheap and reproducible routes of QDśs syntheses are the main goal of many researches in this field. The main objective of this work was to demonstrate the ability of QDs as biolabels for flow cell cytometry analysis. We have synthesized biocompatible water soluble CdS/Cd(OH)2 and CdTe/CdS QDs and applied them as fluorescent labels of hematologic cells. CdTe/CdS QDs was prepared using using a simple aqueous route with mercaptoacetic acid and mercaptopropionic acid as stabilizing agents. The resulting CdTe/CdS QDs can target biological membrane proteins and can also be internalized by cells. We applied the CdTe/CdS QDs as biolabels of human lymphocytes and compared the results obtained for lymphocytes treated and non-treated with permeabilizing agents for cell membranes. Permeabilized cells present higher fluorescence pattern than non permeabilized ones. We associated antibody A to the CdS/Cd(OH)2 QDs to label type A red blood cell (RBC). In this case, the O erythrocytes were used as the negative control. The results demonstrate that QDs were successfully functionalized with antibody A. There was a specific binding of QDs-antibody A to RBC membrane antigen only for A RBCs. We have also monitored QDs-hematologic cell interaction by using fluorescence microscopy. Our method shows that QDs can be conjugated to a variety of specific cell antibodies and can become a potential, highly efficient and low cost diagnostic tool for flow cell cytometry, very compatible with the lasers and filters used in this kind of equipments.

  7. Plasma membrane changes during programmed cell deaths

    PubMed Central

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

    2018-01-01

    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. PMID:29076500

  8. Cytosolic Extract Induces Tir Translocation and Pedestals in EPEC-Infected Red Blood Cells

    PubMed Central

    Swimm, Alyson I; Kalman, Daniel

    2008-01-01

    Enteropathogenic Escherichia coli (EPEC) are deadly contaminants in water and food, and induce protrusion of actin-filled membranous pedestals beneath themselves upon attachment to intestinal epithelia. Pedestal formation requires clustering of Tir and subsequent recruitment of cellular tyrosine kinases including Abl, Arg, and Etk as well as signaling molecules Nck, N-WASP, and Arp2/3 complex. We have developed a cytosolic extract-based cellular system that recapitulates actin pedestal formation in permeabilized red blood cells (RBC) infected with EPEC. RBC support attachment of EPEC and translocation of virulence factors, but not pedestal formation. We show here that extract induces a rapid Ca++-dependent release of Tir from the EPEC Type III secretion system, and that cytoplasmic factor(s) present in the extract facilitate translocation of Tir into the RBC plasma membrane. We show that Abl and related kinases in the extract phosphorylate Tir and that actin polymerization can be reconstituted in infected RBC following addition of cytosolic extract. Reconstitution requires the bacterial virulence factors Tir and intimin, and phosphorylation of Tir on tyrosine residue 474 results in the recruitment of Nck, N-WASP, and Arp2/3 complex beneath attached bacteria at sites of actin polymerization. Together these data describe a biochemical system for dissection of host components that mediate Type III secretion and the mechanisms by which complexes of proteins are recruited to discrete sites within the plasma membrane to initiate localized actin polymerization and morphological changes. PMID:18208322

  9. Controlled lecithin release from a hierarchical architecture on blood-contacting surface to reduce hemolysis of stored red blood cells.

    PubMed

    Shi, Qiang; Fan, Qunfu; Ye, Wei; Hou, Jianwen; Wong, Shing-Chung; Xu, Xiaodong; Yin, Jinghua

    2014-06-25

    Hemolysis of red blood cells (RBCs) caused by implant devices in vivo and nonpolyvinyl chloride containers for RBC preservation in vitro has recently gained much attention. To develop blood-contacting biomaterials with long-term antihemolysis capability, we present a facile method to construct a hydrophilic, 3D hierarchical architecture on the surface of styrene-b-(ethylene-co-butylene)-b-styrene elastomer (SEBS) with poly(ethylene oxide) (PEO)/lecithin nano/microfibers. The strategy is based on electrospinning of PEO/lecithin fibers onto the surface of poly [poly(ethylene glycol) methyl ether methacrylate] [P(PEGMEMA)]-modified SEBS, which renders SEBS suitable for RBC storage in vitro. We demonstrate that the constructed 3D architecture is composed of hydrophilic micro- and nanofibers, which transforms to hydrogel networks immediately in blood; the controlled release of lecithin is achieved by gradual dissolution of PEO/lecithin hydrogels, and the interaction of lecithin with RBCs maintains the membrane flexibility and normal RBC shape. Thus, the blood-contacting surface reduces both mechanical and oxidative damage to RBC membranes, resulting in low hemolysis of preserved RBCs. This work not only paves new way to fabricate high hemocompatible biomaterials for RBC storage in vitro, but provides basic principles to design and develop antihemolysis biomaterials for implantation in vivo.

  10. Validation of the omega-3 fatty acid intake measured by a web-based food frequency questionnaire against omega-3 fatty acids in red blood cells in men with prostate cancer.

    PubMed

    Allaire, J; Moreel, X; Labonté, M-È; Léger, C; Caron, A; Julien, P; Lamarche, B; Fradet, V

    2015-09-01

    The objective of this study was to evaluate the ability of a web-based self-administered food frequency questionnaire (web-FFQ) to assess the omega-3 (ω-3) fatty acids (FAs) intake of men affected with prostate cancer (PCa) against a biomarker. The study presented herein is a sub-study from a phase II clinical trial. Enrolled patients afflicted with PCa were included in the sub-study analysis if the FA profiles from the red blood cell (RBC) membranes and FA intakes at baseline were both determined at the time of the data analysis (n=60). Spearman's correlation coefficients were calculated to estimate the correlations between FA intakes and their proportions in the RBC membranes. Intakes of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were highly correlated with their respective proportions in the RBC membranes (both rs=0.593, P<0.0001). Correlation between alpha-linolenic acid (ALA) intake and its proportion in RBC was not significant (rs=0.130, P=0.332). Correlations were observed between fatty fish intake and total ω-3 FAs (rs=0.304, P=0.02), total long-chain ω-3 FAs (rs=0.290, P=0.03) and DHA (rs=0.328, P=0.01) in RBC membranes. This study has shown that the web-FFQ is an accurate tool to assess total long-chain ω-3 FAs, EPA and DHA but not ALA intake in clinical trials and epidemiological studies carried out in men with PCa.

  11. A novel bioactive membrane by cell electrospinning.

    PubMed

    Chen, Haiping; Liu, Yuanyuan; Hu, Qingxi

    2015-11-01

    Electrospinning permits fabrication of biodegradable matrices that can resemble the both scale and mechanical behavior of the native extracellular matrix. However, achieving high-cellular density and infiltration of cells within matrices with traditional technique remain challenging and time consuming. The cell electrospinning technique presented in this paper can mitigate the problems associated with these limitations. Cells encapsulated by the material in the cell electrospinning technique survived well and distributed homogenously within the nanofibrous membrane, and their vitality was improved to 133% after being cultured for 28 days. The electrospun nanofibrous membrane has a certain degradation property and favorable cell-membrane interaction that supports the active biocompatibility of the membrane. Its properties are helpful for supporting cell attachment and growth, maintaining phenotypic shape, and secreting an ample amount of extracellular matrix (ECM). This novel membrane may be a potential application within the field of tissue engineering. The ability of cell electrospinning to microintegrate cells into a biodegradable fibrous matrix embodies a novel tissue engineering approach that could be applied to fabricate a high cell density elastic tissue mimetic. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Cell cycle dependent changes in the plasma membrane organization of mammalian cells.

    PubMed

    Denz, Manuela; Chiantia, Salvatore; Herrmann, Andreas; Mueller, Peter; Korte, Thomas; Schwarzer, Roland

    2017-03-01

    Lipid membranes are major structural elements of all eukaryotic and prokaryotic organisms. Although many aspects of their biology have been studied extensively, their dynamics and lateral heterogeneity are still not fully understood. Recently, we observed a cell-to-cell variability in the plasma membrane organization of CHO-K1 cells (Schwarzer et al., 2014). We surmised that cell cycle dependent changes of the individual cells from our unsynchronized cell population account for this phenomenon. In the present study, this hypothesis was tested. To this aim, CHO-K1 cells were arrested in different cell cycle phases by chemical treatments, and the order of their plasma membranes was determined by various fluorescent lipid analogues using fluorescence lifetime imaging microscopy. Our experiments exhibit significant differences in the membrane order of cells arrested in the G2/M or S phase compared to control cells. Our single-cell analysis also enabled the specific selection of mitotic cells, which displayed a significant increase of the membrane order compared to the control. In addition, the lipid raft marker GPImYFP was used to study the lateral organization of cell cycle arrested cells as well as mitotic cells and freely cycling samples. Again, significant differences were found between control and arrested cells and even more pronounced between control and mitotic cells. Our data demonstrate a direct correlation between cell cycle progression and plasma membrane organization, underlining that cell-to-cell heterogeneities of membrane properties have to be taken into account in cellular studies especially at the single-cell level. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Stiffening of Red Blood Cells Induced by Cytoskeleton Disorders: A Joint Theory-Experiment Study.

    PubMed

    Lai, Lipeng; Xu, Xiaofeng; Lim, Chwee Teck; Cao, Jianshu

    2015-12-01

    The functions and elasticities of the cell are largely related to the structures of the cytoskeletons underlying the lipid bilayer. Among various cell types, the red blood cell (RBC) possesses a relatively simple cytoskeletal structure. Underneath the membrane, the RBC cytoskeleton takes the form of a two-dimensional triangular network, consisting of nodes of actins (and other proteins) and edges of spectrins. Recent experiments focusing on the malaria-infected RBCs (iRBCs) show that there is a correlation between the elongation of spectrins in the cytoskeletal network and the stiffening of the iRBCs. Here we rationalize the correlation between these two observations by combining the wormlike chain model for single spectrins and the effective medium theory for the network elasticity. We specifically focus on how the disorders in the cytoskeletal network affect its macroscopic elasticity. Analytical and numerical solutions from our model reveal that the stiffness of the membrane increases with increasing end-to-end distances of spectrins, but has a nonmonotonic dependence on the variance of the end-to-end distance distributions. These predictions are verified quantitatively by our atomic force microscopy and micropipette aspiration measurements of iRBCs. The model may, from a molecular level, provide guidelines for future identification of new treatment methods for RBC-related diseases, such as malaria infection. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  14. Effect of Sex and Prior Exposure to a Cafeteria Diet on the Distribution of Sex Hormones between Plasma and Blood Cells

    PubMed Central

    Romero, María del Mar; Fernández-López, José Antonio; Remesar, Xavier; Alemany, Marià

    2012-01-01

    It is generally assumed that steroid hormones are carried in the blood free and/or bound to plasma proteins. We investigated whether blood cells were also able to bind/carry sex-related hormones: estrone, estradiol, DHEA and testosterone. Wistar male and female rats were fed a cafeteria diet for 30 days, which induced overweight. The rats were fed the standard rat diet for 15 additional days to minimize the immediate effects of excess ingested energy. Controls were always kept on standard diet. After the rats were killed, their blood was used for 1) measuring plasma hormone levels, 2) determining the binding of labeled hormones to washed red blood cells (RBC), 3) incubating whole blood with labeled hormones and determining the distribution of label between plasma and packed cells, discounting the trapped plasma volume, 4) determining free plasma hormone using labeled hormones, both through membrane ultrafiltration and dextran-charcoal removal. The results were computed individually for each rat. Cells retained up to 32% estrone, and down to 10% of testosterone, with marked differences due to sex and diet (the latter only for estrogens, not for DHEA and testosterone). Sex and diet also affected the concentrations of all hormones, with no significant diet effects for estradiol and DHEA, but with considerable interaction between both factors. Binding to RBC was non-specific for all hormones. Estrogen distribution in plasma compartments was affected by sex and diet. In conclusion: a) there is a large non-specific RBC-carried compartment for estrone, estradiol, DHEA and testosterone deeply affected by sex; b) Prior exposure to a cafeteria (hyperlipidic) diet induced hormone distribution changes, affected by sex, which hint at sex-related structural differences in RBC membranes; c) We postulate that the RBC compartment may contribute to maintain free (i.e., fully active) sex hormone levels in a way similar to plasma proteins non-specific binding. PMID:22479617

  15. Functional dynamics of cell surface membrane proteins

    NASA Astrophysics Data System (ADS)

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

    2014-04-01

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

  16. An rbcL mRNA-binding protein is associated with C3 to C4 evolution and light-induced production of Rubisco in Flaveria.

    PubMed

    Yerramsetty, Pradeep; Agar, Erin M; Yim, Won C; Cushman, John C; Berry, James O

    2017-07-20

    Nuclear-encoded RLSB protein binds chloroplastic rbcL mRNA encoding the Rubisco large subunit. RLSB is highly conserved across all groups of land plants and is associated with positive post-transcriptional regulation of rbcL expression. In C3 leaves, RLSB and Rubisco occur in all chlorenchyma cell chloroplasts, while in C4 leaves these accumulate only within bundle sheath (BS) chloroplasts. RLSB's role in rbcL expression makes modification of its localization a likely prerequisite for the evolutionary restriction of Rubisco to BS cells. Taking advantage of evolutionarily conserved RLSB orthologs in several C3, C3-C4, C4-like, and C4 photosynthetic types within the genus Flaveria, we show that low level RLSB sequence divergence and modification to BS specificity coincided with ontogeny of Rubisco specificity and Kranz anatomy during C3 to C4 evolution. In both C3 and C4 species, Rubisco production reflected RLSB production in all cell types, tissues, and conditions examined. Co-localization occurred only in photosynthetic tissues, and both proteins were co-ordinately induced by light at post-transcriptional levels. RLSB is currently the only mRNA-binding protein to be associated with rbcL gene regulation in any plant, with variations in sequence and acquisition of cell type specificity reflecting the progression of C4 evolution within the genus Flaveria. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  17. VIEW OF RBC (REFINED BICARBONATE) BUILDING LOOKING NORTHEAST. DEMOLITION IN ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    VIEW OF RBC (REFINED BICARBONATE) BUILDING LOOKING NORTHEAST. DEMOLITION IN PROGRESS. "ARM & HAMMER BAKING SODA WAS MADE HERE FOR OVER 50 YEARS AND THEN SHIPPED ACROSS THE STREET TO THE CHURCH & DWIGHT PLANT ON WILLIS AVE. (ON THE RIGHT IN THIS PHOTO). LAYING ON THE GROUND IN FRONT OF C&D BUILDING IS PART OF AN RBC DRYING TOWER. - Solvay Process Company, Refined Bicarbonate Building, Between Willis & Milton Avenues, Solvay, Onondaga County, NY

  18. Dietary fat intake and red blood cell fatty acid composition of children and women from three different geographical areas in South Africa.

    PubMed

    Ford, Rosalyn; Faber, Mieke; Kunneke, Ernesta; Smuts, Cornelius M

    2016-06-01

    Dietary fat intake, particularly the type of fat, is reflected in the red blood cell (RBC) fatty acid (FA) profile and is vital in growth, development and health maintenance. The FA profile (%wt/wt) of RBC membrane phospholipids (as determined by gas chromatography) and dietary intake (as determined by 24h recall) was assessed in 2-6y old South African children and their caregivers randomly selected from three communities, i.e. an urban Northern Cape community (urban-NC; n=104), an urban coastal Western Cape community (urban-WC; n=93) and a rural Limpopo Province community (rural-LP; n=102). Mean RBC FA values across groups were compared using ANOVA and Bonferroni post-hoc test while controlling for age and gender (children); median dietary intake values were compared using a Kruskal-Wallis test. Dietary intakes for total fat, saturated FAs and polyunsaturated FAs were higher in the two urban areas compared to the rural area. Total fat intake in rural-LP, and omega-3 FA dietary intake in all three areas were lower than the South African adopted guidelines. Dietary SFA intake in both urban areas was higher than recommended by South African guidelines; this was reflected in the RBC membrane FA profile. Rural-LP children had the lowest intake of omega-3 and omega-6 FAs yet presented with the highest RBC docosahexaenoic acid (DHA) profile and highest arachidonic acid percentage. Although differences observed in dietary fat intake between the two urban and the rural area were reflected in the RBC membrane total phospholipid FA profile, the lowest total fat and α-linolenic acid (ALA) intake by rural children that presented with the highest RBC DHA profile warrants further investigation. Copyright © 2016 Elsevier Ltd. All rights reserved.

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

    PubMed

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

    2008-10-01

    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 mechanisms that regulate proliferation and differentiation. The comparison of their membrane proteomes will help unravel the biological principles of pluripotency, and the identification of biomarker proteins in their plasma membranes is considered a crucial step to fully exploit pluripotent cells for therapeutic purposes. For these tasks, membrane proteomics is the method of choice, but as indicated by the scarce identification of membrane and plasma membrane proteins in global proteomic surveys it is not an easy task. In this minireview, we first describe the general challenges of membrane proteomics. We then review current sample preparation steps and discuss protocols that we found particularly beneficial for the identification of large numbers of (plasma) membrane proteins in human tumour- and embryo-derived stem cells. Our optimized assembled protocol led to the identification of a large number of membrane proteins. However, as the composition of cells and membranes is highly variable we still recommend adapting the sample preparation protocol for each individual system.

  20. Functional dynamics of cell surface membrane proteins.

    PubMed

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

    2014-04-01

    Cell surface receptors are integral membrane proteins that receive external stimuli, and transmit signals across plasma membranes. In the conventional view of receptor activation, ligand binding to the extracellular side of the receptor induces conformational changes, which convert the structure of the receptor into an active conformation. However, recent NMR studies of cell surface membrane proteins have revealed that their structures are more dynamic than previously envisioned, and they fluctuate between multiple conformations in an equilibrium on various timescales. In addition, NMR analyses, along with biochemical and cell biological experiments indicated that such dynamical properties are critical for the proper functions of the receptors. In this review, we will describe several NMR studies that revealed direct linkage between the structural dynamics and the functions of the cell surface membrane proteins, such as G-protein coupled receptors (GPCRs), ion channels, membrane transporters, and cell adhesion molecules. Copyright © 2013 Elsevier Inc. All rights reserved.

  1. Preoperative diagnosis of orbital cavernous hemangioma: a 99mTc-RBC SPECT study.

    PubMed

    Burroni, Luca; Borsari, Giulia; Pichierri, Patrizia; Polito, Ennio; Toscano, Olga; Grassetto, Gaia; Al-Nahhas, Adil; Rubello, Domenico; Vattimo, Angelo Giuseppe

    2012-11-01

    This study aimed to describe 99mTc-labeled RBC scintigraphy as a diagnostic method for orbital cavernous hemangiomas and to evaluate this diagnostic tool according to surgical outcomes. Fifty-five patients with clinical and radiological (US, CT, and/or MRI) suspicion of unilateral cavernous hemangioma of the orbit underwent 99mTc-RBC SPECT study.Qualitative and semiquantitative evaluations were performed, and results were statistically analyzed. SPECT images showed focal uptake in the orbital mass in 36 of 55 patients. Nineteen patients had a negative scintigraphic pattern, with concordance of early and late absence of uptake of 99mTc-RBC.Our procedure showed 100% sensitivity and 88.9% specificity for the diagnosis of orbital cavernous hemangioma, with a positive predictive value of 90.9% and a negative predictive value of 100%. 99mTc-RBC imaging is safe, easy to perform, and highly accurate in providing adequate clinical and surgical management. As a noninvasive and highly specific method for diagnosing orbital hemangioma, 99mTc-RBC scintigraphy can avoid more invasive imaging or biopsy.

  2. Fuel cell with ionization membrane

    NASA Technical Reports Server (NTRS)

    Hartley, Frank T. (Inventor)

    2007-01-01

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

  3. Separation of neural stem cells by whole cell membrane capacitance using dielectrophoresis.

    PubMed

    Adams, Tayloria N G; Jiang, Alan Y L; Vyas, Prema D; Flanagan, Lisa A

    2018-01-15

    Whole cell membrane capacitance is an electrophysiological property of the plasma membrane that serves as a biomarker for stem cell fate potential. Neural stem and progenitor cells (NSPCs) that differ in ability to form neurons or astrocytes are distinguished by membrane capacitance measured by dielectrophoresis (DEP). Differences in membrane capacitance are sufficient to enable the enrichment of neuron- or astrocyte-forming cells by DEP, showing the separation of stem cells on the basis of fate potential by membrane capacitance. NSPCs sorted by DEP need not be labeled and do not experience toxic effects from the sorting procedure. Other stem cell populations also display shifts in membrane capacitance as cells differentiate to a particular fate, clarifying the value of sorting a variety of stem cell types by capacitance. Here, we describe methods developed by our lab for separating NSPCs on the basis of capacitance using several types of DEP microfluidic devices, providing basic information on the sorting procedure as well as specific advantages and disadvantages of each device. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. At the border: the plasma membrane-cell wall continuum.

    PubMed

    Liu, Zengyu; Persson, Staffan; Sánchez-Rodríguez, Clara

    2015-03-01

    Plant cells rely on their cell walls for directed growth and environmental adaptation. Synthesis and remodelling of the cell walls are membrane-related processes. During cell growth and exposure to external stimuli, there is a constant exchange of lipids, proteins, and other cell wall components between the cytosol and the plasma membrane/apoplast. This exchange of material and the localization of cell wall proteins at certain spots in the plasma membrane seem to rely on a particular membrane composition. In addition, sensors at the plasma membrane detect changes in the cell wall architecture, and activate cytoplasmic signalling schemes and ultimately cell wall remodelling. The apoplastic polysaccharide matrix is, on the other hand, crucial for preventing proteins diffusing uncontrollably in the membrane. Therefore, the cell wall-plasma membrane link is essential for plant development and responses to external stimuli. This review focuses on the relationship between the cell wall and plasma membrane, and its importance for plant tissue organization. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  5. Functional imaging of microdomains in cell membranes.

    PubMed

    Duggan, James; Jamal, Ghadir; Tilley, Mark; Davis, Ben; McKenzie, Graeme; Vere, Kelly; Somekh, Michael G; O'Shea, Paul; Harris, Helen

    2008-10-01

    The presence of microdomains or rafts within cell membranes is a topic of intense study and debate. The role of these structures in cell physiology, however, is also not yet fully understood with many outstanding problems. This problem is partly based on the small size of raft structures that presents significant problems to their in vivo study, i.e., within live cell membranes. But the structure and dynamics as well as the factors that control the assembly and disassembly of rafts are also of major interest. In this review we outline some of the problems that the study of rafts in cell membranes present as well as describing some views of what are considered the generalised functions of membrane rafts. We point to the possibility that there may be several different 'types' of membrane raft in cell membranes and consider the factors that affect raft assembly and disassembly, particularly, as some researchers suggest that the lifetimes of rafts in cell membranes may be sub-second. We attempt to review some of the methods that offer the ability to interrogate rafts directly as well as describing factors that appear to affect their functionality. The former include both near-field and far-field optical approaches as well as scanning probe techniques. Some of the advantages and disadvantages of these techniques are outlined. Finally, we describe our own views of raft functionality and properties, particularly, concerning the membrane dipole potential, and describe briefly some of the imaging strategies we have developed for their study.

  6. Pharmacological inhibition of calpain-1 prevents red cell dehydration and reduces Gardos channel activity in a mouse model of sickle cell disease

    PubMed Central

    De Franceschi, Lucia; Franco, Robert S.; Bertoldi, Mariarita; Brugnara, Carlo; Matté, Alessandro; Siciliano, Angela; Wieschhaus, Adam J.; Chishti, Athar H.; Joiner, Clinton H.

    2013-01-01

    Sickle cell disease (SCD) is a globally distributed hereditary red blood cell (RBC) disorder. One of the hallmarks of SCD is the presence of circulating dense RBCs, which are important in SCD-related clinical manifestations. In human dense sickle cells, we found reduced calpastatin activity and protein expression compared to either healthy RBCs or unfractionated sickle cells, suggesting an imbalance between activator and inhibitor of calpain-1 in favor of activator in dense sickle cells. Calpain-1 is a nonlysosomal cysteine proteinase that modulates multiple cell functions through the selective cleavage of proteins. To investigate the relevance of this observation in vivo, we evaluated the effects of the orally active inhibitor of calpain-1, BDA-410 (30 mg/kg/d), on RBCs from SAD mice, a mouse model for SCD. In SAD mice, BDA-410 improved RBC morphology, reduced RBC density (D20; from 1106±0.001 to 1100±0.001 g/ml; P<0.05) and increased RBC-K+ content (from 364±10 to 429±12.3 mmol/kg Hb; P<0.05), markedly reduced the activity of the Ca2+-activated K+channel (Gardos channel), and decreased membrane association of peroxiredoxin-2. The inhibitory effect of calphostin C, a specific inhibitor of protein kinase C (PKC), on the Gardos channel was eliminated after BDA-410 treatment, which suggests that calpain-1 inhibition affects the PKC-dependent fraction of the Gardos channel. BDA-410 prevented hypoxia-induced RBC dehydration and K+ loss in SAD mice. These data suggest a potential role of BDA-410 as a novel therapeutic agent for treatment of SCD.—De Franceschi, L., Franco, R. S., Bertoldi, M., Brugnara, C., Matté, A., Siciliano, A., Wieschhaus, A. J., Chishti, A. H., Joiner, C. H. Pharmacological inhibition of calpain-1 prevents red cell dehydration and reduces Gardos channel activity in a mouse model of sickle cell disease. PMID:23085996

  7. Jobelyn® exhibited anti-inflammatory, antioxidant, and membrane-stabilizing activities in experimental models.

    PubMed

    Umukoro, Solomon; Oluwole, Oluwafemi Gabriel; Eduviere, Anthony T; Adrian, Omogbiya Itievere; Ajayi, Abayomi M

    2015-09-01

    Jobelyn® (JB) is an African sorghum-based food supplement claimed to be efficacious for the treatment of rheumatoid arthritis (RA). Although in vitro studies confirmed its anti-inflammatory property, no study had shown the effect of JB using in vivo animal models of inflammation. Thus, its effects on acute and chronic inflammation in rats were evaluated in this study. Its effect on rat red blood cell (RBC) lysis was also assessed. Acute inflammation was induced with intraplanter injection of carrageenan and increase in rat paw volume was measured using plethysmometer. The volume of fluid exudates, number of leukocytes, concentrations of malondialdehyde (MDA), and glutathione (GSH) in the fluid were measured on day 5 after induction of chronic inflammation with carrageenan in the granuloma air pouch model. RBC lysis induced by hypotonic medium as determined by release of hemoglobin was measured spectrophotometerically. JB (50-200 mg/kg) given orally produced a significant inhibition of acute inflammation induced by carrageenan in rats. It reduced the volume and number of leukocytes in inflammatory fluid in the granuloma air pouch model of chronic inflammation. It further decreased the levels of MDA in the fluid suggesting antioxidant property. JB elevated the concentrations of GSH in inflammatory exudates indicating free radical scavenging activity. It also significantly inhibited RBC lysis caused by hypotonic medium, suggesting membrane-stabilizing property. JB has in vivo anti-inflammatory activity, which may be related to its antioxidant and membrane-stabilizing properties, supporting its use for the treatment of arthritic disorder.

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

    NASA Technical Reports Server (NTRS)

    Eisman, G. A.

    1989-01-01

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

  9. Chapter 6: cubic membranes the missing dimension of cell membrane organization.

    PubMed

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

    2009-01-01

    Biological membranes are among the most fascinating assemblies of biomolecules: a bilayer less than 10 nm thick, composed of rather small lipid molecules that are held together simply by noncovalent forces, defines the cell and discriminates between "inside" and "outside", survival, and death. Intracellular compartmentalization-governed by biomembranes as well-is a characteristic feature of eukaryotic cells, which allows them to fulfill multiple and highly specialized anabolic and catabolic functions in strictly controlled environments. Although cellular membranes are generally visualized as flat sheets or closely folded isolated objects, multiple observations also demonstrate that membranes may fold into "unusual", highly organized structures with 2D or 3D periodicity. The obvious correlation of highly convoluted membrane organizations with pathological cellular states, for example, as a consequence of viral infection, deserves close consideration. However, knowledge about formation and function of these highly organized 3D periodic membrane structures is scarce, primarily due to the lack of appropriate techniques for their analysis in vivo. Currently, the only direct way to characterize cellular membrane architecture is by transmission electron microscopy (TEM). However, deciphering the spatial architecture solely based on two-dimensionally projected TEM images is a challenging task and prone to artifacts. In this review, we will provide an update on the current progress in identifying and analyzing 3D membrane architectures in biological systems, with a special focus on membranes with cubic symmetry, and their potential role in physiological and pathophysiological conditions. Proteomics and lipidomics approaches in defined experimental cell systems may prove instrumental to understand formation and function of 3D membrane morphologies.

  10. Tympanic membrane organ culture using cell culture well inserts engrafted with tympanic membrane tissue explants.

    PubMed

    Liew, Lawrence J; Day, Richard M; Dilley, Rodney J

    2017-03-01

    Tissue engineering approaches using growth factors and various materials for repairing chronic perforations of the tympanic membrane are being developed, but there are surprisingly few relevant tissue culture models available to test new treatments. Here, we present a simple three-dimensional model system based on micro-dissecting the rat tympanic membrane umbo and grafting it into the membrane of a cell culture well insert. Cell outgrowth from the graft produced sufficient cells to populate a membrane of similar surface area to the human tympanic membrane within 2 weeks. Tissue grafts from the annulus region also showed cell outgrowth but were not as productive. The umbo organoid supported substantial cell proliferation and migration under the influence of keratinocyte growth medium. Cells from umbo grafts were enzymatically harvested from the polyethylene terephthalate (PET) membrane for expansion in routine culture and cells could be harvested consecutively from the same graft over multiple cycles. We used harvested cells to test cell migration properties and to engraft a porous silk scaffold material as proof-of-principle for tissue engineering applications. This model is simple enough to be widely adopted for tympanic membrane regeneration studies and has promise as a tissue-equivalent model alternative to animal testing.

  11. Evidence of femtosecond-laser pulse induced cell membrane nanosurgery

    NASA Astrophysics Data System (ADS)

    Katchinskiy, Nir; Godbout, Roseline; Elezzabi, Abdulhakem Y.

    2017-02-01

    The mechanism of femtosecond laser nanosurgical attachment is investigated in the following article. Using sub-10 femtosecond laser pulses with 800 nm central wavelength were used to attach retinoblastoma cells. During the attachment process the cell membrane phospholipid bilayers hemifuse into one shared phospholipid bilayer, at the location of attachment. Transmission electron microscopy was used in order to verify the above hypothesis. Based on the imaging results, it was concluded that the two cell membrane coalesce to form one single shared membrane. The technique of cell-cell attachment via femtosecond laser pulses could potentially serve as a platform for precise cell membrane manipulation. Manipulation of the cellular membrane is valuable for studying diseases such as cancer; where the expression level of plasma proteins on the cell membrane is altered.

  12. Polyarylenethioethersulfone Membranes for Fuel Cells (Postprint)

    DTIC Science & Technology

    2010-01-01

    The Electrochemical SocietyProton exchange membrane fuel cells PEMFCs are an attrac- tive power source due to their energy efficiency and...standard in PEMFC technology.3,4 Nafion membranes have a polytetrafluoro- ethylene PTFE backbone, which provides thermal and chemical stability, and...diffusion layers to fabricate MEAs. Single-cell test (H- PEMFC ).— MEAs were positioned in a single-cell fixture with graphite blocks as current

  13. Human and great ape red blood cells differ in plasmalogen levels and composition

    PubMed Central

    2011-01-01

    Background Plasmalogens are ether phospholipids required for normal mammalian developmental, physiological, and cognitive functions. They have been proposed to act as membrane antioxidants and reservoirs of polyunsaturated fatty acids as well as influence intracellular signaling and membrane dynamics. Plasmalogens are particularly enriched in cells and tissues of the human nervous, immune, and cardiovascular systems. Humans with severely reduced plasmalogen levels have reduced life spans, abnormal neurological development, skeletal dysplasia, impaired respiration, and cataracts. Plasmalogen deficiency is also found in the brain tissue of individuals with Alzheimer disease. Results In a human and great ape cohort, we measured the red blood cell (RBC) levels of the most abundant types of plasmalogens. Total RBC plasmalogen levels were lower in humans than bonobos, chimpanzees, and gorillas, but higher than orangutans. There were especially pronounced cross-species differences in the levels of plasmalogens with a C16:0 moiety at the sn-1 position. Humans on Western or vegan diets had comparable total RBC plasmalogen levels, but the latter group showed moderately higher levels of plasmalogens with a C18:1 moiety at the sn-1 position. We did not find robust sex-specific differences in human or chimpanzee RBC plasmalogen levels or composition. Furthermore, human and great ape skin fibroblasts showed only modest differences in peroxisomal plasmalogen biosynthetic activity. Human and chimpanzee microarray data indicated that genes involved in plasmalogen biosynthesis show cross-species differential expression in multiple tissues. Conclusion We propose that the observed differences in human and great ape RBC plasmalogens are primarily caused by their rates of biosynthesis and/or turnover. Gene expression data raise the possibility that other human and great ape cells and tissues differ in plasmalogen levels. Based on the phenotypes of humans and rodents with plasmalogen

  14. 7SL RNA in Vertebrate Red Blood Cells.

    PubMed

    Talhouarne, Gaëlle J S; Gall, Joseph G

    2018-04-23

    We report that 7SL, the RNA component of the signal recognition particle (SRP), is an abundant ncRNA in mature red blood cells (RBCs) of human, mouse, and the frog Xenopus. 7SL RNA in RBCs is not associated with the canonical proteins of the SRP. Instead, it co-immunoprecipitates from a lysate of RBCs with a number of membrane-binding proteins. Human and mouse RBCs also contain a previously undescribed 68 nt RNA, sRN7SL, derived from the "S domain" of 7SL RNA. We discuss the possibility that 7SL RNA is selectively protected from nucleases by association with the RBC membrane. Because 7SL is not associated with the canonical proteins of the SRP, it could represent a non-functional remnant of the protein synthetic machinery. Alternatively, it could play a new, as yet undefined role in RBC metabolism. Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  15. Red blood cells carry out T cell growth and survival bioactivities that are sensitive to cyclosporine A.

    PubMed

    Antunes, Ricardo F; Brandão, Cláudia; Carvalho, Gonçalo; Girão, Cristina; Arosa, Fernando A

    2009-10-01

    Red blood cells (RBC) have emerged as a novel regulatory cell type endowed with bioactivities toward activated human T cells. Herein we show that the RBC bioactivities act on intracellular pathways initiated by T cell receptor (TCR)-dependent and -independent stimuli,including IL-2, IL-15, and the mixture of phorbol dibutyrate and ionomycin. The RBC bioactivities preserve the antioxidant status and are capable of rescuing activated T cells from cell death induced by serum deprivation. They are not mediated by glycosylphosphatidylinositol-linked receptors or sialic acids, and kinetic studies revealed that they hasten the entrance into the cell cycle. By using cyclosporine A (CsA) and rapamycin (Rapa) we show that the RBC bioactivities are calcineurin-dependent. Thus, treatment of T cells with CsA, but not Rapa, impaired RBC bioactivities, and preincubation of RBC with CsA completely abolished their bioactivities. We have demonstrated that RBC carry out bioactivities that are sensitive to CsA.

  16. Fuel cell membrane humidification

    DOEpatents

    Wilson, Mahlon S.

    1999-01-01

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

  17. Does the extracorporeal circulation worsen anemia in hemodialysis patients? Investigation with advanced microscopes of red blood cells drawn at the beginning and end of dialysis.

    PubMed

    Stamopoulos, Dimosthenis; Bakirtzi, Nerantzoula; Manios, Efthymios; Grapsa, Eirini

    2013-01-01

    In hemodialysis (HD) patients, anemia relates to three main factors: insufficient production of erythropoietin; impaired management of iron; and decreased lifespan of red blood cells (RBCs). The third factor can relate to structural deterioration of RBCs due to extrinsic (extracorporeal circuit; biochemical activation and/or mechanical stress during dialysis) and intrinsic (uremic milieu; biochemical interference of the RBC membrane constituents with toxins) mechanisms. Herein, we evaluate information accessed with advanced imaging techniques at the cellular level. Atomic force and scanning electron microscopes were employed to survey intact RBCs (iRBCs) of seven HD patients in comparison to seven healthy donors. The extrinsic factor was investigated by contrasting pre- and post-HD samples. The intrinsic environment was investigated by comparing the microscopy data with the clinical ones. The iRBC membranes of the enrolled HD patients were overpopulated with orifice-like (high incidence; typical size within 100-1,000 nm) and crevice-like (low incidence; typical size within 500-4,000 nm) defects that exhibited a statistically significant (P < 0.05) relative increase (+55% and +350%, respectively) in respect to healthy donors. The relative variation of the orifice and crevice indices (mean population of orifices and crevices per top membrane surface) between pre- and post-HD was not statistically significant (-3.3% and +4.5%, respectively). The orifice index correlates with the concentrations of urea, calcium, and phosphorus, but not, however, with that of creatinine. Extracorporeal circulation is not detrimental to the structural integrity of RBC membranes. Uremic milieu is a candidate cause of RBC membrane deterioration, which possibly worsens anemia.

  18. Fetal Membranes-Derived Stem Cells Microenvironment.

    PubMed

    Favaron, Phelipe Oliveira; Miglino, Maria Angelica

    2017-01-01

    Recently, the regenerative medicine has been trying to congregate different areas such as tissue engineering and cellular therapy, in order to offer effective treatments to overcome several human and veterinary medical problems. In this regard, fetal membranes have been proposed as a powerful source for obtainment of multipotent stem cells with low immunogenicity, anti-inflammatory properties and nontumorigenicity properties for the treatment of several diseases, including replacing cells lost due to tissue injuries or degenerative diseases. Morpho-physiological data have shown that fetal membranes, especially the yolk sac and amnion play different functions according to the gestational period, which are direct related to the features of the microenvironment that their cells are subject. The characteristics of the microenvironment affect or controls important cellular events involved with proliferation, division and maintenance of the undifferentiated stage or differentiation, especially acting on the extracellular matrix components. Considering the importance of the microenvironment and the diversity of embryonic and fetal membrane-derived stem cells, this chapter will addressed advances in the isolation, phenotyping, characteristics of the microenvironment, and applications of yolk sac and amniotic membrane-derived stem cells for human and veterinary regenerative medicine.

  19. Cell Membrane-Cloaked Nanoparticles for Targeted Therapeutics

    NASA Astrophysics Data System (ADS)

    Luk, Brian Tsengchi

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

  20. Cooperative tumour cell membrane targeted phototherapy

    NASA Astrophysics Data System (ADS)

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

    2017-06-01

    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.

  1. Polyarylenethioethersulfone Membranes for Fuel Cells (Postprint)

    DTIC Science & Technology

    2007-09-01

    2007. 0013-4651/2007/1549/B960/9/$20.00 © The Electrochemical SocietyProton exchange membrane fuel cells PEMFCs are an attrac- tive power source...DuPont have become the commercially available standard in PEMFC technology.3,4 Nafion membranes have a polytetrafluoro- ethylene PTFE backbone, which...Inc. were used as anode and cathode gas diffusion layers to fabricate MEAs. Single-cell test (H- PEMFC ).— MEAs were positioned in a single-cell fixture

  2. Chemical camouflage of antigenic determinants: stealth erythrocytes.

    PubMed

    Scott, M D; Murad, K L; Koumpouras, F; Talbot, M; Eaton, J W

    1997-07-08

    In a number of clinical circumstances it would be desirable to artificially conceal cellular antigenic determinants to permit survival of heterologous donor cells. A case in point is the problem encountered in transfusions of patients with rare blood types or chronically transfused patients who become allosensitized to minor blood group determinants. We have tested the possibility that chemical modification of the red blood cell (RBC) membrane might serve to occlude antigenic determinants, thereby minimizing transfusion reactions. To this end, we have covalently bound methoxy(polyethylene glycol) (mPEG) to the surface of mammalian RBC via cyanuric chloride coupling. Human RBC treated with this technique lose ABO blood group reactivity as assessed by solution-phase antisera agglutination. In accord with this, we also find a profound decrease in anti-blood group antibody binding. Furthermore, whereas human monocytes avidly phagocytose untreated sheep RBC, mPEG-derivatized sheep RBC are ineffectively phagocytosed. Surprisingly, human and mouse RBC appear unaffected by this covalent modification of the cell membrane. Thus, mPEG-treated RBC are morphologically normal, have normal osmotic fragility, and mPEG-derivatized murine RBC have normal in vivo survival, even following repeated infusions. Finally, in preliminary experiments, mPEG-modified sheep RBC intraperitoneally transfused into mice show significantly improved (up to 360-fold) survival when compared with untreated sheep RBC. We speculate that similar chemical camouflage of intact cells may have significant clinical applications in both transfusion (e.g., allosensitization and autoimmune hemolytic disease) and transplantation (e.g., endothelial cells and pancreatic beta cells) medicine.

  3. Femtosecond-laser setups for cell-membrane poration

    NASA Astrophysics Data System (ADS)

    Breunig, Hans Georg; Batista, Ana; Sauer, Benjamin; König, Aisada; König, Karsten

    2018-02-01

    Focused femtosecond-laser pulses can create tiny transient holes in cell membranes which temporarily allows foreign genetic material from the outside to reach the cell interior. With suitable laser parameters this all optical "optoporation" allows highly efficient laser-assisted cell transfection by reprogramming the celĺs genetic code with very high cell survival rates. Furthermore, the use of viruses or nanoparticle as carriers which may cause serious side effects can be completely omitted. However, the cell positions need to be precisely determined to allow individual focusing of the laser radiation onto the cell membranes which is for large cell numbers quite elaborate and time consuming. We addressed these issues and present optical microscope add-ons for fast and almost hands-off laserassisted poration of cell membranes with automated determination of the positions of adherent cells in a culture dish or targeting continuously flowing cells in suspension.

  4. Downregulation of Rubisco Activity by Non-enzymatic Acetylation of RbcL.

    PubMed

    Gao, Xiang; Hong, Hui; Li, Wei-Chao; Yang, Lili; Huang, Jirong; Xiao, You-Li; Chen, Xiao-Ya; Chen, Gen-Yun

    2016-07-06

    Atmospheric carbon dioxide (CO2) is assimilated by the most abundant but sluggish enzyme, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco). Here we show that acetylation of lysine residues of the Rubisco large subunit (RbcL), including Lys201 and Lys334 in the active sites, may be an important mechanism in the regulation of Rubisco activities. It is well known that Lys201 reacts with CO2 for carbamylation, a prerequisite for both carboxylase and oxygenase activities of Rubisco, and Lys334 contacts with ribulose-1,5-bisphosphate (RuBP). The acetylation level of RbcL in plants is lower during the day and higher at night, inversely correlating with the Rubisco carboxylation activity. A search of the chloroplast proteome database did not reveal a canonical acetyltransferase; instead, we found that a plant-derived metabolite, 7-acetoxy-4-methylcoumarin (AMC), can non-enzymatically acetylate both native Rubisco and synthesized RbcL peptides spanning Lys334 or Lys201. Furthermore, lysine residues were modified by synthesized 4-methylumbelliferone esters with different electro- and stereo-substitutes, resulting in varied Rubisco activities. 1-Chloroethyl 4-methylcoumarin-7-yl carbonate (ClMC) could transfer the chloroethyl carbamate group to lysine residues of RbcL and completely inactivate Rubisco, whereas bis(4-methylcoumarin-7-yl) carbonate (BMC) improved Rubisco activity through increasing the level of Lys201 carbamylation. Our findings indicate that RbcL acetylation negatively regulates Rubisco activity, and metabolic derivatives can be designed to dissect and improve CO2 fixation efficiency of plants through lysine modification. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

  5. Characterization of Storage-Induced Red Blood Cell Hemolysis Using Raman Spectroscopy.

    PubMed

    Gautam, Rekha; Oh, Joo-Yeun; Marques, Marisa B; Dluhy, Richard A; Patel, Rakesh P

    2018-06-11

    The therapeutic efficacy and safety of stored red blood cells (RBCs) relies on minimal in-bag hemolysis. The accuracy of current methods of measuring hemolysis can suffer as a result of specimen collection and processing artefacts. To test whether Raman spectroscopy could be used to assess hemolysis. RBCs were stored for as long as 42 days. Raman spectra of RBCs were measured before and after washing, and hemolysis was measured in supernatant by visible spectroscopy. Raman spectra indicated increased concentrations of oxyhemoglobin (oxyHb) and methemoglobin (metHb), and decreased membrane fluidity with storage age. Changes in oxyHb and metHb were associated with the intraerythrocytic and extracellular fractions, respectively. Hemolysis increased in a storage age-dependent manner. Changes in Raman bands reflective of oxyHb, metHb, and RBC membranes correlated with hemolysis; the most statistically significant change was an increased intensity of metHb and decreased membrane fluidity. These data suggest that Raman spectroscopy may offer a new label-free modality to assess RBC hemolysis during cold storage.

  6. New High-Temperature Membranes Developed for Proton Exchange Membrane Fuel Cells

    NASA Technical Reports Server (NTRS)

    Kinder, James D.

    2004-01-01

    Fuel cells are receiving a considerable amount of attention for potential use in a variety of areas, including the automotive industry, commercial power generation, and personal electronics. Research at the NASA Glenn Research Center has focused on the development of fuel cells for use in aerospace power systems for aircraft, unmanned air vehicles, and space transportation systems. These applications require fuel cells with higher power densities and better durability than what is required for nonaerospace uses. In addition, membrane cost is a concern for any fuel cell application. The most widely used membrane materials for proton exchange membrane (PEM) fuel cells are based on sulfonated perfluorinated polyethers, typically Nafion 117, Flemion, or Aciplex. However, these polymers are costly and do not function well at temperatures above 80 C. At higher temperatures, conventional membrane materials dry out and lose their ability to conduct protons, essential for the operation of the fuel cell. Increasing the operating temperature of PEM fuel cells from 80 to 120 C would significantly increase their power densities and enhance their durability by reducing the susceptibility of the electrode catalysts to carbon monoxide poisoning. Glenn's Polymers Branch has focused on developing new, low-cost membranes that can operate at these higher temperatures. A new series of organically modified siloxane (ORMOSIL) polymers were synthesized for use as membrane materials in a high-temperature PEM fuel cell. These polymers have an organic portion that can allow protons to transport through the polymer film and a cross-linked silica network that gives the polymers dimensional stability. These flexible xerogel polymer films are thermally stable, with decomposition onset as high as 380 C. Two types of proton-conducting ORMOSIL films have been produced: (1) NASA-A, which can coordinate many highly acid inorganic salts that facilitate proton conduction and (2) NASA-B, which has been

  7. Exacerbation of oxidative stress during sickle vaso-occlusive crisis is associated with decreased anti-band 3 autoantibodies rate and increased red blood cell-derived microparticle level: a prospective study.

    PubMed

    Hierso, Régine; Lemonne, Nathalie; Villaescusa, Rinaldo; Lalanne-Mistrih, Marie-Laure; Charlot, Keyne; Etienne-Julan, Maryse; Tressières, Benoit; Lamarre, Yann; Tarer, Vanessa; Garnier, Yohann; Hernandez, Ada Arce; Ferracci, Serge; Connes, Philippe; Romana, Marc; Hardy-Dessources, Marie-Dominique

    2017-03-01

    Painful vaso-occlusive crisis, a hallmark of sickle cell anaemia, results from complex, incompletely understood mechanisms. Red blood cell (RBC) damage caused by continuous endogenous and exogenous oxidative stress may precipitate the occurrence of vaso-occlusive crises. In order to gain insight into the relevance of oxidative stress in vaso-occlusive crisis occurrence, we prospectively compared the expression levels of various oxidative markers in 32 adults with sickle cell anaemia during vaso-occlusive crisis and steady-state conditions. Compared to steady-state condition, plasma levels of free haem, advanced oxidation protein products and myeloperoxidase, RBC caspase-3 activity, as well as the concentrations of total, neutrophil- and RBC-derived microparticles were increased during vaso-occlusive crises, whereas the reduced glutathione content was decreased in RBCs. In addition, natural anti-band 3 autoantibodies levels decreased during crisis and were negatively correlated with the rise in plasma advanced oxidation protein products and RBC caspase-3 activity. These data showed an exacerbation of the oxidative stress during vaso-occlusive crises in sickle cell anaemia patients and strongly suggest that the higher concentration of harmful circulating RBC-derived microparticles and the reduced anti-band 3 autoantibodies levels may be both related to the recruitment of oxidized band 3 into membrane aggregates. © 2016 John Wiley & Sons Ltd.

  8. Stretching Micropatterned Cells on a PDMS Membrane

    PubMed Central

    Carpi, Nicolas; Piel, Matthieu

    2014-01-01

    Mechanical forces exerted on cells and/or tissues play a major role in numerous processes. We have developed a device to stretch cells plated on a PolyDiMethylSiloxane (PDMS) membrane, compatible with imaging. This technique is reproducible and versatile. The PDMS membrane can be micropatterned in order to confine cells or tissues to a specific geometry. The first step is to print micropatterns onto the PDMS membrane with a deep UV technique. The PDMS membrane is then mounted on a mechanical stretcher. A chamber is bound on top of the membrane with biocompatible grease to allow gliding during the stretch. The cells are seeded and allowed to spread for several hours on the micropatterns. The sample can be stretched and unstretched multiple times with the use of a micrometric screw. It takes less than a minute to apply the stretch to its full extent (around 30%). The technique presented here does not include a motorized device, which is necessary for applying repeated stretch cycles quickly and/or computer controlled stretching, but this can be implemented. Stretching of cells or tissue can be of interest for questions related to cell forces, cell response to mechanical stress or tissue morphogenesis. This video presentation will show how to avoid typical problems that might arise when doing this type of seemingly simple experiment. PMID:24514571

  9. Fuel cell ion-exchange membrane investigation

    NASA Technical Reports Server (NTRS)

    Toy, M. S.

    1972-01-01

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

  10. Dynamic pattern generation in cell membranes: Current insights into membrane organization.

    PubMed

    Raghunathan, Krishnan; Kenworthy, Anne K

    2018-05-09

    It has been two decades since the lipid raft hypothesis was first presented. Even today, whether these nanoscale cholesterol-rich domains are present in cell membranes is not completely resolved. However, especially in the last few years, a rich body of literature has demonstrated both the presence and the importance of non-random distribution of biomolecules on the membrane, which is the focus of this review. These new developments have pushed the experimental limits of detection and have brought us closer to observing lipid domains in the plasma membrane of live cells. Characterization of biomolecules associated with lipid rafts has revealed a deep connection between biological regulation and function and membrane compositional heterogeneities. Finally, tantalizing new developments in the field have demonstrated that lipid domains might not just be associated with the plasma membrane of eukaryotes but could potentially be a ubiquitous membrane-organizing principle in several other biological systems. This article is part of a Special Issue entitled: Emergence of Complex Behavior in Biomembranes edited by Marjorie Longo. Copyright © 2018. Published by Elsevier B.V.

  11. Convection due to an unstable density difference across a permeable membrane

    NASA Astrophysics Data System (ADS)

    Puthenveettil, Baburaj A.; Arakeri, Jaywant H.

    We study natural convection driven by unstable concentration differences of sodium chloride (NaCl) across a horizontal permeable membrane at Rayleigh numbers (Ra) of 1010 to 1011 and Schmidt number (Sc)=600. A layer of brine lies over a layer of distilled water, separated by the membrane, in square-cross-section tanks. The membrane is permeable enough to allow a small flow across it at higher driving potentials. Based on the predominant mode of transport across the membrane, three regimes of convection, namely an advection regime, a diffusion regime and a combined regime, are identified. The near-membrane flow in all the regimes consists of sheet plumes formed from the unstable layers of fluid near the membrane. In the advection regime observed at higher concentration differences (Bb) show a common log-normal probability density function at all Ra. We propose a phenomenology which predicts /line{lambda}_b sqrt{Z_w Z_{V_i}}, where Zw and Z_{V_i} are, respectively, the near-wall length scales in Rayleighnard convection (RBC) and due to the advection velocity. In the combined regime, which occurs at intermediate values of C/2)4/3. At lower driving potentials, in the diffusion regime, the flux scaling is similar to that in turbulent RBC.

  12. Interaction of Defensins with Model Cell Membranes

    NASA Astrophysics Data System (ADS)

    Sanders, Lori K.; Schmidt, Nathan W.; Yang, Lihua; Mishra, Abhijit; Gordon, Vernita D.; Selsted, Michael E.; Wong, Gerard C. L.

    2009-03-01

    Antimicrobial peptides (AMPs) comprise a key component of innate immunity for a wide range of multicellular organisms. For many AMPs, activity comes from their ability to selectively disrupt and lyse bacterial cell membranes. There are a number of proposed models for this action, but the detailed molecular mechanism of selective membrane permeation remains unclear. Theta defensins are circularized peptides with a high degree of selectivity. We investigate the interaction of model bacterial and eukaryotic cell membranes with theta defensins RTD-1, BTD-7, and compare them to protegrin PG-1, a prototypical AMP, using synchrotron small angle x-ray scattering (SAXS). The relationship between membrane composition and peptide induced changes in membrane curvature and topology is examined. By comparing the membrane phase behavior induced by these different peptides we will discuss the importance of amino acid composition and placement on membrane rearrangement.

  13. Application of Controlled Shear Stresses on the Erythrocyte Membrane as a New Approach to Promote Molecule Encapsulation.

    PubMed

    Casagrande, Giustina; Arienti, Flavio; Mazzocchi, Arabella; Taverna, Francesca; Ravagnani, Fernando; Costantino, MariaLaura

    2016-10-01

    Human red blood cells (RBCs) have a remarkable capacity to undergo reversible membrane swelling. Resealed erythrocytes have been proposed as carriers and bioreactors to be used in the treatment of various diseases. This work is aimed at developing a setup allowing the encapsulation of test molecules into erythrocytes by inducing reversible pore formation on the RBC membrane through the application of controlled mechanical shear stresses. The designed setup consists of two reservoirs connected by a glass capillary. Each reservoir is connected to a compressor; during the tests, the reservoirs were in turn pressurized to promote erythrocyte flow through the capillary. The setup was filled with a suspension of erythrocytes, phosphate buffer, and FITC-dextran. Dextran was chosen as the diffusive molecule to check membrane pore dimensions. Samples of the suspension were withdrawn at scheduled times while the setup was operating. Flow cytometry and stereo-optical microscopy analyses were used to evaluate the erythrocyte dextran uptake. The setup was shown to be safe, well controlled, and adjustable. The outcomes of the experimental tests showed significant dextran uptake by RBCs up to 8%. Microscopy observations highlighted the formation of echinocytes in the analyzed samples. Erythrocytes from different donors showed different reactions to mechanical stresses. The experimental outcomes proved the possibility to encapsulate test molecules into erythrocytes by applying controlled mechanical shear stresses on the RBC membrane, encouraging further studies. Copyright © 2016 International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.

  14. DHA and EPA in red blood cell membranes are associated with dietary intakes of omega-3-rich fish in healthy children.

    PubMed

    Parks, Colleen A; Brett, Neil R; Agellon, Sherry; Lavery, Paula; Vanstone, Catherine A; Maguire, Jonathon L; Rauch, Frank; Weiler, Hope A

    2017-09-01

    Omega-3 long-chain polyunsaturated fatty acids (n-3 LCPUFA) are important in child development. The primary objective of this study was to investigate the associations between dietary intakes of n-3 LCPUFA and red blood cell (RBC) n-3 LCPUFA in young children. Healthy children, (2-8y) underwent RBC fatty acid profiling. Dietary intakes were parent-reported over 6 mo using three 24h dietary intake assessments and three 30 d food frequency questionnaires (FFQ). Participants (n = 49, 5.6 ± 1.9y), were 59% male, and had a body mass index (BMI) z-score of 0.65 ± 0.84. Dietary n-3 LCPUFA intakes were not different over time. RBC docosahexaenoic acid (DHA) positively correlated with average DHA from the 24h recalls. RBC DHA and eicosapentaenoic acid (EPA) positively correlated with average n-3 LCPUFA-rich fish intake from the FFQ. RBC appear to reflect long-term stable intakes of n-3 LCPUFA during growth in healthy young children. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Neuroglian-mediated cell adhesion induces assembly of the membrane skeleton at cell contact sites.

    PubMed

    Dubreuil, R R; MacVicar, G; Dissanayake, S; Liu, C; Homer, D; Hortsch, M

    1996-05-01

    The protein ankyrin links integral membrane proteins to the spectrin-based membrane skeleton. Ankyrin is often concentrated within restricted membrane domains of polarized epithelia and neurons, but the mechanisms responsible for membrane targeting and its segregation within a continuous lipid bilayer remain unexplained. We provide evidence that neuroglian, a cell adhesion molecule related to L1 and neurofascin, can transmit positional information directly to ankyrin and thereby polarize its distribution in Drosophila S2 tissue culture cells. Ankyrin was not normally associated with the plasma membrane of these cells. Upon expression of an inducible neuroglian minigene, however, cells aggregated into large clusters and ankyrin became concentrated at sites of cell-cell contact. Spectrin was also recruited to sites of cell contact in response to neuroglian expression. The accumulation of ankyrin at cell contacts required the presence of the cytoplasmic domain of neuroglian since a glycosyl phosphatidylinositol-linked form of neuroglian failed to recruit ankyrin to sites of cell-cell contact. Double-labeling experiments revealed that, whereas ankyrin was strictly associated with sites of cell-cell contact, neuroglian was more broadly distributed over the cell surface. A direct interaction between neuroglian and ankyrin was demonstrated using yeast two-hybrid analysis. Thus, neuroglian appears to be activated by extracellular adhesion so that ankyrin and the membrane skeleton selectively associate with sites of cell contact and not with other regions of the plasma membrane.

  16. Membrane stress increases cation permeability in red cells.

    PubMed

    Johnson, R M

    1994-11-01

    The human red cell is known to increase its cation permeability when deformed by mechanical forces. Light-scattering measurements were used to quantitate the cell deformation, as ellipticity under shear. Permeability to sodium and potassium was not proportional to the cell deformation. An ellipticity of 0.75 was required to increase the permeability of the membrane to cations, and flux thereafter increased rapidly as the limits of cell extension were reached. Induction of membrane curvature by chemical agents also did not increase cation permeability. These results indicate that membrane deformation per se does not increase permeability, and that membrane tension is the effector for increased cation permeability. This may be relevant to some cation permeabilities observed by patch clamping.

  17. Fuel-Cell Structure Prevents Membrane Drying

    NASA Technical Reports Server (NTRS)

    Mcelroy, J.

    1986-01-01

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

  18. Autologus or allogenic uses of umbilical cord blood whole or RBC transfusion - a review.

    PubMed

    Chakrabarty, P; Rudra, S

    2013-01-01

    Once Umbilical Cord with Placenta considered a biological waste product and generally discarded after delivery but now cord blood has emerged as a viable source of hematopoietic stem cell transplantation. High-risk premature infants require red cell transfusions for anemia. A unique property of cord blood (CB) for its high content of immature hematopoietic progenitor cells (HPCs). Placental blood for autologous transfusions can be collected with aseptic precaution/sterilely into citrate-phosphate-dextrose and stored at 4°C. During storage for 8 days, the placental red cell content of adenosine triphosphate remained normal. The 2,3,-diphosphoglycerate concentration of cells stored beyond 8 days declined sharply. So we have to store umbilical cord blood (UCB) within 7 days for its best result. During storage, placental blood underwent an exchange of extra-cellular Na+ and K+, but no change in glutathione content. Hemolysis was less than 1 percent. Bacteriologic and fungal cultures remained sterile. These suggest that human placental blood can be collected safely and preserved effectively for autologous/allogenic transfusion therapy. In neonatal transfusion practice, efforts have been made to provide premature infants with autologous red blood cell (RBC), especially those born before 32 gestational weeks. In India no adverse transfusion effects were seen in a wide variety of patients that received (pooled) allogeneic fresh whole blood / UCB transfusions. The use of UCB for small volume allogeneic transfusions in anaemic children in Africa or in malaria endemic areas has also been proposed. A preclinical study showed that donation and transfusion of UCB would be acceptable to women living in Mombasa, Kenya. In view of the small volumes RBC per unit that can be collected, it is most likely that anaemic children need of a small volume of transfusions. In resource-restricted countries would benefit most from this easily available transfusion product.

  19. Application of image flow cytometry for the characterization of red blood cell morphology

    NASA Astrophysics Data System (ADS)

    Pinto, Ruben N.; Sebastian, Joseph A.; Parsons, Michael; Chang, Tim C.; Acker, Jason P.; Kolios, Michael C.

    2017-02-01

    Red blood cells (RBCs) stored in hypothermic environments for the purpose of transfusion have been documented to undergo structural and functional changes over time. One sign of the so-called RBC storage lesion is irreversible damage to the cell membrane. Consequently, RBCs undergo a morphological transformation from regular, deformable biconcave discocytes to rigid spheroechinocytes. The spherically shaped RBCs lack the deformability to efficiently enter microvasculature, thereby reducing the capacity of RBCs to oxygenate tissue. Blood banks currently rely on microscope techniques that include fixing, staining and cell counting in order to morphologically characterize RBC samples; these methods are labor intensive and highly subjective. This study presents a novel, high-throughput RBC morphology characterization technique using image flow cytometry (IFC). An image segmentation template was developed to process 100,000 images acquired from the IFC system and output the relative spheroechinocyte percentage. The technique was applied on samples extracted from two blood bags to monitor the morphological changes of the RBCs during in vitro hypothermic storage. The study found that, for a given sample of RBCs, the IFC method was twice as fast in data acquisition, and analyzed 250-350 times more RBCs than the conventional method. Over the lifespan of the blood bags, the mean spheroechinocyte population increased by 37%. Future work will focus on expanding the template to segregate RBC images into more subpopulations for the validation of the IFC method against conventional techniques; the expanded template will aid in establishing quantitative links between spheroechinocyte increase and other RBC storage lesion characteristics.

  20. The changes of red blood cell viscoelasticity and sports anemia in male 24-hr ultra-marathoners.

    PubMed

    Liu, Che-Hung; Tseng, Yen-Fang; Lai, Jiun-I; Chen, Yin-Quan; Wang, Shih-Hao; Kao, Wei-Fong; Li, Li-Hua; Chiu, Yu-Hui; How, Chorng-Kuang; Chang, Wen-Han

    2018-05-01

    In endurance sports, stress, dehydration and release of chemical factors have been associated with red blood cell (RBC) alterations of structure and function, which may contribute to sports anemia, a well-observed phenomenon during long-distance running. Until now, the investigation of the changes of viscoelastic properties of RBC membrane, a decisive factor of RBC deformability to avoid hemolysis, is lacking, especially in an Oriental population. nineteen runners were prospectively recruited into our study. Hematological parameters were analyzed before and immediately after the 2015 Taipei 24H Ultra-Marathon Festival, Taiwan. Video particle tracking microrheology was used to determine viscoelastic properties of each RBC sample by calculating the dynamic elastic modulus G'(f) and the viscous modulus G″(f) at frequency f = 20 Hz. Haptoglobin, RBC count, hemoglobin, hematocrit, mean cell hemoglobin, plasma free hemoglobin and unsaturated iron-binding capacity values of the recruited runners showed a statistically significant drop in the post-race values. Blood concentration of reticulocyte and ferritin were significantly higher at post-race compared with pre-race. 15 out of the 19 runners had a concurrent change in the elastic and the viscous moduli of their RBCs. Changes in the elastic and the viscous moduli were correlated with changes in the RBC count, hemoglobin and hematocrit. Viscoelasticity properties, the elastic modulus G'(f) and the viscous modulus G″(f) of RBCs are associated with endurance exercise-induced anemia. Copyright © 2017. Published by Elsevier Taiwan LLC.

  1. The time-course of red blood cell intracellular pH recovery following short-circuiting in relation to venous transit times in rainbow trout, Oncorhynchus mykiss.

    PubMed

    Harter, Till Sebastian; May, Alexandra G; Federspiel, William J; Supuran, Claudiu T; Brauner, Colin J

    2018-04-11

    Accumulating evidence is highlighting the importance of a system of enhanced hemoglobin-oxygen (Hb-O 2 ) unloading for cardiovascular O 2 transport in teleosts. Adrenergically stimulated sodium-proton-exchangers (β-NHE) create H + gradients across the red blood cell (RBC) membrane that are short-circuited in the presence of plasma-accessible carbonic anhydrase (paCA) at the tissues; the result is a large arterial-venous pH shift that greatly enhances O 2 unloading from pH-sensitive Hb. However, RBC intracellular pH (pH i ) must recover during venous transit (31-90 s), to enable O 2 -loading at the gills. The halftimes (t 1/2 ) and magnitudes of RBC β-adrenergic stimulation, short-circuiting with paCA and recovery of RBC pH i , were assessed in vitro, on rainbow trout whole blood, and using changes in closed-system PO 2 as a sensitive indicator for changes in RBC pH i . In addition, the recovery rate of RBC pH i was assessed in a continuous-flow apparatus that more closely mimics RBC transit though the circulation. Results indicate that: i) the t 1/2 of CA short-circuiting is likely within the residence time of blood in the capillaries; ii) the t 1/2 of RBC pH i recovery is 17 s and within the time of RBC venous transit; and iii) after short-circuiting RBCs re-establish the initial H + gradient across the membrane and can potentially undergo repeated cycles of short-circuiting and recovery. Thus, teleosts have evolved a system that greatly enhances O 2 unloading from pH-sensitive Hb at the tissues, while protecting O 2 loading at the gills; the resulting increase in O 2 transport per unit of blood flow may enable the tremendous athletic ability of salmonids.

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

    DOE Office of Scientific and Technical Information (OSTI.GOV)

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

    2015-12-31

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

  3. Lennard-Jones type pair-potential method for coarse-grained lipid bilayer membrane simulations in LAMMPS

    NASA Astrophysics Data System (ADS)

    Fu, S.-P.; Peng, Z.; Yuan, H.; Kfoury, R.; Young, Y.-N.

    2017-01-01

    Lipid bilayer membranes have been extensively studied by coarse-grained molecular dynamics simulations. Numerical efficiencies have been reported in the cases of aggressive coarse-graining, where several lipids are coarse-grained into a particle of size 4 ∼ 6 nm so that there is only one particle in the thickness direction. Yuan et al. proposed a pair-potential between these one-particle-thick coarse-grained lipid particles to capture the mechanical properties of a lipid bilayer membrane, such as gel-fluid-gas phase transitions of lipids, diffusion, and bending rigidity Yuan et al. (2010). In this work we implement such an interaction potential in LAMMPS to simulate large-scale lipid systems such as a giant unilamellar vesicle (GUV) and red blood cells (RBCs). We also consider the effect of cytoskeleton on the lipid membrane dynamics as a model for RBC dynamics, and incorporate coarse-grained water molecules to account for hydrodynamic interactions. The interaction between the coarse-grained water molecules (explicit solvent molecules) is modeled as a Lennard-Jones (L-J) potential. To demonstrate that the proposed methods do capture the observed dynamics of vesicles and RBCs, we focus on two sets of LAMMPS simulations: 1. Vesicle shape transitions with enclosed volume; 2. RBC shape transitions with different enclosed volume. Finally utilizing the parallel computing capability in LAMMPS, we provide some timing results for parallel coarse-grained simulations to illustrate that it is possible to use LAMMPS to simulate large-scale realistic complex biological membranes for more than 1 ms.

  4. Synthesis of Nanogels via Cell Membrane-Templated Polymerization.

    PubMed

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

    2015-09-09

    The synthesis of biomimetic hydrogel nanoparticles coated with a natural cell membrane is described. Compared to the existing strategy of wrapping cell membranes onto pre-formed nanoparticle substrates, this new approach forms the cell membrane-derived vesicles first, followed by growing nanoparticle cores in situ. It adds significant controllability over the nanoparticle properties and opens unique opportunities for a broad range of biomedical applications. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Production of membrane proteins without cells or detergents.

    PubMed

    Rajesh, Sundaresan; Knowles, Timothy; Overduin, Michael

    2011-04-30

    The production of membrane proteins in cellular systems is besieged by several problems due to their hydrophobic nature which often causes misfolding, protein aggregation and cytotoxicity, resulting in poor yields of stable proteins. Cell-free expression has emerged as one of the most versatile alternatives for circumventing these obstacles by producing membrane proteins directly into designed hydrophobic environments. Efficient optimisation of expression and solubilisation conditions using a variety of detergents, membrane mimetics and lipids has yielded structurally and functionally intact membrane proteins, with yields several fold above the levels possible from cell-based systems. Here we review recently developed techniques available to produce functional membrane proteins, and discuss amphipols, nanodisc and styrene maleic acid lipid particle (SMALP) technologies that can be exploited alongside cell-free expression of membrane proteins. Copyright © 2010 Elsevier B.V. All rights reserved.

  6. Short infrared laser pulses increase cell membrane fluidity

    NASA Astrophysics Data System (ADS)

    Walsh, Alex J.; Cantu, Jody C.; Ibey, Bennett L.; Beier, Hope T.

    2017-02-01

    Short infrared laser pulses induce a variety of effects in cells and tissues, including neural stimulation and inhibition. However, the mechanism behind these physiological effects is poorly understood. It is known that the fast thermal gradient induced by the infrared light is necessary for these biological effects. Therefore, this study tests the hypothesis that the fast thermal gradient induced in a cell by infrared light exposure causes a change in the membrane fluidity. To test this hypothesis, we used the membrane fluidity dye, di-4-ANEPPDHQ, to investigate membrane fluidity changes following infrared light exposure. Di-4-ANEPPDHQ fluorescence was imaged on a wide-field fluorescence imaging system with dual channel emission detection. The dual channel imaging allowed imaging of emitted fluorescence at wavelengths longer and shorter than 647 nm for ratiometric assessment and computation of a membrane generalized polarization (GP) value. Results in CHO cells show increased membrane fluidity with infrared light pulse exposure and this increased fluidity scales with infrared irradiance. Full recovery of pre-infrared exposure membrane fluidity was observed. Altogether, these results demonstrate that infrared light induces a thermal gradient in cells that changes membrane fluidity.

  7. Anhydrous Proton-Conducting Membranes for Fuel Cells

    NASA Technical Reports Server (NTRS)

    Narayanan, Sekharipuram; Yen, Shiao-Pin S.

    2005-01-01

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

  8. Influence of mechanical cell salvage on red blood cell aggregation, deformability, and 2,3-diphosphoglycerate in patients undergoing cardiac surgery with cardiopulmonary bypass.

    PubMed

    Gu, Y John; Vermeijden, Wytze J; de Vries, Adrianus J; Hagenaars, J Ans M; Graaff, Reindert; van Oeveren, Willem

    2008-11-01

    Mechanical cell salvage is increasingly used during cardiac surgery. Although this procedure is considered safe, it is unknown whether it affects the red blood cell (RBC) function, especially the RBC aggregation, deformability, and the contents of 2,3-diphosphoglycerate (2,3-DPG). This study examines the following: (1) whether the cell salvage procedure influences RBC function; and (2) whether retransfusion of the salvaged blood affects RBC function in patients. Forty patients undergoing cardiac surgery with cardiopulmonary bypass were randomly allocated to a cell saver group (n = 20) or a control group (n = 20). In the cell saver group, the blood aspirated from the wound area and the residual blood from the heart-lung machine were processed with a continuous-flow cell saver before retransfusion. In the control group this blood was retransfused without processing. The RBC aggregation and deformability were measured with a laser-assisted optical rotational cell analyzer and 2,3,-DPG by conventional laboratory test. The cell saver procedure did not influence the RBC aggregation but significantly reduced the RBC deformability (p = 0.007) and the content of RBC 2,3-DPG (p = 0.032). However, in patients receiving the processed blood, their intraoperative and postoperative RBC aggregation, deformability, and 2,3-DPG content did not differ from those of the control patients. Both groups of patients had a postoperative drop of RBC function as a result of hemodilution. The mechanical cell salvage procedure reduces the RBC deformability and the cell 2,3-DPG content. Retransfusion of the processed blood by cell saver does not further compromise the RBC function in patients undergoing cardiac surgery with cardiopulmonary bypass.

  9. Front-to-rear membrane tension gradient in rapidly moving cells.

    PubMed

    Lieber, Arnon D; Schweitzer, Yonatan; Kozlov, Michael M; Keren, Kinneret

    2015-04-07

    Membrane tension is becoming recognized as an important mechanical regulator of motile cell behavior. Although membrane-tension measurements have been performed in various cell types, the tension distribution along the plasma membrane of motile cells has been largely unexplored. Here, we present an experimental study of the distribution of tension in the plasma membrane of rapidly moving fish epithelial keratocytes. We find that during steady movement the apparent membrane tension is ∼30% higher at the leading edge than at the trailing edge. Similar tension differences between the front and the rear of the cell are found in keratocyte fragments that lack a cell body. This front-to-rear tension variation likely reflects a tension gradient developed in the plasma membrane along the direction of movement due to viscous friction between the membrane and the cytoskeleton-attached protein anchors embedded in the membrane matrix. Theoretical modeling allows us to estimate the area density of these membrane anchors. Overall, our results indicate that even though membrane tension equilibrates rapidly and mechanically couples local boundary dynamics over cellular scales, steady-state variations in tension can exist in the plasma membranes of moving cells. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  10. Neuroglian-mediated cell adhesion induces assembly of the membrane skeleton at cell contact sites

    PubMed Central

    1996-01-01

    The protein ankyrin links integral membrane proteins to the spectrin- based membrane skeleton. Ankyrin is often concentrated within restricted membrane domains of polarized epithelia and neurons, but the mechanisms responsible for membrane targeting and its segregation within a continuous lipid bilayer remain unexplained. We provide evidence that neuroglian, a cell adhesion molecule related to L1 and neurofascin, can transmit positional information directly to ankyrin and thereby polarize its distribution in Drosophila S2 tissue culture cells. Ankyrin was not normally associated with the plasma membrane of these cells. Upon expression of an inducible neuroglian minigene, however, cells aggregated into large clusters and ankyrin became concentrated at sites of cell-cell contact. Spectrin was also recruited to sites of cell contact in response to neuroglian expression. The accumulation of ankyrin at cell contacts required the presence of the cytoplasmic domain of neuroglian since a glycosyl phosphatidylinositol- linked form of neuroglian failed to recruit ankyrin to sites of cell- cell contact. Double-labeling experiments revealed that, whereas ankyrin was strictly associated with sites of cell-cell contact, neuroglian was more broadly distributed over the cell surface. A direct interaction between neuroglian and ankyrin was demonstrated using yeast two-hybrid analysis. Thus, neuroglian appears to be activated by extracellular adhesion so that ankyrin and the membrane skeleton selectively associate with sites of cell contact and not with other regions of the plasma membrane. PMID:8636238

  11. Does the extracorporeal circulation worsen anemia in hemodialysis patients? Investigation with advanced microscopes of red blood cells drawn at the beginning and end of dialysis

    PubMed Central

    Stamopoulos, Dimosthenis; Bakirtzi, Nerantzoula; Manios, Efthymios; Grapsa, Eirini

    2013-01-01

    Background In hemodialysis (HD) patients, anemia relates to three main factors: insufficient production of erythropoietin; impaired management of iron; and decreased lifespan of red blood cells (RBCs). The third factor can relate to structural deterioration of RBCs due to extrinsic (extracorporeal circuit; biochemical activation and/or mechanical stress during dialysis) and intrinsic (uremic milieu; biochemical interference of the RBC membrane constituents with toxins) mechanisms. Herein, we evaluate information accessed with advanced imaging techniques at the cellular level. Methods Atomic force and scanning electron microscopes were employed to survey intact RBCs (iRBCs) of seven HD patients in comparison to seven healthy donors. The extrinsic factor was investigated by contrasting pre- and post-HD samples. The intrinsic environment was investigated by comparing the microscopy data with the clinical ones. Results The iRBC membranes of the enrolled HD patients were overpopulated with orifice-like (high incidence; typical size within 100–1,000 nm) and crevice-like (low incidence; typical size within 500–4,000 nm) defects that exhibited a statistically significant (P < 0.05) relative increase (+55% and +350%, respectively) in respect to healthy donors. The relative variation of the orifice and crevice indices (mean population of orifices and crevices per top membrane surface) between pre- and post-HD was not statistically significant (−3.3% and +4.5%, respectively). The orifice index correlates with the concentrations of urea, calcium, and phosphorus, but not, however, with that of creatinine. Conclusion Extracorporeal circulation is not detrimental to the structural integrity of RBC membranes. Uremic milieu is a candidate cause of RBC membrane deterioration, which possibly worsens anemia. PMID:24143093

  12. ACME: Automated Cell Morphology Extractor for Comprehensive Reconstruction of Cell Membranes

    PubMed Central

    Mosaliganti, Kishore R.; Noche, Ramil R.; Xiong, Fengzhu; Swinburne, Ian A.; Megason, Sean G.

    2012-01-01

    The quantification of cell shape, cell migration, and cell rearrangements is important for addressing classical questions in developmental biology such as patterning and tissue morphogenesis. Time-lapse microscopic imaging of transgenic embryos expressing fluorescent reporters is the method of choice for tracking morphogenetic changes and establishing cell lineages and fate maps in vivo. However, the manual steps involved in curating thousands of putative cell segmentations have been a major bottleneck in the application of these technologies especially for cell membranes. Segmentation of cell membranes while more difficult than nuclear segmentation is necessary for quantifying the relations between changes in cell morphology and morphogenesis. We present a novel and fully automated method to first reconstruct membrane signals and then segment out cells from 3D membrane images even in dense tissues. The approach has three stages: 1) detection of local membrane planes, 2) voting to fill structural gaps, and 3) region segmentation. We demonstrate the superior performance of the algorithms quantitatively on time-lapse confocal and two-photon images of zebrafish neuroectoderm and paraxial mesoderm by comparing its results with those derived from human inspection. We also compared with synthetic microscopic images generated by simulating the process of imaging with fluorescent reporters under varying conditions of noise. Both the over-segmentation and under-segmentation percentages of our method are around 5%. The volume overlap of individual cells, compared to expert manual segmentation, is consistently over 84%. By using our software (ACME) to study somite formation, we were able to segment touching cells with high accuracy and reliably quantify changes in morphogenetic parameters such as cell shape and size, and the arrangement of epithelial and mesenchymal cells. Our software has been developed and tested on Windows, Mac, and Linux platforms and is available

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

    PubMed

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

    2014-07-24

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

  14. An adhesion-based method for plasma membrane isolation: evaluating cholesterol extraction from cells and their membranes.

    PubMed

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

    2009-11-15

    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.

  15. Erythrocyte disorders leading to potassium loss and cellular dehydration.

    PubMed

    Glader, B E; Sullivan, D W

    1979-01-01

    RBC K loss and cellular dehydration are associated with a variety of normal and abnormal erythrocyte conditions. In some cases (normal RBC aging, pyruvate-kinase-deficient RBCs and irreversibly sickled cells) cation and water changes are related to adenosine triphosphate (ATP) depletion and to increased RBC calcium content. In other disorders, such as hereditary xerocytosis, cation depletion and cellular hydration are not related to altered energy or calcium metabolism. Rather, this condition is thought to be due to a structural membrane defect which is manifested by imbalanced cation leaks (K less greater than Na gain) for which the active cation transport is unable to compensate. None of the disorders described here are associated with known structural membrane alterations. The fact that K loss and cellular dehydration are common to several RBC disorders suggests that this phenomenon may have a direct role in membrane injury. This hypothesis is supported by two separate observations: 1)Formation of irreversible sickled cells in vitro is prevented if K and water loss are inhibited, and these effects are independent of ATP depletion and calcium accumulation; 2) the mean critical hemolytic volume is markedly reduced in K- and water-depleted normal RBCs. RBC dehydration without intracellular cation depletion, however, is not associated with changes in mean critical hemolytic volume. These data thus indicate that K loss may have a direct role in RBC membrane injury. The mechanism by which this occurs and the associated alterations in membrane structure, however, remain to be identified.

  16. The role of red blood cell deformability and Na,K-ATPase function in selected risk factors of cardiovascular diseases in humans: focus on hypertension, diabetes mellitus and hypercholesterolemia.

    PubMed

    Radosinska, J; Vrbjar, N

    2016-09-19

    Deformability of red blood cells (RBC) is the ability of RBC to change their shape in order to pass through narrow capillaries in circulation. Deterioration in deformability of RBC contributes to alterations in microcirculatory blood flow and delivery of oxygen to tissues. Several factors are responsible for maintenance of RBC deformability. One of them is the Na,K-ATPase known as crucial enzyme in maintenance of intracellular ionic homeostasis affecting thus regulation of cellular volume and consequently RBC deformability. Decreased deformability of RBC has been found to be the marker of adverse outcomes in cardiovascular diseases (CVD) and the presence of cardiovascular risk factors influences rheological properties of the blood. This review summarizes knowledge concerning the RBC deformability in connection with selected risk factors of CVD, including hypertension, hyperlipidemia, and diabetes mellitus, based exclusively on papers from human studies. We attempted to provide an update on important issues regarding the role of Na,K-ATPase in RBC deformability. In patients suffering from hypertension as well as diabetes mellitus the Na,K-ATPase appears to be responsible for the changes leading to alterations in RBC deformability. The triggering factor for changes of RBC deformability during hypercholesterolemia seems to be the increased content of cholesterol in erythrocyte membranes.

  17. Characterization of femtosecond-laser pulse induced cell membrane nanosurgical attachment.

    PubMed

    Katchinskiy, Nir; Godbout, Roseline; Elezzabi, Abdulhakem Y

    2016-07-01

    This article provides insight into the mechanism of femtosecond laser nanosurgical attachment of cells. We have demonstrated that during the attachment of two retinoblastoma cells using sub-10 femtosecond laser pulses, with 800 nm central wavelength, the phospholipid molecules of both cells hemifuse and form one shared phospholipid bilayer, at the attachment location. In order to verify the hypothesis that hemifusion takes place, transmission electron microscope images of the cell membranes of retinoblastoma cells were taken. It is shown that at the attachment interface, the two cell membranes coalesce and form one single membrane shared by both cells. Thus, further evidence is provided to support the hypothesis that laser-induced ionization process led to an ultrafast reversible destabilization of the phospholipid layer of the cellular membrane, which resulted in cross-linking of the phospholipid molecules in each membrane. This process of hemifusion occurs throughout the entire penetration depth of the femtosecond laser pulse train. Thus, the attachment between the cells takes place across a large surface area, which affirms our findings of strong physical attachment between the cells. The femtosecond laser pulse hemifusion technique can potentially provide a platform for precise molecular manipulation of cellular membranes. Manipulation of the cellular membrane is an important procedure that could aid in studying diseases such as cancer; where the expression level of plasma proteins on the cell membrane is altered.

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

    PubMed

    Chong, Ketpin; Deng, Yuru

    2012-01-01

    Biological membranes are generally perceived as phospholipid bilayer structures that delineate in a lamellar form the cell surface and intracellular organelles. However, much more complex and highly convoluted membrane organizations are ubiquitously present in many cell types under certain types of stress, states of disease, or in the course of viral infections. Their occurrence under pathological conditions make such three-dimensionally (3D) folded and highly ordered membranes attractive biomarkers. They have also stimulated great biomedical interest in understanding the molecular basis of their formation. Currently, the analysis of such membrane arrangements, which include tubulo-reticular structures (TRS) or cubic membranes of various subtypes, is restricted to electron microscopic methods, including tomography. Preservation of membrane structures during sample preparation is the key to understand their true 3D nature. This chapter discusses methods for appropriate sample preparations to successfully examine and analyze well-preserved highly ordered membranes by electron microscopy. Processing methods and analysis conditions for green algae (Zygnema sp.) and amoeba (Chaos carolinense), mammalian cells in culture and primary tissue cells are described. We also discuss methods to identify cubic membranes by transmission electron microscopy (TEM) with the aid of a direct template matching method and by computer simulation. A 3D analysis of cubic cell membrane topology by electron tomography is described as well as scanning electron microscopy (SEM) to investigate surface contours of isolated mitochondria with cubic membrane arrangement. Copyright © 2012 Elsevier Inc. All rights reserved.

  19. Isolation, expression and characterization of rbcL gene from Ulva prolifera J. Agardh (Ulvophyceae, Chlorophyta)

    NASA Astrophysics Data System (ADS)

    Shao, Zhanru; Li, Wei; Guo, Hui; Duan, Delin

    2015-12-01

    Ulva prolifera is a typical green alga in subtidal areas and can grow tremendously fast. A highly efficient Rubisco enzyme which is encoded by UpRbcL gene may contribute to the rapid growth. In this study, the full-length UpRbcL open reading frame (ORF) was identified, which encoded a protein of 474 amino acids. Phylogenetic analysis of UpRbcL sequences revealed that Chlorophyta had a closer genetic relationship with higher plants than with Rhodophyta and Phaeophyta. The two distinct residues (aa11 and aa91) were presumed to be unique for Rubisco catalytic activity. The predicted three-dimensional structure showed that one α/β-barrel existed in the C-terminal region, and the sites for Mg2+ coordination and CO2 fixation were also located in this region. Gene expression profile indicated that UpRbcL was expressed at a higher level under light exposure than in darkness. When the culture temperature reached 35°C, the expression level of UpRbcL was 2.5-fold lower than at 15°C, and the carboxylase activity exhibited 13.8-fold decrease. UpRbcL was heterologously expressed in E. coli and was purified by Ni2+ affinity chromatography. The physiological and biochemical characterization of recombinant Rubisco will be explored in the future.

  20. The role of phosphatidylserine in recognition and removal of erythrocytes.

    PubMed

    Kuypers, F A; de Jong, K

    2004-03-01

    During the time that erythrocytes (RBC) spend in the circulation, a series of progressive events take place that lead to their removal and determine their apparent aging and limited survival. In addition, a fraction of RBC precursors will be removed during erythropoiesis by apoptotic processes, often described as "ineffective erythropoiesis". Both will determine the survival of erythroid cells and play an important role in red cell pathology, including hemoglobinopathies and red cell membrane disorders. The loss of phospholipid asymmetry, and the exposure of phosphatidylserine (PS) on the surface of plasma membranes may be a general trigger by which cells, including aging RBC and apoptotic cells, are removed. Oxidant stress and inactivation of the system that maintains phospholipid asymmetry play a central role in the events that will lead to PS exposure, death and removal.

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

    PubMed

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

    2015-12-15

    Apicomplexa parasites such as Toxoplasma gondii target effectors to and across the boundary of their parasitophorous vacuole (PV), resulting in host cell subversion and potential presentation by MHC class I molecules for CD8 T cell recognition. The host-parasite interface comprises the PV limiting membrane and a highly curved, membranous intravacuolar network (IVN) of uncertain function. Here, using a cell-free minimal system, we dissect how membrane tubules are shaped by the parasite effectors GRA2 and GRA6. We show that membrane association regulates access of the GRA6 protective antigen to the MHC I pathway in infected cells. Although insertion of GRA6 in the PV membrane is key for immunogenicity, association of GRA6 with the IVN limits presentation and curtails GRA6-specific CD8 responses in mice. Thus, membrane deformations of the PV regulate access of antigens to the MHC class I pathway, and the IVN may play a role in immune modulation. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  2. Sequences 5' to translation start regulate expression of petunia rbcS genes.

    PubMed Central

    Dean, C; Favreau, M; Bedbrook, J; Dunsmuir, P

    1989-01-01

    The promoter sequences that contribute to quantitative differences in expression of the petunia genes (rbcS) encoding the small subunit of ribulose bisphosphate carboxylase have been characterized. The promoter regions of the two most abundantly expressed petunia rbcS genes, SSU301 and SSU611, show sequence similarity not present in other rbcS genes. We investigated the significance of these and other sequences by adding specific regions from the SSU301 promoter (the most strongly expressed gene) to equivalent regions in the SSU911 promoter (the least strongly expressed gene) and assaying the expression of the fusions in transgenic tobacco plants. In this way, we characterized an SSU301 promoter region (either from -285 to -178 or -291 to -204) which, when added to SSU911, in either orientation, increased SSU911 expression 25-fold. This increase was equivalent to that caused by addition of the entire SSU301 5'-flanking region. Replacement of SSU911 promoter sequences between -198 and the start codon with sequences from the equivalent region of SSU301 did not increase SSU911 expression significantly. The -291 to -204 SSU301 promoter fragment contributes significantly to quantitative differences in expression between the petunia rbcS genes. PMID:2535543

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

    PubMed

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

    2015-05-18

    Membrane proteins may be influenced by the environment, and they may be unstable in detergents or fail to crystallize. As a result, approaches to characterize structures in a native environment are highly desirable. Here, we report a novel general strategy for precise distance measurements on outer membrane proteins in whole Escherichia coli cells and isolated outer membranes. The cobalamin transporter BtuB was overexpressed and spin-labeled in whole cells and outer membranes and interspin distances were measured to a spin-labeled cobalamin using pulse EPR spectroscopy. A comparative analysis of the data reveals a similar interspin distance between whole cells, outer membranes, and synthetic vesicles. This approach provides an elegant way to study conformational changes or protein-protein/ligand interactions at surface-exposed sites of membrane protein complexes in whole cells and native membranes, and provides a method to validate outer membrane protein structures in their native environment. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

    NASA Technical Reports Server (NTRS)

    Eisman, G. A.

    1989-01-01

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

  5. Red blood cells promote survival and cell cycle progression of human peripheral blood T cells independently of CD58/LFA-3 and heme compounds.

    PubMed

    Fonseca, Ana Mafalda; Pereira, Carlos Filipe; Porto, Graça; Arosa, Fernando A

    2003-07-01

    Red blood cells (RBC) are known to modulate T cell proliferation and function possibly through downregulation of oxidative stress. By examining parameters of activation, division, and cell death in vitro, we show evidence that the increase in survival afforded by RBC is due to the maintenance of the proliferative capacity of the activated T cells. We also show that the CD3+CD8+ T cell subset was preferentially expanded and rescued from apoptosis both in bulk peripheral blood lymphocyte cultures and with highly purified CD8+ T cells. The ability of RBC to induce survival of dividing T cells was not affected by blocking the CD58/CD2 interaction. Moreover, addition of hemoglobin, heme or protoporphyrin IX to cultures of activated T cells did not reproduce the effect of intact RBC. Considering that RBC circulate throughout the body, they could play a biological role in the modulation of T cell differentiation and survival in places of active cell division. Neither CD58 nor the heme compounds studied seem to play a direct relevant role in the modulation of T cell survival.

  6. Mechanism of vaso-occlusion in sickle cell anemia

    NASA Astrophysics Data System (ADS)

    Lei, Huan; Karniadakis, George

    2012-11-01

    Vaso-occlusion crisis is one of the key hallmark of sickle cell anemia. While early studies suggested that the crisis is caused by blockage of a single elongated cell, recent experimental investigations indicate that vaso-occlusion is a complex process triggered by adhesive interactions among different cell groups in multiple stages. Based on dissipative particle dynamics, a multi-scale model for the sickle red blood cells (SS-RBCs), accounting for diversity in both shapes and cell rigidities, is developed to investigate the mechanism of vaso-occlusion crisis. Using this model, the adhesive dynamics of single SS-RBC was investigated in arterioles. Simulation results indicate that the different cell groups (deformable SS2 RBCs, rigid SS4 RBCs, leukocytes, etc.) exhibit heterogeneous adhesive behavior due to the different cell morphologies and membrane rigidities. We further simulate the tube flow of SS-RBC suspensions with different cell fractions. The more adhesive SS2 cells interact with the vascular endothelium and further trap rigid SS4 cells, resulting in vaso-occlusion in vessels less than 15 μm . Under inflammation, adherent leukocytes may also trap SS4 cells, resulting in vaso-occlusion in even larger vessels. This work was supported by the NSF grant CBET-0852948 and the NIH grant R01HL094270.

  7. High red blood cell nitric oxide synthase activation is not associated with improved vascular function and red blood cell deformability in sickle cell anaemia.

    PubMed

    Grau, Marijke; Mozar, Anaïs; Charlot, Keyne; Lamarre, Yann; Weyel, Linda; Suhr, Frank; Collins, Bianca; Jumet, Stéphane; Hardy-Dessources, Marie-Dominique; Romana, Marc; Lemonne, Nathalie; Etienne-Julan, Maryse; Antoine-Jonville, Sophie; Bloch, Wilhelm; Connes, Philippe

    2015-03-01

    Human red blood cells (RBC) express an active and functional endothelial-like nitric oxide (NO) synthase (RBC-NOS). We report studies on RBC-NOS activity in sickle cell anaemia (SCA), a genetic disease characterized by decreased RBC deformability and vascular dysfunction. Total RBC-NOS content was not significantly different in SCA patients compared to healthy controls; however, using phosphorylated RBC-NOS-Ser(1177) as a marker, RBC-NOS activation was higher in SCA patients as a consequence of the greater activation of Akt (phosphorylated Akt-Ser(473) ). The higher RBC-NOS activation in SCA led to higher levels of S-nitrosylated α- and β-spectrins, and greater RBC nitrite and nitrotyrosine levels compared to healthy controls. Plasma nitrite content was not different between the two groups. Laser Doppler flowmetric experiments demonstrated blunted microcirculatory NO-dependent response under hyperthermia in SCA patients. RBC deformability, measured by ektacytometry, was reduced in SCA in contrast to healthy individuals, and pre-shearing RBC in vitro did not improve deformability despite an increase of RBC-NOS activation. RBC-NOS activation is high in freshly drawn blood from SCA patients, resulting in high amounts of NO produced by RBC. However, this does not result in improved RBC deformability and vascular function: higher RBC-NO is not sufficient to counterbalance the enhanced oxidative stress in SCA. © 2014 John Wiley & Sons Ltd.

  8. Microstructured Electrolyte Membranes to Improve Fuel Cell Performance

    NASA Astrophysics Data System (ADS)

    Wei, Xue

    Fuel cells, with the advantages of high efficiency, low greenhouse gas emission, and long lifetime are a promising technology for both portable power and stationary power sources. The development of efficient electrolyte membranes with high ionic conductivity, good mechanical durability and dense structure at low cost remains a challenge to the commercialization of fuel cells. This thesis focuses on exploring novel composite polymer membranes and ceramic electrolytes with the microstructure engineered to improve performance in direct methanol fuel cells (DMFCs) and solid oxide fuel cells (SOFCs), respectively. Polymer/particle composite membranes hold promise to meet the demands of DMFCs at lower cost. The structure of composite membranes was controlled by aligning proton conducting particles across the membrane thickness under an applied electric field. The field-induced structural changes caused the membranes to display an enhanced water uptake, proton conductivity, and methanol permeability in comparison to membranes prepared without an applied field. Although both methanol permeability and proton conductivity are enhanced by the applied field, the permeability increase is relatively lower than the proton conductivity improvement, which results in enhanced proton/methanol selectivity and improved DMFC performance. Apatite ceramics are a new class of fast ion conductors being studied as alternative SOFC electrolytes in the intermediate temperature range. An electrochemical/hydrothermal deposition method was developed to grow fully dense apatite membranes containing well-developed crystals with c-axis alignment to promote ion conductivity. Hydroxyapatite seed crystals were first deposited onto a metal substrate electrochemically. Subsequent ion substitution during the hydrothermal growth process promoted the formation of dense, fully crystalline films with microstructure optimal for ion transport. The deposition parameters were systematically investigated, such as

  9. Analysis of novel sph (spherocytosis) alleles in mice reveals allele-specific loss of band 3 and adducin in α-spectrin–deficient red cells

    PubMed Central

    Robledo, Raymond F.; Lambert, Amy J.; Birkenmeier, Connie S.; Cirlan, Marius V.; Cirlan, Andreea Flavia M.; Campagna, Dean R.; Lux, Samuel E.

    2010-01-01

    Five spontaneous, allelic mutations in the α-spectrin gene, Spna1, have been identified in mice (spherocytosis [sph], sph1J, sph2J, sph2BC, sphDem). All cause severe hemolytic anemia. Here, analysis of 3 new alleles reveals previously unknown consequences of red blood cell (RBC) spectrin deficiency. In sph3J, a missense mutation (H2012Y) in repeat 19 introduces a cryptic splice site resulting in premature termination of translation. In sphIhj, a premature stop codon occurs (Q1853Stop) in repeat 18. Both mutations result in markedly reduced RBC membrane spectrin content, decreased band 3, and absent β-adducin. Reevaluation of available, previously described sph alleles reveals band 3 and adducin deficiency as well. In sph4J, a missense mutation occurs in the C-terminal EF hand domain (C2384Y). Notably, an equally severe hemolytic anemia occurs despite minimally decreased membrane spectrin with normal band 3 levels and present, although reduced, β-adducin. The severity of anemia in sph4J indicates that the highly conserved cysteine residue at the C-terminus of α-spectrin participates in interactions critical to membrane stability. The data reinforce the notion that a membrane bridge in addition to the classic protein 4.1-p55-glycophorin C linkage exists at the RBC junctional complex that involves interactions between spectrin, adducin, and band 3. PMID:20056793

  10. Analysis of novel sph (spherocytosis) alleles in mice reveals allele-specific loss of band 3 and adducin in alpha-spectrin-deficient red cells.

    PubMed

    Robledo, Raymond F; Lambert, Amy J; Birkenmeier, Connie S; Cirlan, Marius V; Cirlan, Andreea Flavia M; Campagna, Dean R; Lux, Samuel E; Peters, Luanne L

    2010-03-04

    Five spontaneous, allelic mutations in the alpha-spectrin gene, Spna1, have been identified in mice (spherocytosis [sph], sph(1J), sph(2J), sph(2BC), sph(Dem)). All cause severe hemolytic anemia. Here, analysis of 3 new alleles reveals previously unknown consequences of red blood cell (RBC) spectrin deficiency. In sph(3J), a missense mutation (H2012Y) in repeat 19 introduces a cryptic splice site resulting in premature termination of translation. In sph(Ihj), a premature stop codon occurs (Q1853Stop) in repeat 18. Both mutations result in markedly reduced RBC membrane spectrin content, decreased band 3, and absent beta-adducin. Reevaluation of available, previously described sph alleles reveals band 3 and adducin deficiency as well. In sph(4J), a missense mutation occurs in the C-terminal EF hand domain (C2384Y). Notably, an equally severe hemolytic anemia occurs despite minimally decreased membrane spectrin with normal band 3 levels and present, although reduced, beta-adducin. The severity of anemia in sph(4J) indicates that the highly conserved cysteine residue at the C-terminus of alpha-spectrin participates in interactions critical to membrane stability. The data reinforce the notion that a membrane bridge in addition to the classic protein 4.1-p55-glycophorin C linkage exists at the RBC junctional complex that involves interactions between spectrin, adducin, and band 3.

  11. Purification and differentiation of human adipose-derived stem cells by membrane filtration and membrane migration methods

    PubMed Central

    Lin, Hong Reng; Heish, Chao-Wen; Liu, Cheng-Hui; Muduli, Saradaprasan; Li, Hsing-Fen; Higuchi, Akon; Kumar, S. Suresh; Alarfaj, Abdullah A.; Munusamy, Murugan A.; Hsu, Shih-Tien; Chen, Da-Chung; Benelli, Giovanni; Murugan, Kadarkarai; Cheng, Nai-Chen; Wang, Han-Chow; Wu, Gwo-Jang

    2017-01-01

    Human adipose-derived stem cells (hADSCs) are easily isolated from fat tissue without ethical concerns, but differ in purity, pluripotency, differentiation ability, and stem cell marker expression, depending on the isolation method. We isolated hADSCs from a primary fat tissue solution using: (1) conventional culture, (2) a membrane filtration method, (3) a membrane migration method where the primary cell solution was permeated through membranes, adhered hADSCs were cultured, and hADSCs migrated out from the membranes. Expression of mesenchymal stem cell markers and pluripotency genes, and osteogenic differentiation were compared for hADSCs isolated by different methods using nylon mesh filter membranes with pore sizes ranging from 11 to 80 μm. hADSCs isolated by the membrane migration method had the highest MSC surface marker expression and efficient differentiation into osteoblasts. Osteogenic differentiation ability of hADSCs and MSC surface marker expression were correlated, but osteogenic differentiation ability and pluripotent gene expression were not. PMID:28071738

  12. Membrane protein synthesis in cell-free systems: from bio-mimetic systems to bio-membranes.

    PubMed

    Sachse, Rita; Dondapati, Srujan K; Fenz, Susanne F; Schmidt, Thomas; Kubick, Stefan

    2014-08-25

    When taking up the gauntlet of studying membrane protein functionality, scientists are provided with a plethora of advantages, which can be exploited for the synthesis of these difficult-to-express proteins by utilizing cell-free protein synthesis systems. Due to their hydrophobicity, membrane proteins have exceptional demands regarding their environment to ensure correct functionality. Thus, the challenge is to find the appropriate hydrophobic support that facilitates proper membrane protein folding. So far, various modes of membrane protein synthesis have been presented. Here, we summarize current state-of-the-art methodologies of membrane protein synthesis in biomimetic-supported systems. The correct folding and functionality of membrane proteins depend in many cases on their integration into a lipid bilayer and subsequent posttranslational modification. We highlight cell-free systems utilizing the advantages of biological membranes. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  13. Photothermal nanoblade for patterned cell membrane cutting

    PubMed Central

    Wu, Ting-Hsiang; Teslaa, Tara; Teitell, Michael A.; Chiou, Pei-Yu

    2010-01-01

    We report a photothermal nanoblade that utilizes a metallic nanostructure to harvest short laser pulse energy and convert it into a highly localized and specifically shaped explosive vapor bubble. Rapid bubble expansion and collapse punctures a lightly-contacting cell membrane via high-speed fluidic flows and induced transient shear stress. The membrane cutting pattern is controlled by the metallic nanostructure configuration, laser pulse polarization, and energy. Highly controllable, sub-micron sized circular hole pairs to half moon-like, or cat-door shaped, membrane cuts were realized in glutaraldehyde treated HeLa cells. PMID:21164656

  14. Biophysics of malarial parasite exit from infected erythrocytes.

    PubMed

    Chandramohanadas, Rajesh; Park, YongKeun; Lui, Lena; Li, Ang; Quinn, David; Liew, Kingsley; Diez-Silva, Monica; Sung, Yongjin; Dao, Ming; Lim, Chwee Teck; Preiser, Peter Rainer; Suresh, Subra

    2011-01-01

    Upon infection and development within human erythrocytes, P. falciparum induces alterations to the infected RBC morphology and bio-mechanical properties to eventually rupture the host cells through parasitic and host derived proteases of cysteine and serine families. We used previously reported broad-spectrum inhibitors (E64d, EGTA-AM and chymostatin) to inhibit these proteases and impede rupture to analyze mechanical signatures associated with parasite escape. Treatment of late-stage iRBCs with E64d and EGTA-AM prevented rupture, resulted in no major RBC cytoskeletal reconfiguration but altered schizont morphology followed by dramatic re-distribution of three-dimensional refractive index (3D-RI) within the iRBC. These phenotypes demonstrated several-fold increased iRBC membrane flickering. In contrast, chymostatin treatment showed no 3D-RI changes and caused elevated fluctuations solely within the parasitophorous vacuole. We show that E64d and EGTA-AM supported PV breakdown and the resulting elevated fluctuations followed non-Gaussian pattern that resulted from direct merozoite impingement against the iRBC membrane. Optical trapping experiments highlighted reduced deformability of the iRBC membranes upon rupture-arrest, more specifically in the treatments that facilitated PV breakdown. Taken together, our experiments provide novel mechanistic interpretations on the role of parasitophorous vacuole in maintaining the spherical schizont morphology, the impact of PV breakdown on iRBC membrane fluctuations leading to eventual parasite escape and the evolution of membrane stiffness properties of host cells in which merozoites were irreversibly trapped, recourse to protease inhibitors. These findings provide a comprehensive, previously unavailable, body of information on the combined effects of biochemical and biophysical factors on parasite egress from iRBCs.

  15. Optomechanical characterization of proton-exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Jalani, Nikhil H.; Mizar, Shivananda P.; Choi, Pyoungho; Furlong, Cosme; Datta, Ravindra

    2004-08-01

    Nafion is widely used as the polymer electrolyte in proton exchange membrane (PEM) fuel cells. The properties that make the Nafion membrane indispensable are the combination of good water uptake, ion-exchange capacity, proton conductivity, gas permeability, and excellent electrochemical stability. The amount of water sorbed in the Nafion membrane is critical as the proton conductivity depends directly on the water content of the membrane which determines the fuel cell performance. The factors which affect the extent of the solvent uptake by Nafion are temperature, ion-exchange capacity, pretreatment of membrane, and the physical state of absorbing water, whether it is in liquid or vapor phase. The water sorption in the membrane is explained in terms of thermodynamic equilibrium of water in the vapor and absorption phases. As the membrane imbibes more water, the membrane matrix expands and exerts a pressure on the pore liquid which affects its chemical potential and limits extent of swelling. The extent of matrix expansion of the membranes depends on the elastic modulus, E, of the membrane, which directly affects the sorption. Hence, it is important to understand the variation of E for Nafion membrane with relative humidity (RH) and temperature. Optoelectronic holography (OEH) techniques are applied to perform quantitative, noninvasive, full field of view investigations to determine temperature and water activity dependence of E. The results obtained confirm that with the increase in temperature, E decreases and the membranes imbibes more water. Such results will allow optimization and realization of fuel cells with improved efficiency and performance.

  16. Correlated alterations in prostate basal cell layer and basement membrane

    PubMed Central

    Liu, Aijun; Wei, Lixin; Gardner, William A.; Deng, Chu-Xia; Man, Yan-Gao

    2009-01-01

    Our recent studies revealed that focal basal cell layer disruption (FBCLD) induced auto-immunoreactions represented a contributing factor for human prostate tumor progression and invasion. As the basement membrane surrounds and attaches to the basal cell layer, our current study assessed whether FBCLD would impact the physical integrity of the associated basement membrane. Paraffin sections from 25-human prostate tumors were subjected to double immunohistochemistry to simultaneously elucidate the basal cell layer and the basement membrane with corresponding biomarkers. The physical integrity of the basement membrane overlying FBCLD was examined to determine the extent of correlated alterations. Of a total of 89 FBCLD encountered, 76 (85 %) showed correlated alterations in the overlying basement membrane, which included distinct focal disruptions or fragmentations. In the remaining 13 (15%) FBCLD, the overlying basement membrane showed significant attenuation or reduction of the immunostaining intensity. The basement membrane in all or nearly all ducts or acini with p63 positive basal cells was substantially thicker and more uniform than that in ducts or acini without p63 positive basal cells, and also, a vast majority of the focal disruptions occurred near basal cells that lack p63 expression. These findings suggest that focal disruptions in the basal cell layer and alterations in the basement membrane are correlated events and that the physical and functional status of the basal cells could significantly impact the physical integrity of the overlying basement membrane. As the degradation of both the basal cell layer and the basement membrane is a pre-requisite for prostate tumor invasion or progression, ducts or acini with focally disrupted basal cell layer and basement membrane are likely at greater risk to develop invasive lesions. Thus, further elucidation of the specific molecules and mechanism associated with these events may lead to the development of a more

  17. Apigenin induced apoptosis in esophageal carcinoma cells by destruction membrane structures.

    PubMed

    Zhu, Haiyan; Jin, Hua; Pi, Jiang; Bai, Haihua; Yang, Fen; Wu, Chaomin; Jiang, Jinhuan; Cai, Jiye

    2016-07-01

    Apigenin has shown to have killing effects on some kinds of solid tumor cells. However, the changes in cell membrane induced by apigenin on subcellular- or nanometer-level were still unclear. In this work, human esophageal cancer cells (EC9706 and KYSE150 cells) were employed as cell model to detect the cytotoxicity of apigenin, including cell growth inhibition, apoptosis induction, membrane toxicity, etc. MTT assay showed that apigenin could remarkably inhibit the growth and proliferation in both types of cells. Annexin V/PI-based flow cytometry analysis showed that the cytotoxic effects of apigenin in KYSE150 cells were mainly through early apoptosis induction, while in EC9706 cells, necrosis, and apoptosis were both involved in cell death. The morphological and ultrastructural properties induced by apigenin were investigated at single cellular- or nanometer-level using atomic force microscopy (AFM). Additionally, lactate dehydrogenase (LDH) leakage was measured to assess the changes in membrane permeability. The results indicated that apigenin increased the membrane permeability and caused leakage of LDH, which was consistent with damages on membrane ultrastructure detected by AFM. Therefore, membrane toxicity, including membrane ultrastructure damages and enhanced membrane permeability, played vital roles in apigenin induced human esophageal cancer cell apoptosis. SCANNING 38:322-328, 2016. © 2015 Wiley Periodicals, Inc. © Wiley Periodicals, Inc.

  18. RED BLOOD CELL STORAGE LESION

    PubMed Central

    Kor, Daryl J.; Van Buskirk, Camille M; Gajic, Ognjen

    2009-01-01

    The past two decades have witnessed increased scrutiny regarding efficacy and risk of the once unquestioned therapy of red blood cell (RBC) transfusion. Simultaneously, a variety of changes have been identified within the RBC and storage media during RBC preservation that are correlated with reduced tissue oxygenation and transfusion-associated adverse effects. These alterations are collectively termed the storage lesion and include extensive biochemical, biomechanical, and immunologic changes involving cells of diverse origin. Time-dependent falls is 2,3-diphosphoglycerate, intracellular RBC adenosine triphosphate, and nitric oxide have been shown to impact RBC deformability and delivery of oxygen to the end-organ. The accumulation of biologic response modifiers such as soluble CD40 ligand (sCD40L), lyso-phosphatidylcholine (lyso-PC), and Regulated on Activation, Normal T-cell Expressed and Secreted (RANTES) have been associated with altered recipient immune function as well. This review will address the alterations occurring within the RBC and storage media during RBC preservation and will address the potential clinical consequence thereof.

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

    NASA Astrophysics Data System (ADS)

    Haryadi, Sugianto, D.; Ristopan, E.

    2015-12-01

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

  20. Polybenzimidazole/Mxene composite membranes for intermediate temperature polymer electrolyte membrane fuel cells.

    PubMed

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

    2018-01-19

    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 Ti 3 C 2 T x -MXene was synthesized and incorporated into polybenzimidazole (PBI)-based membranes by using a solution blending method. The composite membrane with 3 wt% Ti 3 C 2 T 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 Ti 3 C 2 T x -MXene also improved the mechanical properties and thermal stability of PBI membranes. At 3 wt% Ti 3 C 2 T 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.

  1. Polybenzimidazole/Mxene composite membranes for intermediate temperature polymer electrolyte membrane fuel cells

    NASA Astrophysics Data System (ADS)

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

    2018-01-01

    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.

  2. Block Copolymers for Alkaline Fuel Cell Membrane Materials

    DTIC Science & Technology

    2014-07-30

    temperature fuel cells including proton exchange membrane fuel cell ( PEMFC ) and alkaline fuel cell (AFC) with operation temperature usually lower than 120...advantages over proton exchange membrane fuel cells ( PEMFCs ) resulting in the popularity of AFCs in the US space program.[8-11] The primary benefit AFC...offered over PEMFC is better electrochemical kinetics on the anode and cathode under the alkaline environment, which results in the ability to use

  3. Transforming growth factor-β released by apoptotic white blood cells during red blood cell storage promotes transfusion-induced alloimmunomodulation.

    PubMed

    Vallion, Romain; Bonnefoy, Francis; Daoui, Anna; Vieille, Loredane; Tiberghien, Pierre; Saas, Philippe; Perruche, Sylvain

    2015-07-01

    Red blood cell (RBC) alloimmunization is a major immunologic risk of transfusion. However, RBC storage facilitates white blood cell (WBC) apoptosis and apoptotic cells have immunomodulatory properties. We investigated the behavior of WBCs, and apoptosis in particular, in RBC units during storage and then studied the impact of WBC apoptosis on the modulation of posttransfusion alloimmunization in RBC products stored short term. We used a mouse model of alloimmunization to transfused HEL-ovalbumin-Duffy (HOD) surface antigen expressed specifically on RBCs. The presence of circulating anti-HOD immunoglobulin G detected by flow cytometry confirmed immunization to HOD+ RBCs. WBC apoptosis and factors released by apoptotic WBCs during storage were determined and in particular the role of transforming growth factor (TGF)-β was assessed on RBC alloimmunization. In blood stored 72 hours, 30% of WBCs were apoptotic, and transfusion of short-term-stored blood resulted in lesser immunization than did fresh blood or stored leukoreduced (LR) RBCs. WBCs undergoing apoptosis released during short-term storage factors modulating RBC alloimmunization. Indeed apoptotic cell-released factors modulate alloimmunization whereas exogenous apoptotic cells directly transfused with LR RBCs did not. While microparticles released during RBC storage had no immunomodulatory role, TGF-β found in the supernatant of stored blood demonstrated the capacity to favor Treg polarization of naïve CD4+CD25- T cells in vitro and limited RBC alloimmunization in vivo. Indeed, addition of recombinant TGF-β to stored LR RBC transfusion strongly limited posttransfusion RBC alloimmunization. Our findings show that short-term storage of non-LR blood facilitates WBC apoptosis therefore releasing TGF-β that modulates posttransfusion RBC alloimmunization. © 2015 AABB.

  4. Single-cell mechanics--An experimental-computational method for quantifying the membrane-cytoskeleton elasticity of cells.

    PubMed

    Tartibi, M; Liu, Y X; Liu, G-Y; Komvopoulos, K

    2015-11-01

    The membrane-cytoskeleton system plays a major role in cell adhesion, growth, migration, and differentiation. F-actin filaments, cross-linkers, binding proteins that bundle F-actin filaments to form the actin cytoskeleton, and integrins that connect the actin cytoskeleton network to the cell plasma membrane and extracellular matrix are major cytoskeleton constituents. Thus, the cell cytoskeleton is a complex composite that can assume different shapes. Atomic force microscopy (AFM)-based techniques have been used to measure cytoskeleton material properties without much attention to cell shape. A recently developed surface chemical patterning method for long-term single-cell culture was used to seed individual cells on circular patterns. A continuum-based cell model, which uses as input the force-displacement response obtained with a modified AFM setup and relates the membrane-cytoskeleton elastic behavior to the cell geometry, while treating all other subcellular components suspended in the cytoplasmic liquid (gel) as an incompressible fluid, is presented and validated by experimental results. The developed analytical-experimental methodology establishes a framework for quantifying the membrane-cytoskeleton elasticity of live cells. This capability may have immense implications in cell biology, particularly in studies seeking to establish correlations between membrane-cytoskeleton elasticity and cell disease, mortality, differentiation, and migration, and provide insight into cell infiltration through nonwoven fibrous scaffolds. The present method can be further extended to analyze membrane-cytoskeleton viscoelasticity, examine the role of other subcellular components (e.g., nucleus envelope) in cell elasticity, and elucidate the effects of mechanical stimuli on cell differentiation and motility. This is the first study to decouple the membrane-cytoskeleton elasticity from cell stiffness and introduce an effective approach for measuring the elastic modulus. The

  5. Biological effects of the electrostatic field: red blood cell-related alterations of oxidative processes in blood

    NASA Astrophysics Data System (ADS)

    Harutyunyan, Hayk A.; Sahakyan, Gohar V.

    2016-01-01

    The aim of this study was to determine activities of pro-/antioxidant enzymes, reactive oxygen species (ROS) content, and oxidative modification of proteins and lipids in red blood cells (RBCs) and blood plasma of rats exposed to electrostatic field (200 kV/m) during the short (1 h) and the long periods (6 day, 6 h daily). Short-term exposure was characterized by the increase of oxidatively damaged proteins in blood of rats. This was strongly expressed in RBC membranes. After long-term action, RBC content in peripheral blood was higher than in control ( P < 0.01) and the attenuation of prooxidant processes was shown.

  6. In situ single molecule imaging of cell membranes: linking basic nanotechniques to cell biology, immunology and medicine.

    PubMed

    Pi, Jiang; Jin, Hua; Yang, Fen; Chen, Zheng W; Cai, Jiye

    2014-11-07

    The cell membrane, which consists of a viscous phospholipid bilayer, different kinds of proteins and various nano/micrometer-sized domains, plays a very important role in ensuring the stability of the intracellular environment and the order of cellular signal transductions. Exploring the precise cell membrane structure and detailed functions of the biomolecules in a cell membrane would be helpful to understand the underlying mechanisms involved in cell membrane signal transductions, which could further benefit research into cell biology, immunology and medicine. The detection of membrane biomolecules at the single molecule level can provide some subtle information about the molecular structure and the functions of the cell membrane. In particular, information obtained about the molecular mechanisms and other information at the single molecule level are significantly different from that detected from a large amount of biomolecules at the large-scale through traditional techniques, and can thus provide a novel perspective for the study of cell membrane structures and functions. However, the precise investigations of membrane biomolecules prompts researchers to explore cell membranes at the single molecule level by the use of in situ imaging methods, as the exact conformation and functions of biomolecules are highly controlled by the native cellular environment. Recently, the in situ single molecule imaging of cell membranes has attracted increasing attention from cell biologists and immunologists. The size of biomolecules and their clusters on the cell surface are set at the nanoscale, which makes it mandatory to use high- and super-resolution imaging techniques to realize the in situ single molecule imaging of cell membranes. In the past few decades, some amazing imaging techniques and instruments with super resolution have been widely developed for molecule imaging, which can also be further employed for the in situ single molecule imaging of cell membranes. In

  7. In situ single molecule imaging of cell membranes: linking basic nanotechniques to cell biology, immunology and medicine

    NASA Astrophysics Data System (ADS)

    Pi, Jiang; Jin, Hua; Yang, Fen; Chen, Zheng W.; Cai, Jiye

    2014-10-01

    The cell membrane, which consists of a viscous phospholipid bilayer, different kinds of proteins and various nano/micrometer-sized domains, plays a very important role in ensuring the stability of the intracellular environment and the order of cellular signal transductions. Exploring the precise cell membrane structure and detailed functions of the biomolecules in a cell membrane would be helpful to understand the underlying mechanisms involved in cell membrane signal transductions, which could further benefit research into cell biology, immunology and medicine. The detection of membrane biomolecules at the single molecule level can provide some subtle information about the molecular structure and the functions of the cell membrane. In particular, information obtained about the molecular mechanisms and other information at the single molecule level are significantly different from that detected from a large amount of biomolecules at the large-scale through traditional techniques, and can thus provide a novel perspective for the study of cell membrane structures and functions. However, the precise investigations of membrane biomolecules prompts researchers to explore cell membranes at the single molecule level by the use of in situ imaging methods, as the exact conformation and functions of biomolecules are highly controlled by the native cellular environment. Recently, the in situ single molecule imaging of cell membranes has attracted increasing attention from cell biologists and immunologists. The size of biomolecules and their clusters on the cell surface are set at the nanoscale, which makes it mandatory to use high- and super-resolution imaging techniques to realize the in situ single molecule imaging of cell membranes. In the past few decades, some amazing imaging techniques and instruments with super resolution have been widely developed for molecule imaging, which can also be further employed for the in situ single molecule imaging of cell membranes. In

  8. Nanocellulose based asymmetric composite membrane for the multiple functions in cell encapsulation.

    PubMed

    Park, Minsung; Shin, Sungchul; Cheng, Jie; Hyun, Jinho

    2017-02-20

    We describe the nanocomposite membrane for cell encapsulation using nanocelluose hydrogels. One of the surfaces of bacterial cellulose (BC) pellicles was coated with collagen to enhance cell adhesion and the opposite side of the BC pellicles was coated with alginate to protect transplanted cells from immune rejection by the reduced pore size of the composite membrane. The morphology of nanocomposite membrane was observed by scanning electron microscopy and the permeability of the membrane was estimated by the release test using different molecular weights of polymer solution. The nanocomposite membrane was permeable to small molecules but impermeable to large molecules such as IgG antibodies inferring the potential use in cell implantation. In addition, the BC-based nanocomposite membrane showed a superior mechanical property due to the incorporation of compared with alginate membranes. The cells attached efficiently to the surface of BC composite membranes with a high level of cell viability as well as bioactivity. Cells grown on the BC composite membrane kit released dopamine freely to the medium through the membrane, which showed that the BC composite membrane would be a promising cell encapsulation material in implantation. Copyright © 2016 Elsevier Ltd. All rights reserved.

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

    NASA Astrophysics Data System (ADS)

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

    2012-02-01

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

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

    NASA Astrophysics Data System (ADS)

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

    2016-12-01

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

  11. Importance of balancing membrane and electrode water in anion exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

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

    2018-01-01

    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.

  12. Membrane hydraulic permeability changes during cooling of mammalian cells.

    PubMed

    Akhoondi, Maryam; Oldenhof, Harriëtte; Stoll, Christoph; Sieme, Harald; Wolkers, Willem F

    2011-03-01

    In order to predict optimal cooling rates for cryopreservation of cells, the cell-specific membrane hydraulic permeability and corresponding activation energy for water transport need to be experimentally determined. These parameters should preferably be determined at subzero temperatures in the presence of ice. There is, however, a lack of methods to study membrane properties of cells in the presence of ice. We have used Fourier transform infrared spectroscopy to study freezing-induced membrane dehydration of mouse embryonic fibroblast (3T3) cells and derived the subzero membrane hydraulic permeability and the activation energy for water transport from these data. Coulter counter measurements were used to determine the suprazero membrane hydraulic permeability parameters from cellular volume changes of cells exposed to osmotic stress. The activation energy for water transport in the ice phase is about three fold greater compared to that at suprazero temperatures. The membrane hydraulic permeability at 0 °C that was extrapolated from suprazero measurements is about five fold greater compared to that extrapolated from subzero measurements. This difference is likely due to a freezing-induced dehydration of the bound water around the phospholipid head groups. Using Fourier transform infrared spectroscopy, two distinct water transport processes, that of free and membrane bound water, can be identified during freezing with distinct activation energies. Dimethylsulfoxide, a widely used cryoprotective agent, did not prevent freezing-induced membrane dehydration but decreased the activation energy for water transport. Copyright © 2010 Elsevier B.V. All rights reserved.

  13. The structure and function of cell membranes studied by atomic force microscopy.

    PubMed

    Shi, Yan; Cai, Mingjun; Zhou, Lulu; Wang, Hongda

    2018-01-01

    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.

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

    NASA Astrophysics Data System (ADS)

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

    2015-12-01

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

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

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Haryadi,, E-mail: haryadi@polban.ac.id; Sugianto, D.; Ristopan, E.

    2015-12-29

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

  16. Nitric oxide pathology and therapeutics in sickle cell disease.

    PubMed

    Kim-Shapiro, Daniel B; Gladwin, Mark T

    2018-01-01

    Sickle cell disease is caused by a mutant form of hemoglobin that polymerizes under hypoxic conditions which leads to red blood cell (RBC) distortion, calcium-influx mediated RBC dehydration, increased RBC adhesivity, reduced RBC deformability, increased RBC fragility, and hemolysis. These impairments in RBC structure and function result in multifaceted downstream pathology including inflammation, endothelial cell activation, platelet and leukocyte activation and adhesion, and thrombosis, all of which contribute vascular occlusion and substantial morbidity and mortality. Hemoglobin released upon RBC hemolysis scavenges nitric oxide (NO) and generates reactive oxygen species (ROS) and thereby decreases bioavailability of this important signaling molecule. As the endothelium-derived relaxing factor, NO acts as a vasodilator and also decreases platelet, leukocyte, and endothelial cell activation. Thus, low NO bioavailability contributes to pathology in sickle cell disease and its restoration could serve as an effective treatment. Despite its promise, clinical trials based on restoring NO bioavailability have so far been mainly disappointing. However, particular "NO donating" agents such as nitrite, which unlike some other NO donors can improve sickle RBC properties, may yet prove effective.

  17. Measurements of Refractory Black Carbon (rBC) Aerosols in the McMurdo Dry Valleys, Antarctica

    NASA Astrophysics Data System (ADS)

    Khan, A. L.; McMeeking, G. R.; Lyons, W. B.; Schwarz, J. P.; Welch, K. A.; McKnight, D. M.

    2015-12-01

    Measurements of light absorbing particles in the boundary layer of the high southern latitudes are scarce. During the 2013-2014 austral summer field season refractory black carbon (rBC) aerosols were quantified by a single particle soot photometer (SP2) in the McMurdo Dry Valleys, Antarctica. The dark rBC particles absorb more radiation thereby increasing atmospheric heating, as well as reducing surface albedo and enhancing hydrologic melt when deposited on highly reflective surfaces such as snow and ice. Quantifying both local and long-range atmospheric transport of rBC to this region of a remote continent mostly covered by ice and snow would be useful in understanding meltwater generation as climate changes. Although the Dry Valleys are the largest ice-free region of Antarctica, they contain many alpine glaciers, some of which are fed from the East Antarctic Ice Sheet (EAIS). Continuous rBC measurements were collected at Lake Hoare Camp in the Taylor Valley for two months, along with shorter periods at more remote locations within the Dry Valleys. Conditions at the Lake Hoare Camp were dominated by up-valley winds from McMurdo Sound, however, winds also brought air down-valley from the EAIS polar plateau. Here we investigated periods dominated by both up and down-valley winds to explore differences in rBC concentrations, size distributions, and scattering properties. The average background rBC mass concentration was 1ng/m3, though concentrations as high as 50 ng/m3 were observed at times, likely due to local sources.

  18. Three-dimensional image and spatial spectrum analysis of behavior of small animal erythrocytes in optical tweezers

    NASA Astrophysics Data System (ADS)

    Chen, Hui Chi; Shen, Wen-Tai; Kong, Yu-Han; Chuang, Chun-Hao

    2008-02-01

    Because of the softness of membrane, erythrocytes (red blood cell, RBC) have different shapes while being immersed in buffer with different osmotic pressure. While affecting by different viruses and illnesses, RBC may change its shape, or its membrane may become rigid. Moreover, RBC will ford and stretch when it is trapped by optical tweezers. Therefore, the behaviors of RBC in optical tweezers raise more discussion. In this report, we set up an optical tweezers to trap RBC of small animals like feline and canine. By adding a long working distance objective to collect the side-viewing image, a 3-D image system was constructed to detect the motion of trapped RBC. To improve the image quality for side-view, an aperture and narrow glass plate were used. From the video of these images and their spatial spectrum, the shape of trapped RBC was studied.

  19. Proton conducting membrane for fuel cells

    DOEpatents

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

    2005-12-20

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

  20. Proton conducting membrane for fuel cells

    DOEpatents

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

    2007-03-27

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

  1. Shock Wave-Induced Damage and Poration in Eukaryotic Cell Membranes.

    PubMed

    López-Marín, Luz M; Millán-Chiu, Blanca E; Castaño-González, Karen; Aceves, Carmen; Fernández, Francisco; Varela-Echavarría, Alfredo; Loske, Achim M

    2017-02-01

    Shock waves are known to permeabilize eukaryotic cell membranes, which may be a powerful tool for a variety of drug delivery applications. However, the mechanisms involved in shock wave-mediated membrane permeabilization are still poorly understood. In this study, the effects on both the permeability and the ultrastructural features of two human cell lineages were investigated after the application of underwater shock waves in vitro. Scanning Electron Microscopy of cells derived from a human embryo kidney (HEK)-293 and Michigan Cancer Foundation (MCF)-7 cells, an immortalized culture derived from human breast adenocarcinoma, showed a small amount of microvilli (as compared to control cells), the presence of hole-like structures, and a decrease in cell size after shock wave exposure. Interestingly, these effects were accompanied by the permeabilization of acid and macromolecular dyes and gene transfection. Trypan blue exclusion assays indicated that cell membranes were porated during shock wave treatment but resealed after a few seconds. Deformations of the cell membrane lasted for at least 5 min, allowing their observation in fixed cells. For each cell line, different shock wave parameters were needed to achieve cell membrane poration. This difference was correlated to successful gene transfection by shock waves. Our results demonstrate, for the first time, that shock waves induce transient micro- and submicrosized deformations at the cell membrane, leading to cell transfection and cell survival. They also indicate that ultrastructural analyses of cell surfaces may constitute a useful way to match the use of shock waves to different cells and settings.

  2. Binding of /sup 18/F by cell membranes and cell walls of Streptococcus mutans

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yotis, W.W.; Zeb, M.; McNulty, J.

    1983-07-01

    The binding of /sup 18/F to isolated cell membranes and cell walls of Streptococcus mutans GS-5 or other bacteria was assayed. The attachment of /sup 18/F to these cell envelopes proceeded slowly and reached equilibrium within 60 min. /sup 18/F binding was stimulated by Ca/sup 2 +/ (1 mM). The binding of /sup 18/F to cellular components was dependent upon the pH, as well as the amount of /sup 18/F and dose of the binder employed. The binding of /sup 18/F by cell walls prepared from fluoride-sensitive and fluoride-resistant cells of S. salivarius and S. mutans did not differ significantly.more » 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 /sup 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 /sup 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 /sup 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 /sup 18/F per mg (dry weight). /sup 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 /sup 18/F binding by cell membranes and walls of oral flora.« less

  3. Membrane Organization and Cell Fusion During Mating in Fission Yeast Requires Multipass Membrane Protein Prm1

    PubMed Central

    Curto, M.-Ángeles; Sharifmoghadam, Mohammad Reza; Calpena, Eduardo; De León, Nagore; Hoya, Marta; Doncel, Cristina; Leatherwood, Janet; Valdivieso, M.-Henar

    2014-01-01

    The involvement of Schizosaccharomyces pombe prm1+ in cell fusion during mating and its relationship with other genes required for this process have been addressed. S. pombe prm1Δ mutant exhibits an almost complete blockade in cell fusion and an abnormal distribution of the plasma membrane and cell wall in the area of cell–cell interaction. The distribution of cellular envelopes is similar to that described for mutants devoid of the Fig1-related claudin-like Dni proteins; however, prm1+ and the dni+ genes act in different subpathways. Time-lapse analyses show that in the wild-type S. pombe strain, the distribution of phosphatidylserine in the cytoplasmic leaflet of the plasma membrane undergoes some modification before an opening is observed in the cross wall at the cell–cell contact region. In the prm1Δ mutant, this membrane modification does not take place, and the cross wall between the mating partners is not extensively degraded; plasma membrane forms invaginations and fingers that sometimes collapse/retract and that are sometimes strengthened by the synthesis of cell-wall material. Neither prm1Δ nor prm1Δ dniΔ zygotes lyse after cell–cell contact in medium containing and lacking calcium. Response to drugs that inhibit lipid synthesis or interfere with lipids is different in wild-type, prm1Δ, and dni1Δ strains, suggesting that membrane structure/organization/dynamics is different in all these strains and that Prm1p and the Dni proteins exert some functions required to guarantee correct membrane organization that are critical for cell fusion. PMID:24514900

  4. Pyroelectricity as a possible mechanism for cell membrane permeabilization.

    PubMed

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

    2018-02-01

    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.

  5. Compartmental Hollow Fiber Capillary Membrane–Based Bioreactor Technology for In Vitro Studies on Red Blood Cell Lineage Direction of Hematopoietic Stem Cells

    PubMed Central

    Housler, Greggory J.; Miki, Toshio; Schmelzer, Eva; Pekor, Christopher; Zhang, Xiaokui; Kang, Lin; Voskinarian-Berse, Vanessa; Abbot, Stewart; Zeilinger, Katrin

    2012-01-01

    Continuous production of red blood cells (RBCs) in an automated closed culture system using hematopoietic stem cell (HSC) progenitor cell populations is of interest for clinical application because of the high demand for blood transfusions. Previously, we introduced a four-compartment bioreactor that consisted of two bundles of hollow fiber microfiltration membranes for transport of culture medium (forming two medium compartments), interwoven with one bundle of hollow fiber membranes for transport of oxygen (O2), carbon dioxide (CO2), and other gases (forming one gas compartment). Small-scale prototypes were developed of the three-dimensional (3D) perfusion cell culture systems, which enable convection-based mass transfer and integral oxygenation in the cell compartment. CD34+ HSC were isolated from human cord blood units using a magnetic separation procedure. Cells were inoculated into 2- or 8-mL scaled-down versions of the previously designed 800-mL cell compartment devices and perfused with erythrocyte proliferation and differentiation medium. First, using the small-scale 2-mL analytical scale bioreactor, with an initial seeding density of 800,000 cells/mL, we demonstrated approximately 100-fold cell expansion and differentiation after 7 days of culture. An 8-mL laboratory-scale bioreactor was then used to show pseudocontinuous production by intermediately harvesting cells. Subsequently, we were able to use a model to demonstrate semicontinuous production with up to 14,288-fold expansion using seeding densities of 800,000 cells/mL. The down-scaled culture technology allows for expansion of CD34+ cells and stimulating these progenitors towards RBC lineage, expressing approximately 40% CD235+ and enucleation. The 3D perfusion technology provides an innovative tool for studies on RBC production, which is scalable. PMID:21933020

  6. The BSD2 ortholog in Chlamydomonas reinhardtii is a polysome-associated chaperone that co-migrates on sucrose gradients with the rbcL transcript encoding the Rubisco large subunit.

    PubMed

    Doron, Lior; Segal, Na'ama; Gibori, Hadas; Shapira, Michal

    2014-10-01

    The expression of the CO2 -fixation enzyme ribulose-bisphosphate carboxylase/oxygenase (Rubisco), which is affected by light, involves the cysteine-rich protein bundle-sheath defective-2 (BSD2) that was originally identified in maize bundle-sheath cells. We identified the BSD2 ortholog in Chlamydomonas reinhardtii as a small protein (17 kDa) localized to the chloroplast. The algal BSD2-ortholog contains four CXXCXGXG DnaJ-like elements, but lacks the other conserved domains of DnaJ. BSD2 co-migrated with the rbcL transcript on heavy polysomes, and both BSD2 and rbcL mRNA shifted to the lighter fractions under oxidizing conditions that repress the translation of the Rubisco large subunit (RbcL). This profile of co-migration supports the possibility that BSD2 is required for the de novo synthesis of RbcL. Furthermore, BSD2 co-migrated with the rbcL transcript in a C. reinhardtii premature-termination mutant that encodes the first 60 amino acids of RbcL. In both strains, BSD2 shared its migration profile with the rbcL transcript but not with psbA mRNA. The chaperone activity of BSD2 was exemplified by its ability to prevent the aggregation of both citrate synthase (CS) and RbcL in vitro following their chemical denaturation. This activity did not depend on the presence of the thiol groups on BSD2. In contrast, the activity of BSD2 in preventing the precipitation of reduced β-chains in vitro in the insulin turbidity assay was thiol-dependent. We conclude that BSD2 combines a chaperone 'holdase' function with the ability to interact with free thiols, with both activities being required to protect newly synthesized RbcL chains. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

  7. Membrane Purification Cell for Aluminum Recycling

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    David DeYoung; James Wiswall; Cong Wang

    2011-11-29

    Recycling mixed aluminum scrap usually requires adding primary aluminum to the scrap stream as a diluent to reduce the concentration of non-aluminum constituents used in aluminum alloys. Since primary aluminum production requires approximately 10 times more energy than melting scrap, the bulk of the energy and carbon dioxide emissions for recycling are associated with using primary aluminum as a diluent. Eliminating the need for using primary aluminum as a diluent would dramatically reduce energy requirements, decrease carbon dioxide emissions, and increase scrap utilization in recycling. Electrorefining can be used to extract pure aluminum from mixed scrap. Some example applications includemore » 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

  8. Detection of microparticles from human red blood cells by multiparametric flow cytometry

    PubMed Central

    Grisendi, Giulia; Finetti, Elena; Manganaro, Daniele; Cordova, Nicoletta; Montagnani, Giuliano; Spano, Carlotta; Prapa, Malvina; Guarneri, Valentina; Otsuru, Satoru; Horwitz, Edwin M.; Mari, Giorgio; Dominici, Massimo

    2015-01-01

    Background During storage, red blood cells (RBC) undergo chemical and biochemical changes referred to as “storage lesions”. These events determine the loss of RBC integrity, resulting in lysis and release of microparticles. There is growing evidence of the clinical importance of microparticles and their role in blood transfusion-related side effects and pathogen transmission. Flow cytometry is currently one of the most common techniques used to quantify and characterise microparticles. Here we propose multiparametric staining to monitor and quantify the dynamic release of microparticles by stored human RBC. Material and methods RBC units (n=10) were stored under blood bank conditions for up to 42 days. Samples were tested at different time points to detect microparticles and determine the haemolysis rate (HR%). Microparticles were identified by flow cytometry combining carboxyfluorescein diacetate succinimidyl ester (CFSE) dye, annexin V and anti-glycophorin A antibody. Results We demonstrated that CFSE can be successfully used to label closed vesicles with an intact membrane. The combination of CFSE and glycophorin A antibody was effective for monitoring and quantifying the dynamic release of microparticles from RBC during storage. Double staining with CFSE/glycophorin A was a more precise approach, increasing vesicle detection up to 4.7-fold vs the use of glycophorin A/annexin V alone. Moreover, at all the time points tested, we found a robust correlation (R=0.625; p=0.0001) between HR% and number of microparticles detected. Discussion Multiparametric staining, based on a combination of CFSE, glycophorin A antibody and annexin V, was able to detect, characterise and monitor the release of microparticles from RBC units during storage, providing a sensitive approach to labelling and identifying microparticles for transfusion medicine and, more broadly, for cell-based therapies. PMID:25369588

  9. Plasma membrane microorganization of LR73 multidrug-resistant cells revealed by FCS

    NASA Astrophysics Data System (ADS)

    Winckler, Pascale; Jaffiol, Rodolphe; Cailler, Aurélie; Morjani, Hamid; Jeannesson, Pierre; Deturche, Régis

    2011-03-01

    Tumoral cells could present a multidrug resistance (MDR) to chemotherapeutic treatments. This drug resistance would be associated to biomechanisms occurring at the plasma membrane level, involving modification of membrane fluidity, drug permeability, presence of microdomains (rafts, caveolae...), and membrane proteins overexpression such as Pglycoprotein. Fluorescence correlation spectroscopy (FCS) is the relevant method to investigate locally the fluidity of biological membranes through the lateral diffusion of a fluorescent membrane probe. Thus, we use FCS to monitor the plasma membrane local organization of LR73 carcinoma cells and three derived multidrug-resistant cancer cells lines. Measurements were conducted at the single cell level, which enabled us to get a detailed overview of the plasma membrane microviscosity distribution of each cell line studied. Moreover, we propose 2D diffusion simulation based on a Monte Carlo model to investigate the membrane organisation in terms of microdomains. This simulation allows us to relate the differences in the fluidity distributions with microorganization changes in plasma membrane of MDR cells.

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

    DOE Office of Scientific and Technical Information (OSTI.GOV)

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

    2016-04-01

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

  11. Membrane potential dynamics of grid cells

    PubMed Central

    Domnisoru, Cristina; Kinkhabwala, Amina A.; Tank, David W.

    2014-01-01

    During navigation, grid cells increase their spike rates in firing fields arranged on a strikingly regular triangular lattice, while their spike timing is often modulated by theta oscillations. Oscillatory interference models of grid cells predict theta amplitude modulations of membrane potential during firing field traversals, while competing attractor network models predict slow depolarizing ramps. Here, using in-vivo whole-cell recordings, we tested these models by directly measuring grid cell intracellular potentials in mice running along linear tracks in virtual reality. Grid cells had large and reproducible ramps of membrane potential depolarization that were the characteristic signature tightly correlated with firing fields. Grid cells also exhibited intracellular theta oscillations that influenced their spike timing. However, the properties of theta amplitude modulations were not consistent with the view that they determine firing field locations. Our results support cellular and network mechanisms in which grid fields are produced by slow ramps, as in attractor models, while theta oscillations control spike timing. PMID:23395984

  12. Trans-cis isomerization of lipophilic dyes probing membrane microviscosity in biological membranes and in live cells.

    PubMed

    Chmyrov, Volodymyr; Spielmann, Thiemo; Hevekerl, Heike; Widengren, Jerker

    2015-06-02

    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.

  13. Annexins are instrumental for efficient plasma membrane repair in cancer cells.

    PubMed

    Lauritzen, Stine Prehn; Boye, Theresa Louise; Nylandsted, Jesper

    2015-09-01

    Plasma membrane stress can cause damage to the plasma membrane, both when imposed by the extracellular environment and by enhanced oxidative stress. Cells cope with these injuries by rapidly activating their plasma membrane repair system, which is triggered by Ca(2+) influx at the wound site. The repair system is highly dynamic, depends on both lipid and protein components, and include cytoskeletal reorganization, membrane replacements, and membrane fusion events. Cancer cells experience enhanced membrane stress when navigating through dense extracellular matrix, which increases the frequency of membrane injuries. In addition, increased motility and oxidative stress further increase the risk of plasma membrane lesions. Cancer cells compensate by overexpressing Annexin proteins including Annexin A2 (ANXA2). Annexin family members can facilitate membrane fusion events and wound healing by binding to negatively charged phospholipids in the plasma membrane. Plasma membrane repair in cancer cells depends on ANXA2 protein, which is recruited to the wound site and forms a complex with the Ca(2+)-binding EF-hand protein S100A11. Here they regulate actin accumulation around the wound perimeter, which is required for wound closure. In this review, we will discuss the requirement for Annexins, S100 proteins and actin cytoskeleton in the plasma membrane repair response of cancer cells, which reveals a novel avenue for targeting metastatic cancers. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. "NEW MEMBRANE" FORMATION IN AMOEBA PROTEUS UPON INJURY OF INDIVIDUAL CELLS

    PubMed Central

    Szubinska, Barbara

    1971-01-01

    Changes in the plasma membrane complex following the injury of single cells of Amoeba proteus were examined with the electron microscope. Two types of injury were employed in this study; cells were either pinched ("cut") in half or speared with a glass microneedle, and quickly fixed. Speared cells, when fixed in the presence of the ruthenium violet (a derivative of ruthenium red), revealed the presence of an extra trilaminar structure outside of each cell. This structure, called the "new membrane," was separated from the plasma membrane complex by a distance of less than a micron. The trilaminar structure of the new membrane strikingly resembled the image of the plasma membrane in all cells examined, except for its increased width (30%). This new membrane appeared nearly to surround the injured amebae. Attempts were made to demonstrate the possible origin of the new membrane, its reality, and its sensitivity to calcium. Also, some evidence is shown concerning the role of the small dense droplets (100–1200 A in diameter) normally present in the cytoplasm of amebae. Their frequent contact with the plasma membrane of the cell as the result of injury is interpreted as indicating their involvement in the formation and expansion of the plasma membrane. PMID:4103955

  15. Selectivity of Direct Methanol Fuel Cell Membranes.

    PubMed

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

    2015-11-24

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

  16. Selectivity of Direct Methanol Fuel Cell Membranes

    PubMed Central

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

    2015-01-01

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

  17. Radiation Interaction with Therapeutic Drugs and Cell Membranes

    NASA Astrophysics Data System (ADS)

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

    2007-04-01

    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.

  18. Membrane catalyst layer for fuel cells

    DOEpatents

    Wilson, Mahlon S.

    1993-01-01

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

  19. Phospholipid composition of packed red blood cells and that of extracellular vesicles show a high resemblance and stability during storage.

    PubMed

    Laurén, Eva; Tigistu-Sahle, Feven; Valkonen, Sami; Westberg, Melissa; Valkeajärvi, Anne; Eronen, Juha; Siljander, Pia; Pettilä, Ville; Käkelä, Reijo; Laitinen, Saara; Kerkelä, Erja

    2018-01-01

    Red blood cells (RBCs) are stored up to 35-42days at 2-6°C in blood banks. During storage, the RBC membrane is challenged by energy depletion, decreasing pH, altered cation homeostasis, and oxidative stress, leading to several biochemical and morphological changes in RBCs and to shedding of extracellular vesicles (EVs) into the storage medium. These changes are collectively known as RBC storage lesions. EVs accumulate in stored RBC concentrates and are, thus, transfused into patients. The potency of EVs as bioactive effectors is largely acknowledged, and EVs in RBC concentrates are suspected to mediate some adverse effects of transfusion. Several studies have shown accumulation of lipid raft-associated proteins in RBC EVs during storage, whereas a comprehensive phospholipidomic study on RBCs and corresponding EVs during the clinical storage period is lacking. Our mass spectrometric and chromatographic study shows that RBCs maintain their major phospholipid (PL) content well during storage despite abundant vesiculation. The phospholipidomes were largely similar between RBCs and EVs. No accumulation of raft lipids in EVs was seen, suggesting that the primary mechanism of RBC vesiculation during storage might not be raft -based. Nonetheless, a slight tendency of EV PLs for shorter acyl chains was observed. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Red Blood Cell Mechanical Fragility Test for Clinical Research Applications.

    PubMed

    Ziegler, Luke A; Olia, Salim E; Kameneva, Marina V

    2017-07-01

    Red blood cell (RBC) susceptibility to mechanically induced hemolysis, or RBC mechanical fragility (MF), is an important parameter in the characterization of erythrocyte membrane health. The rocker bead test (RBT) and associated calculated mechanical fragility index (MFI) is a simple method for the assessment of RBC MF. Requiring a minimum of 15.5 mL of blood and necessitating adjustment of hematocrit (Ht) to a "standard" value (40%), the current RBT is not suitable for use in most studies involving human subjects. To address these limitations, we propose a 6.5 mL reduced volume RBT and corresponding modified MFI (MMFI) that does not require prior Ht adjustment. This new method was assessed for i) correlation to the existing text, ii) to quantify the effect of Ht on MFI, and iii) validation by reexamining the protective effect of plasma proteins on RBC MF. The reduced volume RBT strongly correlated (r = 0.941) with the established large volume RBT at matched Hts, and an equation was developed to calculate MMFI: a numerical estimation (R 2  = 0.923) of MFI if performed with the reduced volume RBT at "standard" (40%) Ht. An inversely proportional relationship was found between plasma protein concentration and RBC MF using the MMFI-reduced volume method, supporting previous literature findings. The new reduced volume RBT and modified MFI will allow for the measurement of RBC MF in clinical and preclinical studies involving humans or small animals. © 2017 International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.

  1. Radiation pressure on a biconcave human Red Blood Cell and the resulting deformation in a pair of parallel optical traps.

    PubMed

    Liao, Guan-Bo; Chen, Yin-Quan; Bareil, Paul B; Sheng, Yunlong; Chiou, Arthur; Chang, Ming-Shien

    2014-10-01

    We calculated the three-dimensional optical stress distribution and the resulting deformation on a biconcave human red blood cell (RBC) in a pair of parallel optical trap. We assumed a Gaussian intensity distribution with a spherical wavefront for each trapping beam and calculated the optical stress from the momentum transfer associated with the reflection and refraction of the incident photons at each interface. The RBC was modelled as a biconcave thin elastic membrane with uniform elasticity and a uniform thickness of 0.25 μm. The resulting cell deformation was determined from the optical stress distribution by finite element software, Comsol Structure Mechanics Module, with Young's modulus (E) as a fitting parameter in order to fit the theoretical results for cell elongation to our experimental data. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Membrane Tension Inhibits Rapid and Slow Endocytosis in Secretory Cells.

    PubMed

    Wu, Xin-Sheng; Elias, Sharon; Liu, Huisheng; Heureaux, Johanna; Wen, Peter J; Liu, Allen P; Kozlov, Michael M; Wu, Ling-Gang

    2017-12-05

    Endocytosis generates spherical or ellipsoid-like vesicles from the plasma membrane, which recycles vesicles that fuse with the plasma member during exocytosis in neurons and endocrine secretory cells. Although tension in the plasma membrane is generally considered to be an important factor in regulating endocytosis, whether membrane tension inhibits or facilitates endocytosis remains debated in the endocytosis field, and has been rarely studied for vesicular endocytosis in secretory cells. Here we report that increasing membrane tension by adjusting osmolarity inhibited both the rapid (a few seconds) and slow (tens of seconds) endocytosis in calyx-type nerve terminals containing conventional active zones and in neuroendocrine chromaffin cells. We address the mechanism of this phenomenon by computational modeling of the energy barrier that the system must overcome at the stage of membrane budding by an assembling protein coat. We show that this barrier grows with increasing tension, which may slow down or prevent membrane budding. These results suggest that in live secretory cells, membrane tension exerts inhibitory action on endocytosis. Published by Elsevier Inc.

  3. Vitamin E and vitamin E-quinone levels in red blood cells and plasma of newborn infants and their mothers.

    PubMed

    Jain, S K; Wise, R; Bocchini, J J

    1996-02-01

    Vitamin E is a physiological antioxidant and protects cell membranes from oxidative damage. This study has determined whether vitamin E level in RBC of newborns has any relationship with its level in their mothers. We have also examined levels of vitamin E and vitamin E-quinone, an oxidized product of vitamin E, in paired samples of red blood cells (RBC) and plasma of newborns and their mothers. Blood was collected from 26 mothers and their full-term placental cords at delivery. Vitamin E and vitamin E-quinone levels were determined in RBC and plasma by HPLC. Newborn-plasma had significantly lower vitamin E levels compared with maternal-plasma both when expressed as nmole/ml (5.5+/-0.4 vs 26.1+/-1.1, p = 0.0001) or nmole/mumole total lipids (1.9+/-0.1 vs 2.6+/-0.1, p = 0.0001). Vitamin E level in the newborn-RBC was similar to that of maternal-RBC when expressed as nmole/ml packed cells (2.77+/-0.14 vs 2.95+/-0.13), but was significantly lower when expressed as nmole/mumole total lipids (0.56+/-0.03 vs 0.64+/-0.04, p = 0.03) from that of maternal-RBC. Vitamin E-quinone levels are significantly elevated in newborns compared with their mothers both in RBC (29.4+/-2.1 vs 24.1+/-1.2, p = 0.04) and plasma (39.9+/-5.3 vs 25.3+/-4.2, p = 0.006) when expressed as nmole/mmole total lipids but not when expressed as nmole/ml. There was a significant correlation of vitamin E between newborn-plasma and newborn-RBC (r = 0.65, p = 0.0002 for nmole per ml packed RBC;r = 0.63, p = 0.0007 for nmole per mumole total lipids). The relationship between maternal plasma and newborn plasma was significant when vitamin E was normalized with nmole/mumole total lipid (r = 0.54, p = 0.007 but not when expressed as nmole/ml (r = 0.09, p = 0.64). However, vitamin E in the RBC of maternal and newborn had significant correlation when expressed as per ml packed cells (r = 0.61, p = 0.001) and per total lipid (r = 0.46, p = 0.02). There was no relationship of vitamin E-quinone levels between RBC and

  4. Red blood cells as modulators of T cell growth and survival.

    PubMed

    Arosa, Fernando A; Pereira, Carlos F; Fonseca, Ana M

    2004-01-01

    T cell homeostasis is largely controlled by a balance between cell death and survival and anomalies in either process account for a number of diseases linked to excessive or faulty T cell growth. Yet, the influence that cells outside the immunological system have on these processes has only recently received attention. Accumulated evidence indicate that homeostasis of the CD4+ and CD8+ T cell pools is highly dynamic and regulated by signals delivered by cells and molecules present in the different internal microenvironments. The major function of red blood cells (RBC) is generally considered to be oxygen and carbon dioxide transport. In recent years, however, RBC have been implicated in the regulation of basic physiological processes, from vascular contraction and platelet aggregation to T cell growth and survival. Regulation of T cell survival by RBC may influence the response of selected subsets of T cells to internal or external stimuli and may help explaining the immunomodulatory activities of red blood cells. By interfering in the balance between death and survival RBC become potential tools that can be manipulated to improve or reverse pathological situations characterized by anomalies in the control of T cell growth.

  5. Cell-free synthesis of membrane subunits of ATP synthase in phospholipid bicelles: NMR shows subunit a fold similar to the protein in the cell membrane

    PubMed Central

    Uhlemann, Eva-Maria E; Pierson, Hannah E; Fillingame, Robert H; Dmitriev, Oleg Y

    2012-01-01

    NMR structure determination of large membrane proteins is hampered by broad spectral lines, overlap, and ambiguity of signal assignment. Chemical shift and NOE assignment can be facilitated by amino acid selective isotope labeling in cell-free protein synthesis system. However, many biological detergents are incompatible with the cell-free synthesis, and membrane proteins often have to be synthesized in an insoluble form. We report cell-free synthesis of subunits a and c of the proton channel of Escherichia coli ATP synthase in a soluble form in a mixture of phosphatidylcholine derivatives. In comparison, subunit a was purified from the cell-free system and from the bacterial cell membranes. NMR spectra of both preparations were similar, indicating that our procedure for cell-free synthesis produces protein structurally similar to that prepared from the cell membranes. PMID:22162071

  6. Rigid proteins and softening of biological membranes-with application to HIV-induced cell membrane softening.

    PubMed

    Agrawal, Himani; Zelisko, Matthew; Liu, Liping; Sharma, Pradeep

    2016-05-06

    A key step in the HIV-infection process is the fusion of the virion membrane with the target cell membrane and the concomitant transfer of the viral RNA. Experimental evidence suggests that the fusion is preceded by considerable elastic softening of the cell membranes due to the insertion of fusion peptide in the membrane. What are the mechanisms underpinning the elastic softening of the membrane upon peptide insertion? A broader question may be posed: insertion of rigid proteins in soft membranes ought to stiffen the membranes not soften them. However, experimental observations perplexingly appear to show that rigid proteins may either soften or harden membranes even though conventional wisdom only suggests stiffening. In this work, we argue that regarding proteins as merely non-specific rigid inclusions is flawed, and each protein has a unique mechanical signature dictated by its specific interfacial coupling to the surrounding membrane. Predicated on this hypothesis, we have carried out atomistic simulations to investigate peptide-membrane interactions. Together with a continuum model, we reconcile contrasting experimental data in the literature including the case of HIV-fusion peptide induced softening. We conclude that the structural rearrangements of the lipids around the inclusions cause the softening or stiffening of the biological membranes.

  7. Hydrogen-oxygen proton-exchange membrane fuel cells and electrolyzers

    NASA Technical Reports Server (NTRS)

    Baldwin, R.; Pham, M.; Leonida, A.; Mcelroy, J.; Nalette, T.

    1989-01-01

    Hydrogen-oxygen solid polymer electrolyte (SPE) fuel cells and SPE electrolyzers (products of Hamilton Standard) both use a Proton-Exchange Membrane (PEM) as the sole electrolyte. These solid electrolyte devices have been under continuous development for over 30 years. This experience has resulted in a demonstrated ten-year SPE cell life capability under load conditions. Ultimate life of PEM fuel cells and electrolyzers is primarily related to the chemical stability of the membrane. For perfluorocarbon proton exchange membranes an accurate measure of the membrane stability is the fluoride loss rate. Millions of cell hours have contributed to establishing a relationship between fluoride loss rates and average expected ultimate cell life. This relationship is shown. Several features have been introduced into SPE fuel cells and SPE electrolyzers such that applications requiring greater than or equal to 100,000 hours of life can be considered. Equally important as the ultimate life is the voltage stability of hydrogen-oxygen fuel cells and electrolyzers. Here again the features of SPE fuel cells and SPE electrolyzers have shown a cell voltage stability in the order of 1 microvolt per hour. That level of stability has been demonstrated for tens of thousands of hours in SPE fuel cells at up to 500 amps per square foot (ASF) current density.

  8. Red blood cell transfusion and increased length of storage are not associated with deep vein thrombosis in medical and surgical critically ill patients: a prospective observational cohort study

    PubMed Central

    2011-01-01

    Introduction With prolonged storage times, cell membranes of red blood cells (RBCs) undergo morphologic and biochemical changes, termed 'RBC storage lesions'. Storage lesions may promote inflammation and thrombophilia when transfused. In trauma patients, RBC transfusion was an independent risk factor for deep vein thrombosis (DVT), specifically when RBC units were stored > 21 days or when 5 or more units were transfused. The objective of this study was to determine if RBC transfusions or RBC storage age predicts incident DVT in medical or surgical intensive care unit (ICU) patients. Methods Using a database which prospectively enrolled 261 patients over the course of 1 year with an ICU stay of at least 3 days, we analyzed DVT and RBC transfusions using Cox proportional hazards regression. Transfusions were analyzed with 4 thresholds, and storage age using 3 thresholds. DVTs were identified by twice-weekly proximal leg ultrasounds. Multivariable analyses were adjusted for 4 significant DVT predictors in this population (venous thrombosis history, chronic dialysis, platelet transfusion and inotropes). Results Of 261 patients, 126 (48.3%) had at least 1 RBC transfusion; 46.8% of those transfused had ≥ 5 units in ICU. Patients receiving RBCs were older (68.8 vs 64.1 years), more likely to be female (47.0 vs 30.7), sicker (APACHEII 26.8 vs 24.4), and more likely to be surgical (21.4 vs 8.9) (P < 0.05). The total number of RBCs per patient was 1-64, mean was 6.3 (SD 7.5), median was 4 (IQR 2,8). In univariate analyses, there was no association between DVT and RBC exposure (1 day earlier, 3 days earlier, 7 days earlier, or ever) or RBC storage (≤ 7 or > 7 days, ≤ 14 or > 14 days, ≤ 21 or > 21 days). Among patients transfused, no multivariable analyses showed that RBC transfusion or storage age predicted DVT. Trends were counter to the hypothesis (e.g., RBC storage for ≤ 7 days suggested a higher DVT risk compared to > 7 days (HR 5.3; 95% CI 1

  9. Aluminum and temperature alteration of cell membrane permeability of Quercus rubra

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Junping Chen; Sucoff, E.I.; Stadelmann, E.J.

    1991-06-01

    Al toxicity is the major factor limiting plant growth in acid soils. This report extends research on Al-induced changes in membrane behavior of intact root cortex cells of Northern red oak (Quercus rubra). Membrane permeability was determined by the plasmometric method for individual intact cells at temperatures from 2 or 4 to 35 C. Al (0.37 millimolar) significantly increased membrane permeability to urea and monoethyl urea and decreased permeability to water. Al significantly altered the activation energy required to transport water (+ 32%), urea (+ 9%), and monoethyl urea ({minus}7%) across cell membranes. Above 9 C, Al increased the lipidmore » partiality of the cell membranes; below 7 C, Al decreased it. Al narrowed by 6 C the temperature range over which plasmolysis occurred without membrane damage. These changes in membrane behavior are explainable if Al reduced membrane lipid fluidity and kink frequency and increases packing density and the occurrence of straight lipid chains.« less

  10. Advanced membrane electrode assemblies for fuel cells

    DOEpatents

    Kim, Yu Seung; Pivovar, Bryan S.

    2012-07-24

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

  11. Advanced membrane electrode assemblies for fuel cells

    DOEpatents

    Kim, Yu Seung; Pivovar, Bryan S

    2014-02-25

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

  12. Modeling a Membrane: Using Engineering Design to Simulate Cell Transport Processes

    ERIC Educational Resources Information Center

    Mason, Kevin; Evans, Brian

    2017-01-01

    The "plasma membrane," which controls what comes in and goes out of a cell, is integral to maintaining homeostasis. Cell transport of small molecules across the cell membrane happens in several different ways. Some small, nonpolar molecules cross the plasma membrane along the concentration gradient directly through the "phospholipid…

  13. Transfusion of irradiated red blood cell units with a potassium adsorption filter: A randomized controlled trial.

    PubMed

    Cid, Joan; Villegas, Vanessa; Carbassé, Gloria; Alba, Cristina; Perea, Dolores; Lozano, Miguel

    2016-05-01

    The irradiation of red blood cells (RBCs) causes damage of the RBC membrane with increased potassium (K) leak during storage compared with nonirradiated RBC units of similar age. A previous in vitro study showed a mean reduction of K of 94 ± 5% with a potassium adsorption filter (PAF). A prospective, single-center, nonblinded, randomized controlled trial (RCT) was designed to evaluate the safety and efficacy of transfusing irradiated RBC units with the PAF. Patients 18 years of age or older who received irradiated RBC units due to chemotherapy-induced anemia were randomly assigned to receive irradiated RBC units with the PAF (PAF group) or with the standard blood infusion set (control group). Primary outcome measures were safety and efficacy of the PAF (absolute change in hemoglobin [Hb] and K, respectively, in patient's blood values after transfusing the irradiated RBC units with or without the PAF). A total of 63 irradiated RBC units were transfused to 17 patients in the control group, and a total of 56 irradiated RBC units were transfused to 13 patients in the PAF group. The absolute change of Hb (9.3 ± 6.3 g/L vs. 8.1 ± 5.8 g/L; p = 0.3) and the absolute change of K (-0.01 ± 0.4 mmol/L vs. -0.01 ± 0.3 mmol/L; p = 0.2) were comparable between the two groups of the trial. The transfusion of 1 irradiated RBC unit with the PAF was as safe and efficacious as the transfusion of 1 irradiated RBC unit with the standard blood infusion set in patients with chemotherapy-induced anemia. © 2016 AABB.

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

    PubMed Central

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

    2011-01-01

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

  15. Effects of 5-hydroxymethyl-2-furfural on the volume and membrane permeability of red blood cells from patients with sickle cell disease

    PubMed Central

    Hannemann, Anke; Cytlak, Urszula M; Rees, David C; Tewari, Sanjay; Gibson, John S

    2014-01-01

    The heterocyclic aldehyde 5-hydroxymethyl-2-furfural (5HMF) interacts allosterically with the abnormal form of haemoglobin (Hb), HbS, in red blood cells (RBCs) from patients with sickle cell disease (SCD), thereby increasing oxygen affinity and decreasing HbS polymerization and RBC sickling during hypoxia. We hypothesized that should 5HMF also inhibit the main cation pathways implicated in the dehydration of RBCs from SCD patients – the deoxygenation-induced cation pathway (Psickle), the Ca2+-activated K+ channel (the Gardos channel) and the K+–Cl− cotransporter (KCC) – it would have a synergistic effect in protection against sickling, directly through interacting with HbS, and indirectly through maintaining hydration and reducing [HbS]. This study was therefore designed to investigate the effects of 5HMF on RBC volume and K+ permeability in vitro. 5HMF markedly reduced the deoxygenation-induced dehydration of RBCs whether in response to maintained deoxygenation or to cyclical deoxygenation/re-oxygenation. 5HMF was found to inhibit Psickle, an effect which correlated with its effects on sickling. Deoxygenation-induced activation of the Gardos channel and exposure of phosphatidylserine were also inhibited, probably indirectly via reduced entry of Ca2+ through the Psickle pathway. Effects of 5HMF on KCC were more modest with a slight inhibition in N-ethylmaleimide (NEM, 1 mm)-treated RBCs and stimulation in RBCs untreated with NEM. These findings support the hypothesis that 5HMF may also be beneficial through effects on RBC ion and water homeostasis. PMID:25015917

  16. Effects of 5-hydroxymethyl-2-furfural on the volume and membrane permeability of red blood cells from patients with sickle cell disease.

    PubMed

    Hannemann, Anke; Cytlak, Urszula M; Rees, David C; Tewari, Sanjay; Gibson, John S

    2014-09-15

    The heterocyclic aldehyde 5-hydroxymethyl-2-furfural (5HMF) interacts allosterically with the abnormal form of haemoglobin (Hb), HbS, in red blood cells (RBCs) from patients with sickle cell disease (SCD), thereby increasing oxygen affinity and decreasing HbS polymerization and RBC sickling during hypoxia. We hypothesized that should 5HMF also inhibit the main cation pathways implicated in the dehydration of RBCs from SCD patients - the deoxygenation-induced cation pathway (Psickle), the Ca(2+)-activated K(+) channel (the Gardos channel) and the K(+)-Cl(-) cotransporter (KCC) - it would have a synergistic effect in protection against sickling, directly through interacting with HbS, and indirectly through maintaining hydration and reducing [HbS]. This study was therefore designed to investigate the effects of 5HMF on RBC volume and K(+) permeability in vitro. 5HMF markedly reduced the deoxygenation-induced dehydration of RBCs whether in response to maintained deoxygenation or to cyclical deoxygenation/re-oxygenation. 5HMF was found to inhibit Psickle, an effect which correlated with its effects on sickling. Deoxygenation-induced activation of the Gardos channel and exposure of phosphatidylserine were also inhibited, probably indirectly via reduced entry of Ca(2+) through the Psickle pathway. Effects of 5HMF on KCC were more modest with a slight inhibition in N-ethylmaleimide (NEM, 1 mm)-treated RBCs and stimulation in RBCs untreated with NEM. These findings support the hypothesis that 5HMF may also be beneficial through effects on RBC ion and water homeostasis. © 2014 The Authors. The Journal of Physiology © 2014 The Physiological Society.

  17. Cytotopographical specialization of enzymatically isolated rabbit retinal Müller (glial) cells: K+ conductivity of the cell membrane.

    PubMed

    Reichenbach, A; Eberhardt, W

    1988-01-01

    Müller (radial glial) cells were isolated from rabbit retinae by means of papaine and mechanical dissociation. Regional membrane properties of these cells were studied by intracellular microelectrode recordings of potential responses to local application of high K+ solutions. When different parts of the cell membrane were exposed to high K+, the amplitude of the depolarizing responses varied greatly, indicating a strong regional specialization of the membrane properties. Using morphometrical data of isolated rabbit Müller cells, and a simple circuit model, we calculated the endfoot membrane to constitute more than 80% of the total K+ conductance of the cell; the specific resistivity of the endfoot membrane was about 400 omega cm2, i.e., more than 40 times less than that of the membrane of the vitread process, which is immediately adjacent. This kind of regional membrane specialization seems to be optimized in respect to the Müller cells' ability to carry spatial buffering K+ currents.

  18. Evaluation of the RBC Pig-a and PIGRET assays using single doses of hydroxyurea and melphalan in rats.

    PubMed

    Adachi, Hideki; Uematsu, Yasuaki; Yamada, Toru

    2016-11-15

    To evaluate the suitability of the rat Pig-a assay on reticulocytes (PIGRET assay) as a short-term test, red blood cell (RBC) Pig-a and PIGRET assays after single doses with hydroxyurea (HU) and melphalan (L-PAM) were conducted and the results of both assays were compared. HU was administered once orally to male SD rats at 250, 500 and 1000mg/kg, and both assays were conducted using peripheral blood withdrawn from the jugular vein at 1, 2 and 4 weeks after dosing. L-PAM was administered at 1.25, 2.5 and 5mg/kg in the same manner. L-PAM produced significant dose-dependent increases in mutant frequencies in the PIGRET assay after single oral doses, but did not produce dose-dependent increases in mutant frequencies in the RBC Pig-a assay. These results suggest that the PIGRET assay is more sensitive for the evaluation of the mutagenic potential of L-PAM than the RBC Pig-a assay. In contrast, HU, a clastogenic but not DNA-reactive compound, gave negative results in both assays. The results with these 2 chemicals indicate that the single-dose PIGRET assay in rats has the potential to properly detect DNA-reactive compounds that directly cause DNA damage in a short-term assay. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Aging of cell membranes: facts and theories.

    PubMed

    Zs-Nagy, Imre

    2014-01-01

    This chapter is intended to outline the main results of a research trend realized by the author during the last 45 years, focused on the main role played by the cell membrane in the aging process. It is a very wide field; therefore, the reader cannot expect in this limited space a detailed description, but will be given a wide, interdisciplinary insight into the main facts and theories regarding cellular aging. The central idea described here is the concept called the membrane hypothesis of aging (MHA). The history, the chemical roots, physicochemical facts, biophysical processes, as well as the obligatory biochemical consequences are all touched in by indicating the most important sources of detailed knowledge for those who are more interested in the basic biology of the aging process. This chapter covers also the available anti-aging interventions on the cell membrane by means of the centrophenoxine treatment based on the MHA. It also briefly interprets the possibilities of a just developing anti-aging method by using the recombinant human growth hormone, essential basis of which is the species specificity, and the general presence of receptors of this hormone in the plasma membrane of all types of cells.

  20. Effect of Thuya occidentalis on the labeling of red blood cells and plasma proteins with technetium-99m.

    PubMed Central

    Oliveira, J. F.; Braga, A. C.; Avila, A. S.; Fonseca, L. M.; Gutfilen, B.; Bernardo-Filho, M.

    1996-01-01

    Thuya occidentalis is used in popular medicine in the treatment of condyloma and has antibacterial action. Red blood cells (RBC) labeled with technetium-99m (99mTc) are used for several evaluations in nuclear medicine. This labeling depends on a reducing agent, usually stannous ion. Any drug which alters the labeling of the tracer could be expected to modify the disposition of the radiopharmaceutical. We have evaluated the influence of T. occidentalis extract on the labeling of RBC and plasma proteins with 99mTc. Blood was withdrawn and incubated with T. occidentalis (0.25; 2.5; 20.5; and 34.1 percent v/v). Stannous chloride (1.2 micrograms/ml) was added and then 99mTc was added. Plasma (P) and blood cells (BC) were isolated, also precipitated with trichloroacetic acid and soluble (SF) and insoluble fractions (IF) separated. The analysis of the results shows that there is a decrease in radioactivity (from 97.64 to 75.89 percent) in BC with 34.1 percent of the drug. In the labeling process of RBC with 99mTc, the stannous and pertechnetate ions pass through the membrane, so we suggest that the T. occidentalis effect can be explained (i) by an inhibition of the transport of these ions, (ii) by damage in membrane, (iii) by competition with the cited ions for the same binding sites, or (iv) by possible generation of reactive oxygen species that could oxidize the stannous ion. PMID:9436292

  1. Effect of Thuya occidentalis on the labeling of red blood cells and plasma proteins with technetium-99m.

    PubMed

    Oliveira, J F; Braga, A C; Avila, A S; Fonseca, L M; Gutfilen, B; Bernardo-Filho, M

    1996-01-01

    Thuya occidentalis is used in popular medicine in the treatment of condyloma and has antibacterial action. Red blood cells (RBC) labeled with technetium-99m (99mTc) are used for several evaluations in nuclear medicine. This labeling depends on a reducing agent, usually stannous ion. Any drug which alters the labeling of the tracer could be expected to modify the disposition of the radiopharmaceutical. We have evaluated the influence of T. occidentalis extract on the labeling of RBC and plasma proteins with 99mTc. Blood was withdrawn and incubated with T. occidentalis (0.25; 2.5; 20.5; and 34.1 percent v/v). Stannous chloride (1.2 micrograms/ml) was added and then 99mTc was added. Plasma (P) and blood cells (BC) were isolated, also precipitated with trichloroacetic acid and soluble (SF) and insoluble fractions (IF) separated. The analysis of the results shows that there is a decrease in radioactivity (from 97.64 to 75.89 percent) in BC with 34.1 percent of the drug. In the labeling process of RBC with 99mTc, the stannous and pertechnetate ions pass through the membrane, so we suggest that the T. occidentalis effect can be explained (i) by an inhibition of the transport of these ions, (ii) by damage in membrane, (iii) by competition with the cited ions for the same binding sites, or (iv) by possible generation of reactive oxygen species that could oxidize the stannous ion.

  2. S4(13)-PV cell-penetrating peptide induces physical and morphological changes in membrane-mimetic lipid systems and cell membranes: implications for cell internalization.

    PubMed

    Cardoso, Ana M S; Trabulo, Sara; Cardoso, Ana L; Lorents, Annely; Morais, Catarina M; Gomes, Paula; Nunes, Cláudia; Lúcio, Marlene; Reis, Salette; Padari, Kärt; Pooga, Margus; Pedroso de Lima, Maria C; Jurado, Amália S

    2012-03-01

    The present work aims to gain insights into the role of peptide-lipid interactions in the mechanisms of cellular internalization and endosomal escape of the S4(13)-PV cell-penetrating peptide, which has been successfully used in our laboratory as a nucleic acid delivery system. A S4(13)-PV analogue, S4(13)-PVscr, displaying a scrambled amino acid sequence, deficient cell internalization and drug delivery inability, was used in this study for comparative purposes. Differential scanning calorimetry, fluorescence polarization and X-ray diffraction at small and wide angles techniques showed that both peptides interacted with anionic membranes composed of phosphatidylglycerol or a mixture of this lipid with phosphatidylethanolamine, increasing the lipid order, shifting the phase transition to higher temperatures and raising the correlation length between the bilayers. However, S4(13)-PVscr, in contrast to the wild-type peptide, did not promote lipid domain segregation and induced the formation of an inverted hexagonal lipid phase instead of a cubic phase in the lipid systems assayed. Electron microscopy showed that, as opposed to S4(13)-PVscr, the wild-type peptide induced the formation of a non-lamellar organization in membranes of HeLa cells. We concluded that lateral phase separation and destabilization of membrane lamellar structure without compromising membrane integrity are on the basis of the lipid-driven and receptor-independent mechanism of cell entry of S4(13)-PV peptide. Overall, our results can contribute to a better understanding of the role of peptide-lipid interactions in the mechanisms of cell-penetrating peptide membrane translocation, helping in the future design of more efficient cell-penetrating peptide-based drug delivery systems. Copyright © 2011 Elsevier B.V. All rights reserved.

  3. 3D visualization of membrane failures in fuel cells

    NASA Astrophysics Data System (ADS)

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

    2017-03-01

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

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

    PubMed

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

    2012-12-01

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

  5. Membrane Protein Mobility and Orientation Preserved in Supported Bilayers Created Directly from Cell Plasma Membrane Blebs.

    PubMed

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

    2016-03-29

    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

  6. Rupturing Giant Plasma Membrane Vesicles to Form Micron-sized Supported Cell Plasma Membranes with Native Transmembrane Proteins.

    PubMed

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

    2017-11-09

    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.

  7. Discriminatory power of rbcL barcode locus for authentication of some of United Arab Emirates (UAE) native plants.

    PubMed

    Maloukh, Lina; Kumarappan, Alagappan; Jarrar, Mohammad; Salehi, Jawad; El-Wakil, Houssam; Rajya Lakshmi, T V

    2017-06-01

    DNA barcoding of United Arab Emirates (UAE) native plants is of high practical and scientific value as the plants adapt to very harsh environmental conditions that challenge their identification. Fifty-one plant species belonged to 22 families, 2 monocots, and 20 eudicots; a maximum number of species being legumes and grasses were collected. To authenticate the morphological identification of the wild plant taxa, rbcL and matK regions were used in the study. The primer universality and discriminatory power of rbcL is 100%, while it is 35% for matK locus for these plant species. The sequences were submitted to GenBank; accession numbers were obtained for all the rbcL sequences and for 6 of matK sequences. We suggest rbcL as a promising barcode locus for the tested group of 51 plants. In the present study, an inexpensive, simple method of identification of rare desert plant taxa through rbcL barcode is being reported.

  8. Bacillus subtilis MreB orthologs self-organize into filamentous structures underneath the cell membrane in a heterologous cell system.

    PubMed

    Dempwolff, Felix; Reimold, Christian; Reth, Michael; Graumann, Peter L

    2011-01-01

    Actin-like bacterial cytoskeletal element MreB has been shown to be essential for the maintenance of rod cell shape in many bacteria. MreB forms rapidly remodelling helical filaments underneath the cell membrane in Bacillus subtilis and in other bacterial cells, and co-localizes with its two paralogs, Mbl and MreBH. We show that MreB localizes as dynamic bundles of filaments underneath the cell membrane in Drosophila S2 Schneider cells, which become highly stable when the ATPase motif in MreB is modified. In agreement with ATP-dependent filament formation, the depletion of ATP in the cells lead to rapid dissociation of MreB filaments. Extended induction of MreB resulted in the formation of membrane protrusions, showing that like actin, MreB can exert force against the cell membrane. Mbl also formed membrane associated filaments, while MreBH formed filaments within the cytosol. When co-expressed, MreB, Mbl and MreBH built up mixed filaments underneath the cell membrane. Membrane protein RodZ localized to endosomes in S2 cells, but localized to the cell membrane when co-expressed with Mbl, showing that bacterial MreB/Mbl structures can recruit a protein to the cell membrane. Thus, MreB paralogs form a self-organizing and dynamic filamentous scaffold underneath the membrane that is able to recruit other proteins to the cell surface.

  9. Bacillus subtilis MreB Orthologs Self-Organize into Filamentous Structures underneath the Cell Membrane in a Heterologous Cell System

    PubMed Central

    Dempwolff, Felix; Reimold, Christian; Reth, Michael; Graumann, Peter L.

    2011-01-01

    Actin-like bacterial cytoskeletal element MreB has been shown to be essential for the maintenance of rod cell shape in many bacteria. MreB forms rapidly remodelling helical filaments underneath the cell membrane in Bacillus subtilis and in other bacterial cells, and co-localizes with its two paralogs, Mbl and MreBH. We show that MreB localizes as dynamic bundles of filaments underneath the cell membrane in Drosophila S2 Schneider cells, which become highly stable when the ATPase motif in MreB is modified. In agreement with ATP-dependent filament formation, the depletion of ATP in the cells lead to rapid dissociation of MreB filaments. Extended induction of MreB resulted in the formation of membrane protrusions, showing that like actin, MreB can exert force against the cell membrane. Mbl also formed membrane associated filaments, while MreBH formed filaments within the cytosol. When co-expressed, MreB, Mbl and MreBH built up mixed filaments underneath the cell membrane. Membrane protein RodZ localized to endosomes in S2 cells, but localized to the cell membrane when co-expressed with Mbl, showing that bacterial MreB/Mbl structures can recruit a protein to the cell membrane. Thus, MreB paralogs form a self-organizing and dynamic filamentous scaffold underneath the membrane that is able to recruit other proteins to the cell surface. PMID:22069484

  10. Molecular machines open cell membranes

    NASA Astrophysics Data System (ADS)

    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.

    2017-08-01

    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.

  11. Molecular machines open cell membranes.

    PubMed

    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

    2017-08-30

    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.

  12. Structural associations between organelle membranes in nectary parenchyma cells.

    PubMed

    Machado, Silvia Rodrigues; Gregório, Elisa A; Rodrigues, Tatiane M

    2018-05-01

    The close association between membranes and organelles, and the intense chloroplast remodeling in parenchyma cells of extrafloral nectaries occurred only at the secretion time and suggest a relationship with the nectar secretion. Associations between membranes and organelles have been well documented in different tissues and cells of plants, but poorly explored in secretory cells. Here, we described the close physical juxtaposition between membranes and organelles, mainly with chloroplasts, in parenchyma cells of Citharexylum myrianthum (Verbenaeceae) extrafloral nectaries under transmission electron microscopy, using conventional and microwave fixation. At the time of nectar secretion, nectary parenchyma cells exhibit a multitude of different organelle and membrane associations as mitochondria-mitochondria, mitochondria-endoplasmic reticulum, mitochondria-chloroplast, chloroplast-nuclear envelope, mitochondria-nuclear envelope, chloroplast-plasmalemma, chloroplast-chloroplast, chloroplast-tonoplast, chloroplast-peroxisome, and mitochondria-peroxisome. These associations were visualized as amorphous electron-dense material, a network of dense fibrillar material and/or dense bridges. Chloroplasts exhibited protrusions variable in shape and extension, which bring them closer to each other and to plasmalemma, tonoplast, and nuclear envelope. Parenchyma cells in the pre- and post-secretory stages did not exhibit any association or juxtaposition of membranes and organelles, and chloroplast protrusions were absent. Chloroplasts had peripheral reticulum that was more developed in the secretory stage. We propose that such subcellular phenomena during the time of nectar secretion optimize the movement of signaling molecules and the exchange of metabolites. Our results open new avenues on the potential mechanisms of organelle contact in parenchyma nectary cells, and reveal new attributes of the secretory cells on the subcellular level.

  13. Importance of methodological standardization for the ektacytometric measures of red blood cell deformability in sickle cell anemia.

    PubMed

    Renoux, Céline; Parrow, Nermi; Faes, Camille; Joly, Philippe; Hardeman, Max; Tisdale, John; Levine, Mark; Garnier, Nathalie; Bertrand, Yves; Kebaili, Kamila; Cuzzubbo, Daniela; Cannas, Giovanna; Martin, Cyril; Connes, Philippe

    2016-01-01

    Red blood cell (RBC) deformability is severely decreased in patients with sickle cell anemia (SCA), which plays a role in the pathophysiology of the disease. However, investigation of RBC deformability from SCA patients demands careful methodological considerations. We assessed RBC deformability by ektacytometry (LORRCA MaxSis, Mechatronics, The Netherlands) in 6 healthy individuals and 49 SCA patients and tested the effects of different heights of the RBC diffraction patterns, obtained by altering the camera gain of the LORRCA, on the result of RBC deformability measurements, expressed as Elongation Index (EI). Results indicate that the pattern of RBCs from control subjects adopts an elliptical shape under shear stress, whereas the pattern of RBCs from individuals with SCA adopts a diamond shape arising from the superposition of elliptical and circular patterns. The latter represent rigid RBCs. While the EI measures did not change with the variations of the RBC diffraction pattern heights in the control subjects, we observed a decrease of EI when the RBC diffraction pattern height is increased in the SCA group. The differences in SCA EI values measured at 5 Pa between the different diffraction pattern heights correlated with the percent of hemoglobin S and the percent of sickled RBC observed by microscopy. Our study confirms that the camera gain or aperture of the ektacytometer should be used to standardize the size of the RBC diffraction pattern height when measuring RBC deformability in sickle cell patients and underscores the potential clinical utility of this technique.

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

    PubMed

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

    2017-04-01

    An efficient system for direct delivery of integral membrane proteins is successfully developed using a new biocompatible exosome-based platform. Fusogenic exosomes harboring viral fusogen, vascular stomatitis virus (VSV)-G protein, can fuse with and modify plasma membranes in a process called "membrane editing." This can facilitate the transfer of biologically active membrane proteins into the target cell membranes both in vitro and in vivo. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. FORMATION OF INTRACYTOPLASMIC MEMBRANE SYSTEM OF MYCOBACTERIA RELATED TO CELL DIVISION

    PubMed Central

    Imaeda, Tamotsu; Ogura, Mituo

    1963-01-01

    Imaeda, Tamotsu (Instituto Venezolano de Investigaciones Científicas, Caracas, Venezuela) and Mitua Ogura. Formation of intracytoplasmic membrane system of mycobacteria related to cell division. J. Bacteriol. 85:150–163. 1963.—Mycobacterium leprae, M. lepraemurium, and a Mycobacterium sp. were observed with an electron microscope. In these bacilli, the three-dimensional structure of the intracytoplasmic membrane system consists of tubular infoldings of the invaginated plasma membrane. The moderately dense substance, presumably representing the cell-wall precursor, is found in the membranous system, especially in the rapid growth phase of mycobacteria. This system always shows an intimate relationship with cell division. A low-density zone, probably corresponding to the low-density substance which coats the cell wall, appears in the connecting regions of the system and in the longitudinal portion of the cell wall. These zones extend centripetally, and the separation of the cell wall occurs after the two zones meet. Based on these results, we hypothesize that the intracytoplasmic membrane system may produce cell-wall material during cell division of mycobacteria. Images PMID:13956365

  16. The fluidity of Chinese hamster ovary cell and bull sperm membranes after cholesterol addition.

    PubMed

    Purdy, P H; Fox, M H; Graham, J K

    2005-08-01

    Cell plasma membrane fluidity is affected by membrane lipid and protein composition as well as temperature. Altering the cholesterol content of a membrane can change membrane fluidity at different temperatures and this may affect cell survival during cryopreservation. In these experiments, we examined the effect that adding cholesterol to the membranes of Chinese hamster ovary cells (CHO) and bull sperm had on cell plasma membrane fluidity and cell survival when cells were cooled to 5 degrees C or were cryopreserved. Cells were treated with 0, 1.5 or 5.0mg cholesterol-loaded cyclodextrin (CLC), stained with N-((4-(6-phenyl-1,3,5-hexatrienyl)phenyl)propyl)trimethylammonium-p-toluenesulfonate (TMAP-DPH) to evaluate membrane fluidity and with propidium iodide to evaluate cell viability, prior to analysis by flow cytometry at 23, 5 degrees C, and after cryopreservation. CHO cells exhibited a single cell population with all cells having similar membrane fluidity. Membrane fluidity did not change when temperature had been reduced and then returned to 23 degrees C (P<0.05), however, adding cholesterol to the cells induced membranes to become more rigid (P<0.05). Bull sperm samples consisted of two cell subpopulations, one having relatively higher membrane fluidity than the other, regardless of cholesterol treatment or temperature. In addition, cells possessing the highest membrane fluidity did not survive cooling or cryopreservation efficiently. CLC treatment did not significantly alter membrane fluidity after temperature changes, but did maintain higher percentages of spermatozoa surviving cooling to 5 degrees C and cryopreservation (P<0.05). In conclusion, adding cholesterol to cell resulted in detectable membrane fluidity changes in CHO cells and increased survival of bull sperm after cooling to 5 degrees C and after cryopreservation.

  17. In vitro evaluation of the human gingival fibroblast/gingival mesenchymal stem cell dynamics through perforated guided tissue membranes: cell migration, proliferation and membrane stiffness assay.

    PubMed

    Gamal, A Y; Al-Berry, N N; Hassan, A A; Rashed, L A; Iacono, V J

    2017-06-01

    Migration of gingival fibroblasts/gingival mesenchymal stem cells through macro-perforated barrier membranes may allow them to participate positively in periodontal regeneration. The optimal guided tissue membrane perforation diameter that could favor maximum cell migration into the defect area and at the same time act as an occlusive barrier for gingival epithelium and its associated gingival extracellular matrix component is not yet identified. Cultured human gingival fibroblasts/gingival mesenchymal stem cells were placed in the upper chambers of 12-well collagen-coated polytetrafluoroethylene transwells, which were manually perforated with 0.2, 0.4 and 0.7 mm sized pores. The lower chambers of the transwells received blood clot as an attraction medium. The number of cells that have migrated to the lower chambers was calculated. Proliferation of these cells was evaluated using MTT assay. Scanning electron microscopy images were obtained for the lower surfaces of the transwell membranes. Perforated bovine collagen membranes (Tutopatch ® ) were subjected to mechanical testing to determine the tensile strength and modulus of elasticity. Group 3 (0.7 mm) showed significantly higher values for cell migration and proliferation. All groups showed a small degree of extracellular matrix migration through membrane perforations. Scanning electron microscopy evaluation revealed variable numbers of cells in fibrin matrices located mainly around the pore edges. There were non-significant differences between groups regarding mechanical properties. The present study demonstrated that macro-membrane perforations of 0.2, 0.4 and 0.7 mm are suitable pore diameters that could maintain membrane stiffness and allow for cellular migration. However, these membrane perforation diameters did not allow for total gingival connective tissue isolation. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  18. Red blood cells upregulate cytoprotective proteins and the labile iron pool in dividing human T cells despite a reduction in oxidative stress.

    PubMed

    Fonseca, Ana Mafalda; Pereira, Carlos F; Porto, Graça; Arosa, Fernando A

    2003-12-01

    We have recently reported that red blood cells (RBC) promote T cell growth and survival by inhibiting activation-induced T cell death. In the present study, we have examined parameters of oxidative stress and intracellular iron in activated T cells and correlated these data with the expression of ferritin, heme oxygenase-1 (HO-1), and the transferrin receptor CD71. T cells growing in the presence of RBC had reduced levels of reactive oxygen species (ROS) and oxidatively modified proteins, suggesting that RBC efficiently counteracted ROS production on the activated T cells. Flow cytometry and immunodetection demonstrated that T cells dividing in the presence of RBC had increased levels of intracellular ferritin rich in L-subunits and HO-1 along with a downmodulation in CD71 expression. Finally, using the fluorescent iron indicator calcein and flow cytometry analysis, we were able to show that a relative amount of the labile iron pool (LIP) was upregulated in T cells growing in the presence of RBC. These findings are consistent with a typical response to iron overload. However, neither heme compounds nor ferric iron reproduced the levels of expansion and survival of T cells induced by intact RBC. Altogether, these data suggest that RBC inhibit apoptosis of activated T cells by a combination of ROS scavenging and upregulation of cytoprotective proteins such as ferritin and HO-1, which may counteract a possible toxic effect of the increased intracellular free iron.

  19. Influence of red blood cell-derived microparticles upon vasoregulation

    PubMed Central

    Said, Ahmed S.; Doctor, Allan

    2017-01-01

    Here we review recent data and the evolving understanding of the role of red blood cell-derived microparticles (RMPs) in normal physiology and in disease progression. Microparticles (MPs) are small membrane vesicles derived from various parent cell types. MPs are produced in response to a variety of stimuli through several cytoskeletal and membrane phospholipid changes. MPs have been investigated as potential biomarkers for multiple disease processes and are thought to have biological effects, most notably in: promotion of coagulation, production and handling of reactive oxygen species, immune modulation, angiogenesis, and in apoptosis. Specifically, RMPs are produced normally during RBC maturation and their production is accelerated during processing and storage for transfusion. Several factors during RBC storage are known to trigger RMP production, including: increased intracellular calcium, increased potassium leakage, and energy failure with ATP depletion. Of note, RMP composition differs from that of intact RBCs, and the nature and composition of RMP components are affected by both storage duration and the character of storage solutions. Recognised RMP bioactivities include: promotion of coagulation, immune modulation, and promotion of endothelial adhesion, as well as influence upon vasoregulation via nitric oxide (NO) scavenging. Of particular relevance, RMPs are more avid NO scavengers than intact RBCs and this feature has been proposed as a mechanism for the impaired oxygen delivery homeostasis that has been observed following transfusion. Preliminary human studies demonstrate that circulating RMP abundance increases with RBC transfusion and is associated with altered plasma vasoactivity and abnormal vasoregulation. In summary, RMPs are submicron particles released from stored RBCs, with demonstrated vasoactive properties that appear to disturb oxygen delivery homeostasis. The clinical impact of RMPs in transfusion recipients is an area of continued

  20. Influence of red blood cell-derived microparticles upon vasoregulation.

    PubMed

    Said, Ahmed S; Doctor, Allan

    2017-10-01

    Here we review recent data and the evolving understanding of the role of red blood cell-derived microparticles (RMPs) in normal physiology and in disease progression. Microparticles (MPs) are small membrane vesicles derived from various parent cell types. MPs are produced in response to a variety of stimuli through several cytoskeletal and membrane phospholipid changes. MPs have been investigated as potential biomarkers for multiple disease processes and are thought to have biological effects, most notably in: promotion of coagulation, production and handling of reactive oxygen species, immune modulation, angiogenesis, and in apoptosis. Specifically, RMPs are produced normally during RBC maturation and their production is accelerated during processing and storage for transfusion. Several factors during RBC storage are known to trigger RMP production, including: increased intracellular calcium, increased potassium leakage, and energy failure with ATP depletion. Of note, RMP composition differs from that of intact RBCs, and the nature and composition of RMP components are affected by both storage duration and the character of storage solutions. Recognised RMP bioactivities include: promotion of coagulation, immune modulation, and promotion of endothelial adhesion, as well as influence upon vasoregulation via nitric oxide (NO) scavenging. Of particular relevance, RMPs are more avid NO scavengers than intact RBCs and this feature has been proposed as a mechanism for the impaired oxygen delivery homeostasis that has been observed following transfusion. Preliminary human studies demonstrate that circulating RMP abundance increases with RBC transfusion and is associated with altered plasma vasoactivity and abnormal vasoregulation. In summary, RMPs are submicron particles released from stored RBCs, with demonstrated vasoactive properties that appear to disturb oxygen delivery homeostasis. The clinical impact of RMPs in transfusion recipients is an area of continued

  1. Reassessing ecdysteroidogenic cells from the cell membrane receptors' perspective.

    PubMed

    Alexandratos, Alexandros; Moulos, Panagiotis; Nellas, Ioannis; Mavridis, Konstantinos; Dedos, Skarlatos G

    2016-02-05

    Ecdysteroids secreted by the prothoracic gland (PG) cells of insects control the developmental timing of their immature life stages. These cells have been historically considered as carrying out a single function in insects, namely the biochemical conversion of cholesterol to ecdysteroids and their secretion. A growing body of evidence shows that PG cells receive multiple cues during insect development so we tested the hypothesis that they carry out more than just one function in insects. We characterised the molecular nature and developmental profiles of cell membrane receptors in PG cells of Bombyx mori during the final larval stage and determined what receptors decode nutritional, developmental and physiological signals. Through iterative approaches we identified a complex repertoire of cell membrane receptors that are expressed in intricate patterns and activate previously unidentified signal transduction cascades in PG cells. The expression patterns of some of these receptors explain precisely the mechanisms that are known to control ecdysteroidogenesis. However, the presence of receptors for the notch, hedgehog and wingless signalling pathways and the expression of innate immunity-related receptors such as phagocytosis receptors, receptors for microbial ligands and Toll-like receptors call for a re-evaluation of the role these cells play in insects.

  2. Analysis of RBC-microparticles in stored whole blood bags - a promising marker to detect blood doping in sports?

    PubMed

    Voss, Sven Christian; Jaganjac, Morana; Al-Thani, Amna Mohamed; Grivel, Jean-Charles; Raynaud, Christophe Michel; Al-Jaber, Hind; Al-Menhali, Afnan Saleh; Merenkov, Zeyed Ahmad; Alsayrafi, Mohammed; Latiff, Aishah; Georgakopoulos, Costas

    2017-11-01

    Blood doping in sports is prohibited by the World Anti-Doping Agency (WADA). To find a possible biomarker for the detection of blood doping, we investigated the changes in blood stored in CPDA-1 blood bags of eight healthy subjects who donated one unit of blood. Aliquots were taken on days 0, 14, and 35. Platelet-free plasma was prepared and stored at -80°C until analysis on a flow cytometer dedicated for the analysis of microparticles (MPs). Changes in the number of red blood cell (RBC) -MPs were highly significant (p < 0.0001) with a mean of 219 (10^3/μL) on day 0 changing to 23 120 (10^3/μL) on day 14 and 29 310 (10^3/μL) on day 35. We conclude that RBC-MPs seem to be a promising biomarker for doping control but confirmation by a transfusion study is necessary. Copyright © 2017 John Wiley & Sons, Ltd.

  3. Introducing catalyst in alkaline membrane for improved performance direct borohydride fuel cells

    NASA Astrophysics Data System (ADS)

    Qin, Haiying; Lin, Longxia; Chu, Wen; Jiang, Wei; He, Yan; Shi, Qiao; Deng, Yonghong; Ji, Zhenguo; Liu, Jiabin; Tao, Shanwen

    2018-01-01

    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.

  4. The First Cell Membranes

    NASA Technical Reports Server (NTRS)

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

    2004-01-01

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

  5. Electrospun fiber membranes enable proliferation of genetically modified cells

    PubMed Central

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

    2013-01-01

    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

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

    PubMed

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

    2014-04-01

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

  7. The actin homologue MreB organizes the bacterial cell membrane

    PubMed Central

    Strahl, Henrik; Bürmann, Frank; Hamoen, Leendert W.

    2014-01-01

    The eukaryotic cortical actin cytoskeleton creates specific lipid domains, including lipid rafts, which determine the distribution of many membrane proteins. Here we show that the bacterial actin homologue MreB displays a comparable activity. MreB forms membrane-associated filaments that coordinate bacterial cell wall synthesis. We noticed that the MreB cytoskeleton influences fluorescent staining of the cytoplasmic membrane. Detailed analyses combining an array of mutants, using specific lipid staining techniques and spectroscopic methods, revealed that MreB filaments create specific membrane regions with increased fluidity (RIFs). Interference with these fluid lipid domains (RIFs) perturbs overall lipid homeostasis and affects membrane protein localization. The influence of MreB on membrane organization and fluidity may explain why the active movement of MreB stimulates membrane protein diffusion. These novel MreB activities add additional complexity to bacterial cell membrane organization and have implications for many membrane-associated processes. PMID:24603761

  8. The actin homologue MreB organizes the bacterial cell membrane.

    PubMed

    Strahl, Henrik; Bürmann, Frank; Hamoen, Leendert W

    2014-03-07

    The eukaryotic cortical actin cytoskeleton creates specific lipid domains, including lipid rafts, which determine the distribution of many membrane proteins. Here we show that the bacterial actin homologue MreB displays a comparable activity. MreB forms membrane-associated filaments that coordinate bacterial cell wall synthesis. We noticed that the MreB cytoskeleton influences fluorescent staining of the cytoplasmic membrane. Detailed analyses combining an array of mutants, using specific lipid staining techniques and spectroscopic methods, revealed that MreB filaments create specific membrane regions with increased fluidity (RIFs). Interference with these fluid lipid domains (RIFs) perturbs overall lipid homeostasis and affects membrane protein localization. The influence of MreB on membrane organization and fluidity may explain why the active movement of MreB stimulates membrane protein diffusion. These novel MreB activities add additional complexity to bacterial cell membrane organization and have implications for many membrane-associated processes.

  9. Erythrocyte shape abnormalities, membrane oxidative damage, and β-actin alterations: an unrecognized triad in classical autism.

    PubMed

    Ciccoli, Lucia; De Felice, Claudio; Paccagnini, Eugenio; Leoncini, Silvia; Pecorelli, Alessandra; Signorini, Cinzia; Belmonte, Giuseppe; Guerranti, Roberto; Cortelazzo, Alessio; Gentile, Mariangela; Zollo, Gloria; Durand, Thierry; Valacchi, Giuseppe; Rossi, Marcello; Hayek, Joussef

    2013-01-01

    Autism spectrum disorders (ASDs) are a complex group of neurodevelopment disorders steadily rising in frequency and treatment refractory, where the search for biological markers is of paramount importance. Although red blood cells (RBCs) membrane lipidomics and rheological variables have been reported to be altered, with some suggestions indicating an increased lipid peroxidation in the erythrocyte membrane, to date no information exists on how the oxidative membrane damage may affect cytoskeletal membrane proteins and, ultimately, RBCs shape in autism. Here, we investigated RBC morphology by scanning electron microscopy in patients with classical autism, that is, the predominant ASDs phenotype (age range: 6-26 years), nonautistic neurodevelopmental disorders (i.e., "positive controls"), and healthy controls (i.e., "negative controls"). A high percentage of altered RBCs shapes, predominantly elliptocytes, was observed in autistic patients, but not in both control groups. The RBCs altered morphology in autistic subjects was related to increased erythrocyte membrane F2-isoprostanes and 4-hydroxynonenal protein adducts. In addition, an oxidative damage of the erythrocyte membrane β-actin protein was evidenced. Therefore, the combination of erythrocyte shape abnormalities, erythrocyte membrane oxidative damage, and β-actin alterations constitutes a previously unrecognized triad in classical autism and provides new biological markers in the diagnostic workup of ASDs.

  10. Erythrocyte Shape Abnormalities, Membrane Oxidative Damage, and β-Actin Alterations: An Unrecognized Triad in Classical Autism

    PubMed Central

    Ciccoli, Lucia; De Felice, Claudio; Pecorelli, Alessandra; Belmonte, Giuseppe; Guerranti, Roberto; Cortelazzo, Alessio; Durand, Thierry; Valacchi, Giuseppe; Rossi, Marcello; Hayek, Joussef

    2013-01-01

    Autism spectrum disorders (ASDs) are a complex group of neurodevelopment disorders steadily rising in frequency and treatment refractory, where the search for biological markers is of paramount importance. Although red blood cells (RBCs) membrane lipidomics and rheological variables have been reported to be altered, with some suggestions indicating an increased lipid peroxidation in the erythrocyte membrane, to date no information exists on how the oxidative membrane damage may affect cytoskeletal membrane proteins and, ultimately, RBCs shape in autism. Here, we investigated RBC morphology by scanning electron microscopy in patients with classical autism, that is, the predominant ASDs phenotype (age range: 6–26 years), nonautistic neurodevelopmental disorders (i.e., “positive controls”), and healthy controls (i.e., “negative controls”). A high percentage of altered RBCs shapes, predominantly elliptocytes, was observed in autistic patients, but not in both control groups. The RBCs altered morphology in autistic subjects was related to increased erythrocyte membrane F2-isoprostanes and 4-hydroxynonenal protein adducts. In addition, an oxidative damage of the erythrocyte membrane β-actin protein was evidenced. Therefore, the combination of erythrocyte shape abnormalities, erythrocyte membrane oxidative damage, and β-actin alterations constitutes a previously unrecognized triad in classical autism and provides new biological markers in the diagnostic workup of ASDs. PMID:24453417

  11. Imidazolium-Based Polymeric Materials as Alkaline Anion-Exchange Fuel Cell Membranes

    NASA Technical Reports Server (NTRS)

    Narayan, Sri R.; Yen, Shiao-Ping S.; Reddy, Prakash V.; Nair, Nanditha

    2012-01-01

    Polymer electrolyte membranes that conduct hydroxide ions have potential use in fuel cells. A variety of polystyrene-based quaternary ammonium hydroxides have been reported as anion exchange fuel cell membranes. However, the hydrolytic stability and conductivity of the commercially available membranes are not adequate to meet the requirements of fuel cell applications. When compared with commercially available membranes, polystyrene-imidazolium alkaline membrane electrolytes are more stable and more highly conducting. At the time of this reporting, this has been the first such usage for imidazolium-based polymeric materials for fuel cells. Imidazolium salts are known to be electrochemically stable over wide potential ranges. By controlling the relative ratio of imidazolium groups in polystyrene-imidazolium salts, their physiochemical properties could be modulated. Alkaline anion exchange membranes based on polystyrene-imidazolium hydroxide materials have been developed. The first step was to synthesize the poly(styrene-co-(1-((4-vinyl)methyl)-3- methylimidazolium) chloride through a free-radical polymerization. Casting of this material followed by in situ treatment of the membranes with sodium hydroxide solutions provided the corresponding hydroxide salts. Various ratios of the monomers 4-chloromoethylvinylbenzine (CMVB) and vinylbenzine (VB) provided various compositions of the polymer. The preferred material, due to the relative ease of casting the film, and its relatively low hygroscopic nature, was a 2:1 ratio of CMVB to VB. Testing confirmed that at room temperature, the new membranes outperformed commercially available membranes by a large margin. With fuel cells now in use at NASA and in transportation, and with defense potential, any improvement to fuel cell efficiency is a significant development.

  12. Chemical Imaging of the Cell Membrane by NanoSIMS

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Weber, P K; Kraft, M L; Frisz, J F

    2010-02-23

    The existence of lipid microdomains and their role in cell membrane organization are currently topics of great interest and controversy. The cell membrane is composed of a lipid bilayer with embedded proteins that can flow along the two-dimensional surface defined by the membrane. Microdomains, known as lipid rafts, are believed to play a central role in organizing this fluid system, enabling the cell membrane to carry out essential cellular processes, including protein recruitment and signal transduction. Lipid rafts are also implicated in cell invasion by pathogens, as in the case of the HIV. Therefore, understanding the role of lipid raftsmore » 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

  13. Listeria membrane protrusion collapse: Requirement of Cyclophilin A for Listeria cell-to-cell spreading.

    PubMed

    Dhanda, Aaron S; Lulic, Katarina T; Vogl, A Wayne; Mc Gee, Margaret M; Chiu, Robert H; Guttman, Julian A

    2018-05-04

    Listeria generate actin-rich tubular protrusions at the plasma membrane that propel the bacteria into neighbouring cells. The precise molecular mechanisms governing the formation of these protrusions remain poorly defined. Here we demonstrate that the PPIase Cyclophilin A (CypA) is hijacked by Listeria at membrane protrusions used for cell-to-cell spreading. CypA localizes within the F-actin of these structures and is crucial for their proper formation, as in cells depleted of CypA, these extended actin-rich structures are mis-shaped and collapsed due to changes within the F-actin network. The lack of structural integrity within the Listeria membrane protrusions hampers the microbes from spreading from CypA null cells. Our results demonstrate a crucial role for CypA during Listeria infections.

  14. Enhancement of the blood compatibility of dialyzer membranes by the physical adsorption of human thrombomodulin (ART-123).

    PubMed

    Omichi, Masaaki; Matsusaki, Michiya; Kato, Shinya; Maruyama, Ikuro; Akashi, Mitsuru

    2010-11-01

    ART-123 is a recombinant soluble human thrombomodulin (hTM) with excellent anticoagulant activity. We focused on improving the blood compatibility of the polysulfone-polyvinylpyrrolidone dialyzer surface by the physical adsorption of ART-123 onto the surface. The blood compatibility of the dialyzer with the hTM adsorbed membrane was evaluated by measuring the differential pressure between the arterial and the venous pressures and by blood parameters during blood circulation. The hTM adsorbed dialyzer membrane inhibited blood clot formation without heparin administration due to the anticoagulant activity of hTM for over 4 h. The physically adsorbed hTM was stable during blood circulation, and it did not affect activated clotting time, which is significant drawback of heparin administration, and blood cell counts of RBC, WBC, or platelets. The physical adsorption of hTM onto the dialyzer membrane will be a simple and safe method to prevent blood coagulation during dialysis instead of heparin administration. © 2010 Wiley Periodicals, Inc.

  15. Mature red blood cells: from optical model to inverse light-scattering problem.

    PubMed

    Gilev, Konstantin V; Yurkin, Maxim A; Chernyshova, Ekaterina S; Strokotov, Dmitry I; Chernyshev, Andrei V; Maltsev, Valeri P

    2016-04-01

    We propose a method for characterization of mature red blood cells (RBCs) morphology, based on measurement of light-scattering patterns (LSPs) of individual RBCs with the scanning flow cytometer and on solution of the inverse light-scattering (ILS) problem for each LSP. We considered a RBC shape model, corresponding to the minimal bending energy of the membrane with isotropic elasticity, and constructed an analytical approximation, which allows rapid simulation of the shape, given the diameter and minimal and maximal thicknesses. The ILS problem was solved by the nearest-neighbor interpolation using a preliminary calculated database of 250,000 theoretical LSPs. For each RBC in blood sample we determined three abovementioned shape characteristics and refractive index, which also allows us to calculate volume, surface area, sphericity index, spontaneous curvature, hemoglobin concentration and content.

  16. Mature red blood cells: from optical model to inverse light-scattering problem

    PubMed Central

    Gilev, Konstantin V.; Yurkin, Maxim A.; Chernyshova, Ekaterina S.; Strokotov, Dmitry I.; Chernyshev, Andrei V.; Maltsev, Valeri P.

    2016-01-01

    We propose a method for characterization of mature red blood cells (RBCs) morphology, based on measurement of light-scattering patterns (LSPs) of individual RBCs with the scanning flow cytometer and on solution of the inverse light-scattering (ILS) problem for each LSP. We considered a RBC shape model, corresponding to the minimal bending energy of the membrane with isotropic elasticity, and constructed an analytical approximation, which allows rapid simulation of the shape, given the diameter and minimal and maximal thicknesses. The ILS problem was solved by the nearest-neighbor interpolation using a preliminary calculated database of 250,000 theoretical LSPs. For each RBC in blood sample we determined three abovementioned shape characteristics and refractive index, which also allows us to calculate volume, surface area, sphericity index, spontaneous curvature, hemoglobin concentration and content. PMID:27446656

  17. Measurement of blood coagulation with considering RBC aggregation through a microchip-based light transmission aggregometer.

    PubMed

    Lim, Hyunjung; Nam, Jeonghun; Xue, Shubin; Shin, Sehyun

    2011-01-01

    Even though blood coagulation can be tested by various methods and techniques, the effect of RBC aggregation on blood coagulation is not fully understood. The present study monitored clot formation in a microchip-based light transmission aggregometer. Citrated blood samples with and without the addition of calcium ion solution were initially disaggregated by rotating a stirrer in the microchip. After abrupt stop of the rotating stirrer, the transmitted light intensity over time was recorded. The syllectogram (light intensity vs. time graph) manifested a rapid increase that is associated with RBC aggregation followed by a decrease that is associated with blood coagulation. The time to reach the peak point was used as a new index of coagulation time (CT) and ranged from 200 to 500 seconds in the present measurements. The CT was inversely proportional to the concentration of fibrinogen, which enhances RBC aggregation. In addition, the CT was inversely proportional to the hematocrit, which is similar to the case of the prothrombin time (PT), as measured by a commercial coagulometer. Thus, we carefully concluded that RBC aggregation should be considered in tests of blood coagulation.

  18. Manipulating membrane lipid profiles to restore T-cell function in autoimmunity.

    PubMed

    Waddington, Kirsty E; Jury, Elizabeth C

    2015-08-01

    Plasma membrane lipid rafts are heterogeneous cholesterol and glycosphingolipid (GSL)-enriched microdomains, within which the tight packing of cholesterol with the saturated-acyl chains of GSLs creates a region of liquid-order relative to the surrounding disordered membrane. Thus lipid rafts govern the lateral mobility and interaction of membrane proteins and regulate a plethora of signal transduction events, including T-cell antigen receptor (TCR) signalling. The pathways regulating homoeostasis of membrane cholesterol and GSLs are tightly controlled and alteration of these metabolic processes coincides with immune cell dysfunction as is evident in atherosclerosis, cancer and autoimmunity. Indeed, membrane lipid composition is emerging as an important factor influencing the ability of cells to respond appropriately to microenvironmental stimuli. Consequently, there is increasing interest in targeting membrane lipids or their metabolic control as a novel therapeutic approach to modulate immune cell behaviour and our recent work demonstrates that this is a promising strategy in T-cells from patients with the autoimmune disease systemic lupus erythematosus (SLE). © 2015 Authors; published by Portland Press Limited.

  19. Hybrid continuum-coarse-grained modeling of erythrocytes

    NASA Astrophysics Data System (ADS)

    Lyu, Jinming; Chen, Paul G.; Boedec, Gwenn; Leonetti, Marc; Jaeger, Marc

    2018-06-01

    The red blood cell (RBC) membrane is a composite structure, consisting of a phospholipid bilayer and an underlying membrane-associated cytoskeleton. Both continuum and particle-based coarse-grained RBC models make use of a set of vertices connected by edges to represent the RBC membrane, which can be seen as a triangular surface mesh for the former and a spring network for the latter. Here, we present a modeling approach combining an existing continuum vesicle model with a coarse-grained model for the cytoskeleton. Compared to other two-component approaches, our method relies on only one mesh, representing the cytoskeleton, whose velocity in the tangential direction of the membrane may be different from that of the lipid bilayer. The finitely extensible nonlinear elastic (FENE) spring force law in combination with a repulsive force defined as a power function (POW), called FENE-POW, is used to describe the elastic properties of the RBC membrane. The mechanical interaction between the lipid bilayer and the cytoskeleton is explicitly computed and incorporated into the vesicle model. Our model includes the fundamental mechanical properties of the RBC membrane, namely fluidity and bending rigidity of the lipid bilayer, and shear elasticity of the cytoskeleton while maintaining surface-area and volume conservation constraint. We present three simulation examples to demonstrate the effectiveness of this hybrid continuum-coarse-grained model for the study of RBCs in fluid flows.

  20. Cooperative binding of Annexin A5 to phosphatidylserine on apoptotic cell membranes

    NASA Astrophysics Data System (ADS)

    Janko, Christina; Jeremic, Ivica; Biermann, Mona; Chaurio, Ricardo; Schorn, Christine; Muñoz, Luis E.; Herrmann, Martin

    2013-12-01

    Healthy cells exhibit an asymmetric plasma membrane with phosphatidylserine (PS) located on the cytoplasmic leaflet of the plasma membrane bilayer. Annexin A5-FITC, a PS binding protein, is commonly used to evaluate apoptosis in flow cytometry. PS exposed by apoptotic cells serves as a major ‘eat-me’ signal for phagocytes. Although exposition of PS has been observed after alternative stimuli, no clearance of viable, PS exposing cells has been detected. Thus, besides PS exposure, membranes of viable and apoptotic cells might exhibit specific characteristics. Here, we show that Annexin A5 binds in a cooperative manner to different types of dead cells. Shrunken apoptotic cells thereby showed the highest Hill coefficient values. Contrarily, parafomaldehyde fixation of apoptotic cells completely abrogates the cooperativity effect seen with dead and dying cells. We tend to speculate that the cooperative binding of Annexin A5 to the membranes of apoptotic cells reflects higher fluidity of the exposed membranes facilitating PS clustering.

  1. Light-scattering analysis of ultrasonic wave's influence on the RBC agglutination in vitro

    NASA Astrophysics Data System (ADS)

    Doubrovski, Valeri A.; Dvoretski, Costanten N.

    1999-04-01

    Elastic light scattering is one of the most often used optical methods to analyze the cells agglutination reaction - the base of a great number of medical diagnostic test and biomedical investigations. The increase of the resolution of methods and apparatus towards the induced cells aggregation - the foundation of the reaction of agglutination, is quite an actual problem. The solution of this problem increases the reliability of the diagnostic test and gives an opportunity to achieve the diagnostic information in the cases when the traditional approaches do not lead to the diagnostic results. The attempt to increase the resolution of the immune reaction analyzer by means of ultrasonic waves action on the reagent mixture in vitro is taken in this paper. The RBC agglutination reaction which is usually used for the blood group type examination is chosen as an example of an object of the investigation. Different laser optical trains of the devices based on the turbidimetric and nephelometric methods and their combination are analyzed here. The influence of the ultrasonic wave time interval action and of the features of the sample preparation procedure on the resolution towards the agglutination process was investigated in this work. It is shown that the ultrasonic wave action on the reagent mixture leads to a large gain in the resolution of the device towards the RBC agglutination process. The experiments showed that the resolution of the device was enough to register the agglutination process even for the erythrocytes with weak agglutination ability when the reaction was invisible without ultrasonic action. It occurred that the diagnostic test time was more than by an order shortened due to the ultrasonic wave action. The optimal ultrasonic time interval action, the sample preparation technology and experimental technique were defined. The principle of the ultrasonic wave action on the cells agglutination process suggested here can be spread out on the immune

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

    NASA Technical Reports Server (NTRS)

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

    1996-01-01

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

  3. An AFM-based pit-measuring method for indirect measurements of cell-surface membrane vesicles

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhang, Xiaojun; Department of Biotechnology, Nanchang University, Nanchang, Jiangxi 330031; Chen, Yuan

    2014-03-28

    Highlights: • Air drying induced the transformation of cell-surface membrane vesicles into pits. • An AFM-based pit-measuring method was developed to measure cell-surface vesicles. • Our method detected at least two populations of cell-surface membrane vesicles. - Abstract: Circulating membrane vesicles, which are shed from many cell types, have multiple functions and have been correlated with many diseases. Although circulating membrane vesicles have been extensively characterized, the status of cell-surface membrane vesicles prior to their release is less understood due to the lack of effective measurement methods. Recently, as a powerful, micro- or nano-scale imaging tool, atomic force microscopy (AFM)more » has been applied in measuring circulating membrane vesicles. However, it seems very difficult for AFM to directly image/identify and measure cell-bound membrane vesicles due to the similarity of surface morphology between membrane vesicles and cell surfaces. Therefore, until now no AFM studies on cell-surface membrane vesicles have been reported. In this study, we found that air drying can induce the transformation of most cell-surface membrane vesicles into pits that are more readily detectable by AFM. Based on this, we developed an AFM-based pit-measuring method and, for the first time, used AFM to indirectly measure cell-surface membrane vesicles on cultured endothelial cells. Using this approach, we observed and quantitatively measured at least two populations of cell-surface membrane vesicles, a nanoscale population (<500 nm in diameter peaking at ∼250 nm) and a microscale population (from 500 nm to ∼2 μm peaking at ∼0.8 μm), whereas confocal microscopy only detected the microscale population. The AFM-based pit-measuring method is potentially useful for studying cell-surface membrane vesicles and for investigating the mechanisms of membrane vesicle formation/release.« less

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

    PubMed

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

    2015-09-01

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

  5. Low-autofluorescence fluoropolymer membrane filters for cell filtration

    NASA Astrophysics Data System (ADS)

    Kihara, Naoto; Kuboyama, Daiki; Onoshima, Daisuke; Ishikawa, Kenji; Tanaka, Hiromasa; Ozawa, Naoya; Hase, Tetsunari; Koguchi, Ryohei; Yukawa, Hiroshi; Odaka, Hidefumi; Hasegawa, Yoshinori; Baba, Yoshinobu; Hori, Masaru

    2018-06-01

    A fluoropolymer membrane filter with through-holes was fabricated by photolithographic patterning and the dry etching method. 380,000 highly packed through-holes, each with a diameter of 7 µm were able to cover a whole area with a diameter of 13 mm. Ethylene tetrafluoroethylene (ETFE) was used as the membrane, which was suitable for the fluorescence detection of rare cells such as circulating tumor cells (CTCs) in human blood. The device fabrication for the size based capture of rare cells in blood such as CTCs is realized in this study.

  6. Effect of plasma membrane fluidity on serotonin transport by endothelial cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Block, E.R.; Edwards, D.

    1987-11-01

    To evaluate the effect of plasma membrane fluidity of lung endothelial cells on serotonin transport, porcine pulmonary artery endothelial cells were incubated for 3 h with either 0.1 mM cholesterol hemisuccinate, 0.1 mM cis-vaccenic acid, or vehicle (control), after which plasma membrane fluidity and serotinin transport were measured. Fluorescence spectroscopy was used to measure fluidity in the plasma membrane. Serotonin uptake was calculated from the disappearance of ({sup 14}C)-serotonin from the culture medium. Cholesterol decreased fluidity in the subpolar head group and central and midacyl side-chain regions of the plasma membrane and decreased serotonin transport, whereas cis-vaccenic acid increased fluiditymore » in the central and midacyl side-chain regions of the plasma membrane and also increased serotonin transport. Cis-vaccenic acid had no effect of fluidity in the subpolar head group region of the plasma membrane. These results provide evidence that the physical state of the central and midacyl chains within the pulmonary artery endothelial cell plasma membrane lipid bilayer modulates transmembrane transport of serotonin by these cells.« less

  7. Nanoscale manipulation of membrane curvature for probing endocytosis in live cells.

    PubMed

    Zhao, Wenting; Hanson, Lindsey; Lou, Hsin-Ya; Akamatsu, Matthew; Chowdary, Praveen D; Santoro, Francesca; Marks, Jessica R; Grassart, Alexandre; Drubin, David G; Cui, Yi; Cui, Bianxiao

    2017-08-01

    Clathrin-mediated endocytosis (CME) involves nanoscale bending and inward budding of the plasma membrane, by which cells regulate both the distribution of membrane proteins and the entry of extracellular species. Extensive studies have shown that CME proteins actively modulate the plasma membrane curvature. However, the reciprocal regulation of how the plasma membrane curvature affects the activities of endocytic proteins is much less explored, despite studies suggesting that membrane curvature itself can trigger biochemical reactions. This gap in our understanding is largely due to technical challenges in precisely controlling the membrane curvature in live cells. In this work, we use patterned nanostructures to generate well-defined membrane curvatures ranging from +50 nm to -500 nm radius of curvature. We find that the positively curved membranes are CME hotspots, and that key CME proteins, clathrin and dynamin, show a strong preference towards positive membrane curvatures with a radius <200 nm. Of ten CME-related proteins we examined, all show preferences for positively curved membrane. In contrast, other membrane-associated proteins and non-CME endocytic protein caveolin1 show no such curvature preference. Therefore, nanostructured substrates constitute a novel tool for investigating curvature-dependent processes in live cells.

  8. Microdomains in the membrane landscape shape antigen-presenting cell function.

    PubMed

    Zuidscherwoude, Malou; de Winde, Charlotte M; Cambi, Alessandra; van Spriel, Annemiek B

    2014-02-01

    The plasma membrane of immune cells is a highly organized cell structure that is key to the initiation and regulation of innate and adaptive immune responses. It is well-established that immunoreceptors embedded in the plasma membrane have a nonrandom spatial distribution that is important for coupling to components of intracellular signaling cascades. In the last two decades, specialized membrane microdomains, including lipid rafts and TEMs, have been identified. These domains are preformed structures ("physical entities") that compartmentalize proteins, lipids, and signaling molecules into multimolecular assemblies. In APCs, different microdomains containing immunoreceptors (MHC proteins, PRRs, integrins, among others) have been reported that are imperative for efficient pathogen recognition, the formation of the immunological synapse, and subsequent T cell activation. In addition, recent work has demonstrated that tetraspanin microdomains and lipid rafts are involved in BCR signaling and B cell activation. Research into the molecular mechanisms underlying membrane domain formation is fundamental to a comprehensive understanding of membrane-proximal signaling and APC function. This review will also discuss the advances in the microscopy field for the visualization of the plasma membrane, as well as the recent progress in targeting microdomains as novel, therapeutic approach for infectious and malignant diseases.

  9. Low-Cost Proton Conducting Membranes for PEM Fuel Cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hu, Hongxing

    Proton exchange membrane (PEM) is the key component in PEM fuel cells that critically determines the system performance and its economic viability. Presently, the state-of-the-art PEMs, such as Nafion membranes, are based on perfluorosulfonic acid (PFSA) ionomers. But these ionomer materials are expensive, particularly at the low volumes that will be needed for initial commercialization. Besides, they are not suitable for fuel cells operated beyond 100°C, because of the limitations connected to the humidification requirement of such membrane materials, limiting the maximum operating temperature to about 90°C. Fuel cells for transportation applications are required to operate in a wide temperaturemore » range from –20°C to 120°C. Low-cost PEMs with capabilities in a range of temperature and humidity conditions are urgently needed to meet the DOE fuel cell targets for transportation applications. Amsen Technologies LLC chooses to address the DOE call with a novel reinforced PEM approach based on new, non-PFSA proton conducting ionomers developed from our previous DOE SBIR projects. Along with this approach is the use of very cheap, ultra thin and highly porous microporous polymer meshes as the support for the membrane. The new PEM is expected to have significant cost advantages over traditional PEMs. The microporous polyolefin support costs $2-3/m 2; and the new ionomers that Amsen has developed are estimated at ~$250/kg at the higher end including material costs and labor costs (which may go down in the future as the processing is optimized and production scaled up). These have led to an estimate of total material cost for the membrane at $11 to $12/m 2, offering high potential of meeting the DOE cost targets (≤$20/m 2) after adding processing cost and profit margin. The Phase I results have successfully demonstrated that it is very promising to develop the intended low-cost, high-performance PEM membrane. Suitable material system has been identified, and

  10. Hawthorn special extract WS® 1442 increases red blood cell NO-formation without altering red blood cell deformability.

    PubMed

    Rieckeheer, Eva; Schwinger, Robert H G; Bloch, Wilhelm; Brixius, Klara

    2011-12-15

    WS(®) 1442 is a special extract of hawthorn leaves with flowers used for the treatment of mild cardiac failure. The activation of endothelial nitric oxide synthase (eNOS) has been shown to contribute to its vasodilating properties. Quite recently it has been demonstrated that red blood cells (RBCs) express a functional NO-synthase (rbcNOS) and rbcNOS activation has been associated with increased RBC deformability. The aim of the present study was to determine whether WS(®) 1442 is able to activate rbcNOS, to induce NO-formation in RBC and to alter RBC-deformability. Blood from healthy volunteers was incubated with WS(®) 1442 (25-100 μg/ml) for up to 30 min. RbcNOS activation was detected by immunohistochemical staining of phosphorylated rbcNOS and NO-formation was examined by diaminofluorescein (DAF) fluorescence. RBC deformability was measured by a laser assisted optical rotational cell analyzer. Serine 1177 of RbcNOS (rbcNOS Ser(1177)) was time- and concentration-dependently phosphorylated by WS(®) 1442. Rates of rbcNOS Ser(1177) phosphorylation were up to 149% higher in RBCs treated with WS(®) 1442 in comparison to control (DMSO 0.05%). WS(®) 1442 induced a time-dependent increase in NO-formation in RBCs which reached its maximum after 5 min. An increase in shear stress (0.3-50 Pa) caused an increase in RBC deformability. WS(®) 1442 did not change either basal or maximal RBC-deformability or shear stress sensitivity of RBC at normoxia. WS(®) 1442 activates rbcNOS and causes NO-formation in RBCs. WS(®) 1442-dependent NO-formation however does not affect RBC-deformability at normoxia. Copyright © 2011. Published by Elsevier GmbH.

  11. Separation of red blood cells in deep deterministic lateral displacement devices

    NASA Astrophysics Data System (ADS)

    Kabacaoglu, Gokberk; Biros, George

    2017-11-01

    Microfluidic cell separation techniques are of great interest since they help rapid medical diagnoses and tests. Deterministic lateral displacement (DLD) is one of them. A DLD device consists of arrays of pillars. Main flow and alignment of the pillars define two different directions. Size-based separation of rigid spherical particles is possible as they follow one of these directions depending on their sizes. However, the separation of non-spherical deformable particles such as red blood cells (RBCs) is more complicated than that due to their intricate dynamics. We study the separation of RBCs in DLD using an in-house integral equation solver. We systematically investigate the effects of the interior fluid viscosity and the membrane elasticity of an RBC on its behavior. These mechanical properties of a cell determine its deformability, which can be altered by several diseases. We particularly consider deep devices in which an RBC can show rich dynamics such as tank-treading and tumbling. It turns out that strong hydrodynamic lift force moves the tank-treading cells along the pillars and downward force leads the tumbling ones to move with the flow. Thereby, deformability-based separation of RBCs is possible.

  12. Human mesenchymal stem cells promote CD34+ hematopoietic stem cell proliferation with preserved red blood cell differentiation capacity.

    PubMed

    Lau, Show Xuan; Leong, Yin Yee; Ng, Wai Hoe; Ng, Albert Wee Po; Ismail, Ida Shazrina; Yusoff, Narazah Mohd; Ramasamy, Rajesh; Tan, Jun Jie

    2017-06-01

    Studies showed that co-transplantation of mesenchymal stem cells (MSCs) and cord blood-derived CD34 + hematopoietic stem cells (HSCs) offered greater therapeutic effects but little is known regarding the effects of human Wharton's jelly derived MSCs on HSC expansion and red blood cell (RBC) generation in vitro. This study aimed to investigate the effects of MSCs on HSC expansion and differentiation. HSCs were co-cultured with MSCs or with 10% MSCs-derived conditioned medium, with HSCs cultured under standard medium served as a control. Cell expansion rates, number of mononuclear cell post-expansion and number of enucleated cells post-differentiation were evaluated. HSCs showed superior proliferation in the presence of MSC with mean expansion rate of 3.5 × 10 8  ± 1.8 × 10 7 after day 7 compared to the conditioned medium and the control group (8.9 × 10 7  ± 1.1 × 10 8 and 7.0 × 10 7  ± 3.3 × 10 6 respectively, P < 0.001). Although no significant differences in RBC differentiation were observed between groups at passage IV, the number of enucleated cell was greater compared to earlier passages, indicating successful RBC differentiation. Cord blood-derived CD34 + HSCs can be greatly expanded by co-culturing with MSCs without affecting the RBC differentiation capability, suggesting the importance of direct MSC-HSCs contact in HSC expansion and RBC differentiation. © 2017 International Federation for Cell Biology.

  13. Plasma membrane organization and dynamics is probe and cell line dependent.

    PubMed

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

    2017-09-01

    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. Fuel cell subassemblies incorporating subgasketed thrifted membranes

    DOEpatents

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

    2014-01-28

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

  15. Fuel cell subassemblies incorporating subgasketed thrifted membranes

    DOEpatents

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

    2016-03-01

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

  16. A Lattice Boltzmann Fictitious Domain Method for Modeling Red Blood Cell Deformation and Multiple-Cell Hydrodynamic Interactions in Flow

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Shi, Xing; Lin, Guang; Zou, Jianfeng

    To model red blood cell (RBC) deformation in flow, the recently developed LBM-DLM/FD method ([Shi and Lim, 2007)29], derived from the lattice Boltzmann method and the distributed Lagrange multiplier/fictitious domain methodthe fictitious domain method, is extended to employ the mesoscopic network model for simulations of red blood cell deformation. The flow is simulated by the lattice Boltzmann method with an external force, while the network model is used for modeling red blood cell deformation and the fluid-RBC interaction is enforced by the Lagrange multiplier. To validate parameters of the RBC network model, sThe stretching numerical tests on both coarse andmore » fine meshes are performed and compared with the corresponding experimental data to validate the parameters of the RBC network model. In addition, RBC deformation in pipe flow and in shear flow is simulated, revealing the capacity of the current method for modeling RBC deformation in various flows.« less

  17. The role of cell membranes in the regulation of lignification in pine cells

    NASA Technical Reports Server (NTRS)

    Hendrix, D. L.

    1978-01-01

    The identity of pine cell membranes bearing PAL enzyme activity, the isolation of a plasma membrane preparation from pine cells for testing as a regulatory barrier in lignification, and the measurement of the geopotential effect in pine stems are presented. A model to describe and predict the interaction of gravity and lignification of higher plants was developed.

  18. Aluminum-induced cell death of barley-root border cells is correlated with peroxidase- and oxalate oxidase-mediated hydrogen peroxide production.

    PubMed

    Tamás, L; Budíková, S; Huttová, J; Mistrík, I; Simonovicová, M; Siroká, B

    2005-06-01

    The function of root border cells (RBC) during aluminum (Al) stress and the involvement of oxalate oxidase, peroxidase and H(2)O(2) generation in Al toxicity were studied in barley roots. Our results suggest that RBC effectively protect the barley root tip from Al relative to the situation in roots cultivated in hydroponics where RBC are not sustained in the area surrounding the root tip. The removal of RBC from Al-treated roots increased root growth inhibition, Al and Evans blue uptake, inhibition of RBC production, the level of dead RBC, peroxidase and oxalate oxidase activity and the production of H(2)O(2). Our results suggest that even though RBC actively produce active oxygen species during Al stress, their role in the protection of root tips against Al toxicity is to chelate Al in their dead cell body.

  19. 40 CFR 35.2035 - Rotating biological contractor (RBC) replacement grants.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... AND OTHER FEDERAL ASSISTANCE STATE AND LOCAL ASSISTANCE Grants for Construction of Treatment Works... a grant for 100 percent of the cost, including planning and design costs, of modification or...) The RBC failure has significantly increased the project's capital or operation and maintenance costs...

  20. 40 CFR 35.2035 - Rotating biological contractor (RBC) replacement grants.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... AND OTHER FEDERAL ASSISTANCE STATE AND LOCAL ASSISTANCE Grants for Construction of Treatment Works... a grant for 100 percent of the cost, including planning and design costs, of modification or...) The RBC failure has significantly increased the project's capital or operation and maintenance costs...

  1. A high-performance polydimethylsiloxane electrospun membrane for cell culture in lab-on-a-chip.

    PubMed

    Moghadas, Hajar; Saidi, Mohammad Said; Kashaninejad, Navid; Nguyen, Nam-Trung

    2018-03-01

    Thin porous membranes are important components in a microfluidic device, serving as separators, filters, and scaffolds for cell culture. However, the fabrication and the integration of these membranes possess many challenges, which restrict their widespread applications. This paper reports a facile technique to fabricate robust membrane-embedded microfluidic devices. We integrated an electrospun membrane into a polydimethylsiloxane (PDMS) device using the simple plasma-activated bonding technique. To increase the flexibility of the membrane and to address the leakage problem, the electrospun membrane was fabricated with the highest weight ratio of PDMS to polymethylmethacrylate (i.e., 6:1 w/w). The membrane-integrated microfluidic device could withstand a flow rate of up to 50  μ l/min. As a proof of concept, we demonstrated that such a compartmentalized microfluidic platform could be successfully used for cell culture with the capability of providing a more realistic in vivo -like condition. Human lung cancer epithelial cells (A549) were seeded on the membrane from the top microchannel, while the continuous flow of the culture medium through the bottom microchannel provided a shear-free cell culture condition. The tortuous micro-/nanofibers of the membrane immobilized the cells within the hydrophobic micropores and with no need of extracellular matrix for cell adhesion and cell growth. The hydrophobic surface conditions of the membrane were suitable for anchorage-independent cell types. To further extend the application of the device, we qualitatively showed that rinsing the membrane with ethanol prior to cell seeding could temporarily render the membrane hydrophilic and the platform could also be used for anchorage-dependent cells. Due to the three-dimensional (3D) topography of the membranes, three different configurations were observed, including individual single cells, monolayer cells, and 3D cell clusters. This cost-effective and robust compartmentalized

  2. In vitro membrane protein synthesis inside Sec translocon-reconstituted cell-sized liposomes

    PubMed Central

    Ohta, Naoki; Kato, Yasuhiko; Watanabe, Hajime; Mori, Hirotada; Matsuura, Tomoaki

    2016-01-01

    Protein synthesis using an in vitro transcription-translation system (IVTT) inside cell-sized liposomes has become a valuable tool to study the properties of biological systems under cell-mimicking conditions. However, previous liposome systems lacked the machinery for membrane protein translocation. Here, we reconstituted the translocon consisting of SecYEG from Escherichia coli inside cell-sized liposomes. The cell-sized liposomes also carry the reconstituted IVTT, thereby providing a cell-mimicking environment for membrane protein synthesis. By using EmrE, a multidrug transporter from E. coli, as a model membrane protein, we found that both the amount and activity of EmrE synthesized inside the liposome is increased approximately three-fold by incorporating the Sec translocon. The topological change of EmrE induced by the translocon was also identified. The membrane integration of 6 out of 9 E. coli inner membrane proteins that was tested was increased by incorporation of the translocon. By introducing the Sec translocon, the membrane integration efficiency of the membrane protein of interest was increased, and enabled the integration of membrane proteins that otherwise cannot be inserted. In addition, this work represents an essential step toward the construction of an artificial cell through a bottom-up approach. PMID:27808179

  3. Choroid plexus epithelial cells express the adhesion protein P-cadherin at cell-cell contacts and syntaxin-4 in the luminal membrane domain.

    PubMed

    Christensen, Inga Baasch; Mogensen, Esben Nees; Damkier, Helle Hasager; Praetorius, Jeppe

    2018-05-01

    The choroid plexus epithelial cells (CPECs) belong to a small group of polarized cells, where the Na + -K + -ATPase is expressed in the luminal membrane. The basic polarity of the cells is, therefore, still debated. We investigated the subcellular distribution of an array of proteins known to play fundamental roles either in establishing and maintaining basic cell polarity or in the polarized delivery and recycling of plasma membrane proteins. Immunofluorescence histochemical analysis was applied to determine the subcellular localization of apical and basolateral membrane determinants. Mass spectrometry analysis of CPECs isolated by fluorescence-activated cell sorting was applied to determine the expression of specific forms of the proteins. CPECs mainly express the cell-adhesive P-cadherin, which is localized to the lateral membranes. Proteins belonging to the Crumbs and partitioning defective (Par) protein complexes were all localized to the luminal membrane domain. Par-1 and the Scribble complex were localized to the basolateral membrane domain. Lethal(2) giant larvae homolog 2 (Lgl2) labeling was preferentially observed in the luminal membrane domain. Phosphatidylinositol 3,4,5-trisphosphate (PIP 3 ) was immunolocalized to the basolateral membrane domain, while phosphatidylinositol 4,5-bisphosphate (PIP 2 ) staining was most prominent in the luminal membrane domain along with the PIP 3 phosphatase, Pten. The apical target-SNARE syntaxin-3 and the basolateral target-SNARE syntaxin-4 were both localized to the apical membrane domain in CPECs, which lack cellular expression of the clathrin adaptor protein AP-1B for basolateral protein recycling. In conclusion, the CPECs are conventionally polarized, but express P-cadherin at cell-cell contacts, and Lgl2 and syntaxin-4 in the luminal plasma membrane domain.

  4. Cell membrane temperature rate sensitivity predicted from the Nernst equation.

    PubMed

    Barnes, F S

    1984-01-01

    A hyperpolarized current is predicted from the Nernst equation for conditions of positive temperature derivatives with respect to time. This ion current, coupled with changes in membrane channel conductivities, is expected to contribute to a transient potential shift across the cell membrane for silent cells and to a change in firing rate for pacemaker cells.

  5. The stress-free shape of the red blood cell membrane.

    PubMed Central

    Fischer, T M; Haest, C W; Stöhr-Liesen, M; Schmid-Schönbein, H; Skalak, R

    1981-01-01

    The two main proposals found in the literature for the stress-free shape of the red cell membrane are (a) the bioconcave shape and (b) the sphere of the same surface area. These possibilities are evaluated in this paper using theoretical modeling of equilibrium membrane shapes according to Zarda et al. (1977. J. Biomech. 10:211-221) and by comparison to experiments on red cells whose membrane shear modulus has been increased by treatment with diamide. Neither proposal is found to be compatible with all the experimental behaviour of native red cells. Neither proposal is found to be compatible with all the experimental behaviour of native red cells. To account for this discrepancy we propose that either the shear modulus of the native membrane is dependent on the membrane strain or that the bending stiffness is higher than estimated by Evans (1980. Biophys. J. 30:265-286). These studies suggest that the bioconcave disk is the more likely possibility for the stress-free shape. Images FIGURE 4 FIGURE 5 FIGURE 6 FIGURE 7 PMID:7248469

  6. Influence of the membrane ion exchange capacity on the catalyst layer of proton exchange membrane fuel cell

    NASA Astrophysics Data System (ADS)

    Navessin, Titichai

    2005-07-01

    This work investigated the effect of ion exchange capacity (IEC) of polymer electrolyte membranes (PEM) on the PEM fuel cell cathode catalyst layer. A series of radiation grafted ethylene tetrafluoroethylene-g-polystyrene sulfonic acid (ETFE-g-PSSA) membranes was used to provide a systematic variation of IEC. A method to fabricate gas diffusion electrodes (GDEs) was adapted and custom-made GDEs with known compositions were prepared. Oxygen electrochemistry, mass transport properties, water absorption behaviour and proton conductivity were studied in relation to the IEC. Electrochemical characterization including cyclic voltammetry, electrochemical impedance spectroscopy and linear sweep voltammetry were employed. The agglomerate model for cathodes was adapted and used to extract mass transport parameters from experimental results. Prior to investigation in fuel cell systems, studies were performed in a half-fuel cell, which simplified complicating parameters associated with fuel cell operation. It was found that membranes with higher IEC resulted in a higher active surface area of electrode. In contrast, they exhibited lower oxygen reduction performance. The extracted effective diffusion coefficient of oxygen and O2 solubility in the catalyst layer was used to estimate the extent of flooding, which revealed that ˜67--70% of void space was filled with water. The membrane's IEC regulates the extent of flooding of the cathode, which in turn affects its electrochemical characteristics. The investigation under operating fuel cell conditions revealed an increase in fuel cell performance with increasing IEC---a contradicting trend to that found for the half-fuel cell. This is explained by the interplay of electroosmotic flux and hydraulic counterflux in the membrane which affects water management in the membrane electrode assembly (MEA). The influence was most significant in the cathode catalyst layer, where it affects mass transport and electrochemical characteristics

  7. Red blood cell alloimmunization among sickle cell Kuwaiti Arab patients who received red blood cell transfusion.

    PubMed

    Ameen, Reem; Al Shemmari, Salem; Al-Bashir, Abdulaziz

    2009-08-01

    Sickle cell disease (SCD) is common in the Arabian Gulf region. Most cases require a red blood cell (RBC) transfusion, increasing the potential for RBC alloantibody development. The incidence of RBC alloimmunization among Kuwaiti Arab SCD patients is not yet known. This study retrospectively assessed the effect of using two different matching protocols on the incidence of alloimmunization among multiply transfused Kuwaiti Arab SCD patients. A total of 233 Kuwaiti Arab SCD patients were divided into two groups: Group 1 (n = 110) received RBC transfusion through standard ABO- and D-matched nonleukoreduced blood; Group 2 (n = 123) received RBCs matched for ABO, Rh, and K1 poststorage-leukoreduced blood. Multivariate analysis was performed on the factors associated with RBC alloimmunization and antibody specificity. Sixty-five percent of patients in Group 1 developed clinically significant RBC alloantibody with an increased prevalence in females; in patients in Group 2, 23.6% developed RBC alloantibodies (p = 0.01). In Group 1, 72 patients (65.5%) had alloantibodies directed against Rh and Kell systems (p = 0.01). Multivariate analysis further confirmed the results, showing that blood transfusion type and sex have significant effects on the rate of alloimmunizations. This study confirms the importance of selecting RBCs matched for Rh and Kell to reduce the risk of alloimmunizations among Kuwaiti Arab SCD patients.

  8. Nanocomposite membranes based on polybenzimidazole and ZrO2 for high-temperature proton exchange membrane fuel cells.

    PubMed

    Nawn, Graeme; Pace, Giuseppe; Lavina, Sandra; Vezzù, Keti; Negro, Enrico; Bertasi, Federico; Polizzi, Stefano; Di Noto, Vito

    2015-04-24

    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.

  9. A Red Blood Cell Membrane-Camouflaged Nanoparticle Counteracts Streptolysin O-Mediated Virulence Phenotypes of Invasive Group A Streptococcus

    PubMed Central

    Escajadillo, Tamara; Olson, Joshua; Luk, Brian T.; Zhang, Liangfang; Nizet, Victor

    2017-01-01

    Group A Streptococcus (GAS), an important human-specific Gram-positive bacterial pathogen, is associated with a broad spectrum of disease, ranging from mild superficial infections such as pharyngitis and impetigo, to serious invasive infections including necrotizing fasciitis and streptococcal toxic shock syndrome. The GAS pore-forming streptolysin O (SLO) is a well characterized virulence factor produced by nearly all GAS clinical isolates. High level expression of SLO is epidemiologically linked to intercontinental dissemination of hypervirulent clonotypes and poor clinical outcomes. SLO can trigger macrophage and neutrophil cell death and/or the inactivation of immune cell functions, and promotes tissue injury and bacterial survival in animal models of infection. In the present work, we describe how the pharmacological presentation of red blood cell (RBC) derived biomimetic nanoparticles (“nanosponges”) can sequester SLO and block the ability of GAS to damage host cells, thereby preserving innate immune function and increasing bacterial clearance in vitro and in vivo. Nanosponge administration protected human neutrophils, macrophages, and keratinocytes against SLO-mediated cytotoxicity. This therapeutic intervention prevented SLO-induced macrophage apoptosis and increased neutrophil extracellular trap formation, allowing increased GAS killing by the respective phagocytic cell types. In a murine model of GAS necrotizing skin infection, local administration of the biomimetic nanosponges was associated with decreased lesion size and reduced bacterial colony-forming unit recovery. Utilization of a toxin decoy and capture platform that inactivates the secreted SLO before it contacts the host cell membrane, presents a novel virulence factor targeted strategy that could be a powerful adjunctive therapy in severe GAS infections where morbidity and mortality are high despite antibiotic treatment. PMID:28769806

  10. Plasma Membrane Sterol Distribution Resembles the Surface Topography of Living Cells

    PubMed Central

    2007-01-01

    Cholesterol is an important constituent of cellular membranes. It has been suggested that cholesterol segregates into sterol-rich and -poor domains in the plasma membrane, although clear evidence for this is lacking. By fluorescence imaging of the natural sterol dehydroergosterol (DHE), the lateral sterol distribution has been visualized in living cells. The spatial labeling pattern of DHE coincided with surface structures such as ruffles, microvilli, and filopodia with correlation lengths in the range of 0.8–2.5 μm. DHE staining of branched tubules and of nanotubes connecting two cells was detected. Dynamics of DHE in folded and plane membrane regions was comparable as determined by fluorescence recovery after photobleaching. DHE colocalized with fluid membrane-preferring phospholipids in surface structures and at sites of cell attachment as well as in the cleavage furrow of dividing cells, but it was not particularly enriched in those regions. Fluorescent sterol showed homogeneous staining in membrane blebs induced by F-actin disruption. Cross-linking the ganglioside GM1—a putative raft marker—did not affect the cell surface distribution of DHE. The results suggest that spatial heterogeneities of plasma membrane staining of DHE resolvable by light microscopy reflect the cell surface topography but not phase-separated sterol domains in the bilayer plane. PMID:17065557

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

    PubMed

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

    2016-06-01

    Multipotent progenitor cells were mobilized during pediatric extracorporeal membrane oxygenation (ECMO). We hypothesize that these cells also adhered onto polymethylpentene (PMP) fibers within the membrane oxygenator (MO) during adult ECMO support. Mononuclear cells were removed from the surface of explanted PMP-MOs (n = 16). Endothelial-like outgrowth and mesenchymal-like cells were characterized by flow cytometric analysis using different surface markers. Spindle-shaped attaching cells were identified early, but without proliferative activity. After long-term cultivation palisading type or cobblestone-type outgrowth cells with high proliferative activity appeared and were characterized as (i) leukocytoid CD45+/CD31+ (CD133+/VEGFR-II+/CD90+/CD14+/CD146dim/CD105dim); (ii) endothelial-like CD45-/CD31+ (VEGF-RII+/CD146+/CD105+/CD133-/CD14-/CD90-); and (iii) mesenchymal-like cells CD45-/CD31- (CD105+/CD90+/CD133dim/VEGFR-II-/CD146-/CD14-). The distribution of the cell populations depended on the MO and cultivation time. Endothelial-like cells formed capillary-like structures and did uptake Dil-acetylated low-density lipoprotein. Endothelial- and mesenchymal-like cells adhered on the surface of PMP-MOs. Further research is needed to identify the clinical relevance of these cells. Copyright © 2015 The Authors. Artificial Organs published by Wiley Periodicals, Inc. on behalf of International Center for Artificial Organs and Transplantation (ICAOT).

  12. Of macrophages and red blood cells; a complex love story.

    PubMed

    de Back, Djuna Z; Kostova, Elena B; van Kraaij, Marian; van den Berg, Timo K; van Bruggen, Robin

    2014-01-01

    Macrophages tightly control the production and clearance of red blood cells (RBC). During steady state hematopoiesis, approximately 10(10) RBC are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with interactions that are essential for erythroid development. New evidence suggests that not only under homeostasis but also under stress conditions, macrophages play an important role in promoting erythropoiesis. Once RBC have matured, these cells remain in circulation for about 120 days. At the end of their life span, RBC are cleared by macrophages residing in the spleen and the liver. Current theories about the removal of senescent RBC and the essential role of macrophages will be discussed as well as the role of macrophages in facilitating the removal of damaged cellular content from the RBC. In this review we will provide an overview on the role of macrophages in the regulation of RBC production, maintenance and clearance. In addition, we will discuss the interactions between these two cell types during transfer of immune complexes and pathogens from RBC to macrophages.

  13. Effects of cholesterol depletion on membrane nanostructure in MCF-7 cells by atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Yuhua; Jiang, Ningcheng; Shi, Aisi; Zheng, Liqin; Yang, Hongqin; Xie, Shusen

    2017-02-01

    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.

  14. Selective effect of cell membrane on synaptic neurotransmission

    NASA Astrophysics Data System (ADS)

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

    2016-01-01

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

  15. Chitosan and alginate types of bio-membrane in fuel cell application: An overview

    NASA Astrophysics Data System (ADS)

    Shaari, N.; Kamarudin, S. K.

    2015-09-01

    The major problems of polymer electrolyte membrane fuel cell technology that need to be highlighted are fuel crossovers (e.g., methanol or hydrogen leaking across fuel cell membranes), CO poisoning, low durability, and high cost. Chitosan and alginate-based biopolymer membranes have recently been used to solve these problems with promising results. Current research in biopolymer membrane materials and systems has focused on the following: 1) the development of novel and efficient biopolymer materials; and 2) increasing the processing capacity of membrane operations. Consequently, chitosan and alginate-based biopolymers seek to enhance fuel cell performance by improving proton conductivity, membrane durability, and reducing fuel crossover and electro-osmotic drag. There are four groups of chitosan-based membranes (categorized according to their reaction and preparation): self-cross-linked and salt-complexed chitosans, chitosan-based polymer blends, chitosan/inorganic filler composites, and chitosan/polymer composites. There are only three alginate-based membranes that have been synthesized for fuel cell application. This work aims to review the state-of-the-art in the growth of chitosan and alginate-based biopolymer membranes for fuel cell applications.

  16. S-layer and cytoplasmic membrane - exceptions from the typical archaeal cell wall with a focus on double membranes.

    PubMed

    Klingl, Andreas

    2014-01-01

    The common idea of typical cell wall architecture in archaea consists of a pseudo-crystalline proteinaceous surface layer (S-layer), situated upon the cytoplasmic membrane. This is true for the majority of described archaea, hitherto. Within the crenarchaea, the S-layer often represents the only cell wall component, but there are various exceptions from this wall architecture. Beside (glycosylated) S-layers in (hyper)thermophilic cren- and euryarchaea as well as halophilic archaea, one can find a great variety of other cell wall structures like proteoglycan-like S-layers (Halobacteria), glutaminylglycan (Natronococci), methanochondroitin (Methanosarcina) or double layered cell walls with pseudomurein (Methanothermus and Methanopyrus). The presence of an outermost cellular membrane in the crenarchaeal species Ignicoccus hospitalis already gave indications for an outer membrane similar to Gram-negative bacteria. Although there is just limited data concerning their biochemistry and ultrastructure, recent studies on the euryarchaeal methanogen Methanomassiliicoccus luminyensis, cells of the ARMAN group, and the SM1 euryarchaeon delivered further examples for this exceptional cell envelope type consisting of two membranes.

  17. Label-free evanescent microscopy for membrane nano-tomography in living cells.

    PubMed

    Bon, Pierre; Barroca, Thomas; Lévèque-Fort, Sandrine; Fort, Emmanuel

    2014-11-01

    We show that through-the-objective evanescent microscopy (epi-EM) is a powerful technique to image membranes in living cells. Readily implementable on a standard inverted microscope, this technique enables full-field and real-time tracking of membrane processes without labeling and thus signal fading. In addition, we demonstrate that the membrane/interface distance can be retrieved with 10 nm precision using a multilayer Fresnel model. We apply this nano-axial tomography of living cell membranes to retrieve quantitative information on membrane invagination dynamics. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

  18. Elastic Properties of Pore-Spanning Apical Cell Membranes Derived from MDCK II Cells.

    PubMed

    Nehls, Stefan; Janshoff, Andreas

    2017-10-17

    The mechanical response of adherent, polarized cells to indentation is frequently attributed to the presence of an endogenous actin cortex attached to the inner leaflet of the plasma membrane. Here, we scrutinized the elastic properties of apical membranes separated from living cells and attached to a porous mesh in the absence of intracellular factors originating from the cytosol, organelles, the substrate, neighbors, and the nucleus. We found that a tension-based model describes the data very well providing essentially the prestress of the shell generated by adhesion of the apical membrane patches to the pore rim and the apparent area compressibility modulus, an intrinsic elastic modulus modulated by the surface excess stored in membrane reservoirs. Removal of membrane-associated proteins by proteases decreases the area compressibility modulus, whereas fixation and cross-linking of proteins with glutaraldehyde increases it. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  19. Proteomic identification of erythrocyte membrane protein deficiency in hereditary spherocytosis.

    PubMed

    Peker, Selen; Akar, Nejat; Demiralp, Duygu Ozel

    2012-03-01

    Hereditary spherocytosis (HS) is the most common congenital hemolytic anemia in Caucasians, with an estimated prevalence ranging from 1:2000 to 1:5000. The molecular defect in one of the erythrocytes (RBC) membrane proteins underlying HS like; spectrin-α, spectrin-β, ankyrin, band 3 and protein 4.2 that lead to membrane destabilization and vesiculation, may change the RBCs into denser and more rigid cells (spherocytes), which are removed by the spleen, leading to the development of hemolytic anemia. It is classified as mild, moderate and severe, according to the degree of the hemolytic anemia and the associated symptoms. Two-dimensional gel electrophoresis (2-DE) is potentially valuable method for studying heritable disorders as HS that involve membrane proteins. This separation technique of proteins based upon two biophysically unrelated parameters; molecular weight and charge, is a good option in clinical proteomics in terms of ability to separate complex mixtures, display post-translational modifications and changes after phosphorylation. In this study, we have used contemporary methods with some modifications for the solubilisation, separation and identification of erythrocyte membrane proteins in normal and in HS RBCs. Spectrin alpha and beta chain, ankyrin and band 3 proteins expression differences were found with PDQuest software 8.0.1. and peptide mass fingerprinting (PMF) analysis performed for identification of proteins in this study.

  20. A comprehensive framework for functional diversity patterns of marine chromophytic phytoplankton using rbcL phylogeny

    PubMed Central

    Samanta, Brajogopal; Bhadury, Punyasloke

    2016-01-01

    Marine chromophytes are taxonomically diverse group of algae and contribute approximately half of the total oceanic primary production. To understand the global patterns of functional diversity of chromophytic phytoplankton, robust bioinformatics and statistical analyses including deep phylogeny based on 2476 form ID rbcL gene sequences representing seven ecologically significant oceanographic ecoregions were undertaken. In addition, 12 form ID rbcL clone libraries were generated and analyzed (148 sequences) from Sundarbans Biosphere Reserve representing the world’s largest mangrove ecosystem as part of this study. Global phylogenetic analyses recovered 11 major clades of chromophytic phytoplankton in varying proportions with several novel rbcL sequences in each of the seven targeted ecoregions. Majority of OTUs was found to be exclusive to each ecoregion, whereas some were shared by two or more ecoregions based on beta-diversity analysis. Present phylogenetic and bioinformatics analyses provide a strong statistical support for the hypothesis that different oceanographic regimes harbor distinct and coherent groups of chromophytic phytoplankton. It has been also shown as part of this study that varying natural selection pressure on form ID rbcL gene under different environmental conditions could lead to functional differences and overall fitness of chromophytic phytoplankton populations. PMID:26861415

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

    NASA Astrophysics Data System (ADS)

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

    2016-01-01

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

  2. Composite polymer membranes for proton exchange membrane fuel cells operating at elevated temperatures and reduced humidities

    NASA Astrophysics Data System (ADS)

    Zhang, Tao

    Proton Exchange Membrane Fuel Cells (PEMFCs) are the leading candidate in the fuel cell technology due to the high power density, solid electrolyte, and low operational temperature. However, PEMFCs operating in the normal temperature range (60-80°C) face problems including poor carbon monoxide tolerance and heat rejection. The poisoning effect can be significantly relieved by operating the fuel cell at elevated temperature, which also improves the heat rejection and electrochemical kinetics. Low relative humidity (RH) operation is also desirable to simplify the reactant humidification system. However, at elevated temperatures, reduced RH PEMFC performance is seriously impaired due to irreversible water loss from presently employed state-of-the-art polymer membrane, Nafion. This thesis focuses on developing polymer electrolyte membranes with high water retention ability for operation in elevated temperature (110-150°C), reduced humidity (˜50%RH) PEMFCs. One approach is to alter Nafion by adding inorganic particles such as TiO2, SiO2, Zr(HPO 4)2, etc. While the presence of these materials in Nafion has proven beneficial, a reduction or no improvement in the PEMFC performance of Nafion/TiO2 and Nafion/Zr(HPO4)2 membranes is observed with reduced particle sizes or increased particle loadings in Nafion. It is concluded that the PEMFC performance enhancement associated with addition of these inorganic particles was not due to the particle hydrophilicity. Rather, the particle, partially located in the hydrophobic region of the membrane, benefits the cell performance by altering the membrane structure. Water transport properties of some Nafion composite membranes were investigated by NMR methods including pulsed field gradient spin echo diffusion, spin-lattice relaxation, and spectral measurements. Compared to unmodified Nafion, composite membranes materials exhibit longer longitudinal relaxation time constant T1. In addition to the Nafion material, sulfonated styrene

  3. Hydrogen-oxygen proton-exchange membrane fuel cells and electrolyzers

    NASA Technical Reports Server (NTRS)

    Baldwin, R.; Pham, M.; Leonida, A.; Mcelroy, J.; Nalette, T.

    1989-01-01

    Hydrogen-oxygen SPE fuel cells and SPE electrolyzers (products of Hamilton Standard) both use a Proton-Exchange Membrane (PEM) as the sole electrolyte. The SPE cells have demonstrated a ten year life capability under load conditions. Ultimate life of PEM fuel cells and electrolyzers is primarily related to the chemical stability of the membrane. For perfluorocarbon proton-exchange membranes an accurate measure of the membrane stability is the fluoride loss rate. Millions of cell hours have contributed to establishing a relationship between fluroride loss rates and average expected ultimate cell life. Several features were introduced into SPE fuel cells and SPE electrolyzers such that applications requiring greater than or equal to 100,000 hours of life can be considered. Equally important as the ultimate life is the voltage stability of hydrogen-oxygen fuel cells and electrolyzers. Here again the features of SPE fuel cells and SPE electrolyzers have shown a cell voltage stability in the order of 1 microvolt per hour. That level of stability were demonstrated for tens of thousands of hours in SPE fuel cells at up to 500 amps per square foot (ASF) current density. The SPE electrolyzers have demonstrated the same at 1000 ASF. Many future extraterrestrial applications for fuel cells require that they be self recharged. To translate the proven SPE cell life and stability into a highly reliable extraterrestrial electrical energy storage system, a simplification of supporting equipment is required. Static phase separation, static fluid transport and static thermal control will be most useful in producting required system reliability. Although some 200,000 SPE fuel cell hours were recorded in earth orbit with static fluid phase separation, no SPE electrolyzer has, as yet, operated in space.

  4. Arenavirus budding resulting from viral-protein-associated cell membrane curvature

    PubMed Central

    Schley, David; Whittaker, Robert J.; Neuman, Benjamin W.

    2013-01-01

    Viral replication occurs within cells, with release (and onward infection) primarily achieved through two alternative mechanisms: lysis, in which virions emerge as the infected cell dies and bursts open; or budding, in which virions emerge gradually from a still living cell by appropriating a small part of the cell membrane. Virus budding is a poorly understood process that challenges current models of vesicle formation. Here, a plausible mechanism for arenavirus budding is presented, building on recent evidence that viral proteins embed in the inner lipid layer of the cell membrane. Experimental results confirm that viral protein is associated with increased membrane curvature, whereas a mathematical model is used to show that localized increases in curvature alone are sufficient to generate viral buds. The magnitude of the protein-induced curvature is calculated from the size of the amphipathic region hypothetically removed from the inner membrane as a result of translation, with a change in membrane stiffness estimated from observed differences in virion deformation as a result of protein depletion. Numerical results are based on experimental data and estimates for three arenaviruses, but the mechanisms described are more broadly applicable. The hypothesized mechanism is shown to be sufficient to generate spontaneous budding that matches well both qualitatively and quantitatively with experimental observations. PMID:23864502

  5. Alloimmunization is associated with older age of transfused red blood cells in sickle cell disease.

    PubMed

    Desai, Payal C; Deal, Allison M; Pfaff, Emily R; Qaqish, Bahjat; Hebden, Leyna M; Park, Yara A; Ataga, Kenneth I

    2015-08-01

    Red blood cell (RBC) alloimmunization is a significant clinical complication of sickle cell disease (SCD). It can lead to difficulty with cross-matching for future transfusions and may sometimes trigger life-threatening delayed hemolytic transfusion reactions. We conducted a retrospective study to explore the association of clinical complications and age of RBC with alloimmunization in patients with SCD followed at a single institution from 2005 to 2012. One hundred and sixty six patients with a total of 488 RBC transfusions were evaluated. Nineteen patients (11%) developed new alloantibodies following blood transfusions during the period of review. The median age of RBC units was 20 days (interquartile range: 14-27 days). RBC antibody formation was significantly associated with the age of RBC units (P = 0.002), with a hazard ratio of 3.5 (95% CI: 1.71-7.11) for a RBC unit that was 7 days old and 9.8 (95% CI: 2.66-35.97) for a unit that was 35 days old, 28 days after the blood transfusion. No association was observed between RBC alloimmunization and acute vaso-occlusive complications. Although increased echocardiography-derived tricuspid regurgitant jet velocity (TRV) was associated with the presence of RBC alloantibodies (P = 0.02), TRV was not significantly associated with alloimmunization when adjusted for patient age and number of transfused RBC units. Our study suggests that RBC antibody formation is significantly associated with older age of RBCs at the time of transfusion. Prospective studies in patients with SCD are required to confirm this finding. © 2015 Wiley Periodicals, Inc.

  6. Basolateral membrane K+ channels in renal epithelial cells

    PubMed Central

    Devor, Daniel C.

    2012-01-01

    The major function of epithelial tissues is to maintain proper ion, solute, and water homeostasis. The tubule of the renal nephron has an amazingly simple structure, lined by epithelial cells, yet the segments (i.e., proximal tubule vs. collecting duct) of the nephron have unique transport functions. The functional differences are because epithelial cells are polarized and thus possess different patterns (distributions) of membrane transport proteins in the apical and basolateral membranes of the cell. K+ channels play critical roles in normal physiology. Over 90 different genes for K+ channels have been identified in the human genome. Epithelial K+ channels can be located within either or both the apical and basolateral membranes of the cell. One of the primary functions of basolateral K+ channels is to recycle K+ across the basolateral membrane for proper function of the Na+-K+-ATPase, among other functions. Mutations of these channels can cause significant disease. The focus of this review is to provide an overview of the basolateral K+ channels of the nephron, providing potential physiological functions and pathophysiology of these channels, where appropriate. We have taken a “K+ channel gene family” approach in presenting the representative basolateral K+ channels of the nephron. The basolateral K+ channels of the renal epithelia are represented by members of the KCNK, KCNJ, KCNQ, KCNE, and SLO gene families. PMID:22338089

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

    PubMed

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

    2015-04-21

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

  8. Review of cell performance in anion exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Dekel, Dario R.

    2018-01-01

    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.

  9. Membrane protein stoichiometry studied in intact mammalian cells using liquid-phase electron microscopy.

    PubMed

    DE Jonge, N

    2018-02-01

    Receptor membrane proteins in the plasma membranes of cells respond to extracellular chemical signals by conformational changes, spatial redistribution, and (re-)assembly into protein complexes, for example, into homodimers (pairs of the same protein type). The functional state of the proteins can be determined from information about how subunits are assembled into protein complexes. Stoichiometric information about the protein complex subunits, however, is generally not obtained from intact cells but from pooled material extracted from many cells, resulting in a lack of fundamental knowledge about the functioning of membrane proteins. First, functional states may dramatically differ from cell to cell on account of cell heterogeneity. Second, extracting the membrane proteins from the plasma membrane may lead to many artefacts. Liquid-phase scanning transmission electron microscopy (STEM), in short liquid STEM, is a new technique capable of determining the locations of individual membrane proteins within the intact plasma membranes of cells in liquid. Many tens of whole cells can readily be imaged. It is possible to analyse the stoichiometry of membrane proteins in single cells while accounting for heterogenic cell populations. Liquid STEM was used to image epidermal growth factor receptors in whole COS7 cells. A study of the dimerisation of the HER2 protein in breast cancer cells revealed the presence of rare cancer cells in which HER2 was in a different functional state than in the bulk cells. Stoichiometric information about receptors is essential not only for basic science but also for biomedical application because they present many important pharmaceutical targets. © 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.

  10. Comparison of human mesenchymal stromal cells from four neonatal tissues: Amniotic membrane, chorionic membrane, placental decidua and umbilical cord.

    PubMed

    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

    2017-05-01

    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.

  11. Analysis of the 3D structure og agglutinated erythrocyte using CellScan and confocal microscopy: characterization by FLIM-FRET

    NASA Astrophysics Data System (ADS)

    Riquelme, Bibiana D.; Dumas, Dominique; Valverde de Rasia, Juana; Rasia, Rodolfo J.; Stoltz, Jean Francois

    2003-10-01

    We report the adhesion of human erythrocyte membranes mediated by monoclonal antibodies anti-glycophorin. The distribution of the linked antibodies on membrane was identified with selective fluorescence labels. To analyze the antibody distribution on interfacial region between two cells agglutinated and on its surface, three types of fluorescence marked strategy were evaluated. The 3D images were obtained in a CellScan and Confocal Laser Scanning Microscopy CLSM. We considered the FRET signal to characterize the agglutination of Red Blood Cells (RBC) by specific monoclonal antibodies (anti-glycophorin A or B). The fluorescence labeling demonstrated that distribution of antibody on erythrocyte membranes is not homogeneous. The fluorescence intensity on contact region in the agglutinated is bigger than the intensity on exterior surface. Tentatively, we interpreted these intensity differences in terms of the mobility of antibody linked to the glycocalix on cell surface. Such mobility has a large consequence in the morphology of cellular agglutinated.

  12. Electroporating Fields Target Oxidatively Damaged Areas in the Cell Membrane

    PubMed Central

    Vernier, P. Thomas; Levine, Zachary A.; Wu, Yu-Hsuan; Joubert, Vanessa; Ziegler, Matthew J.; Mir, Lluis M.; Tieleman, D. Peter

    2009-01-01

    Reversible electropermeabilization (electroporation) is widely used to facilitate the introduction of genetic material and pharmaceutical agents into living cells. Although considerable knowledge has been gained from the study of real and simulated model membranes in electric fields, efforts to optimize electroporation protocols are limited by a lack of detailed understanding of the molecular basis for the electropermeabilization of the complex biomolecular assembly that forms the plasma membrane. We show here, with results from both molecular dynamics simulations and experiments with living cells, that the oxidation of membrane components enhances the susceptibility of the membrane to electropermeabilization. Manipulation of the level of oxidative stress in cell suspensions and in tissues may lead to more efficient permeabilization procedures in the laboratory and in clinical applications such as electrochemotherapy and electrotransfection-mediated gene therapy. PMID:19956595

  13. Membrane Crossing and Membranotropic Activity of Cell-Penetrating Peptides: Dangerous Liaisons?

    PubMed

    Walrant, Astrid; Cardon, Sébastien; Burlina, Fabienne; Sagan, Sandrine

    2017-12-19

    Living organisms have to maintain a stable balance in molecules and ions in the changing environment in which they are living, a process known as homeostasis. At the level of cells, the plasma membrane has a major role in homeostasis, since this hydrophobic film prevents passive diffusion of large and hydrophilic molecules between the extracellular and intracellular milieu. Living organisms have evolved with highly sophisticated transport systems to control exchanges across this barrier: import of nutrients and fuel essential for their survival; recognition of chemical or physical messengers allowing information interchanges with surrounding cells. Besides specialized proteins, endocytosis mechanisms at the level of the lipid bilayer can transport molecules from the outside across the cell membrane, in an energy-dependent manner. The cell membrane is highly heterogeneous in its molecular composition (tens of different lipids, proteins, polysaccharides, and combinations of these) and dynamic with bending, deformation, and elastic properties that depend on the local composition of membrane domains. Many viruses, microorganisms, and toxins exploit the plasma membrane to enter into cells. Chemists develop strategies to target the plasma membrane with molecules capable of circumventing this hydrophobic barrier, in particular to transport and deliver nonpermeable drugs in cells for biotechnological or pharmaceutical purposes. Drug delivery systems are numerous and include lipid-, sugar-, protein-, and peptide-based delivery systems, since these biomolecules generally have good biocompatibility, biodegradability, environmental sustainability, cost effectiveness, and availability. Among those, cell-penetrating peptides (CPPs), reported for the first time in the early 1990s, are attracting major interest not only as potential drug delivery systems but also at the level of fundamental research. It was indeed demonstrated very early that these peptides, which generally

  14. Endothelial cell membrane vesicles in the study of organ preference of metastasis.

    PubMed

    Johnson, R C; Augustin-Voss, H G; Zhu, D Z; Pauli, B U

    1991-01-01

    Many malignancies exhibit distinct patterns of metastasis that appear to be mediated by receptor/ligand-like interactions between tumor cells and organ-specific vascular endothelium. In order to study endothelial cell surface molecules involved in the binding of metastatic cells, we developed a perfusion method to isolate outside-out membrane vesicles from the lumenal surface of rat lung microvascular endothelium. Lungs were perfused in situ for 4 h at 37 degrees C with a solution of 100 mM formaldehyde, 2 mM dithiothreitol in phosphate-buffered saline to induce endothelial cell vesiculation. Radioiodinated rat lung endothelial cell membrane vesicles bound lung-metastatic tumor cells (B16F10, R323OAC-MET) in significantly higher numbers than their low or nonmetastatic counterparts (B16F0, R323OAC-LR). In contrast, leg endothelial membrane vesicle showed no binding preference for either cell line. Neuraminidase treatment of vesicles abolished specificity of adhesion of lung-derived vesicles to lung metastatic tumor cells. These results demonstrate that in situ perfusion is an appropriate technique to obtain pure endothelial cell membrane vesicles containing functionally active adhesion molecules. The preferential binding of lung-derived endothelial cell membrane vesicles by lung metastatic tumor cells is evidence of the importance of endothelial cell adhesion molecules in the formation of metastases.

  15. Biophysical Studies of Nanosecond Pulsed Electric Field Induced Cell Membrane Permeabilization

    NASA Astrophysics Data System (ADS)

    Wu, Yu-Hsuan

    Nanosecond megavolts-per-meter pulsed electric field (nsPEF) offers a non-invasive manipulation of intracellular organelles and functions of biological cells. Accordingly, nsPEF is a potential technique for biophysical research and cancer therapy, and is of growing interest. Although, the application of nsPEF has shown electroperturbation on cell plasma membranes and intracellular membranes as well, the mechanisms underlying the electropermeabilization are still not clear. In this thesis, we systematically study nsPEFs (5 and 30 ns) induced membrane permeability change in biological cell in-vitro with different pulse parameters. In Chapter 3, we investigate the nsPEF-induced intracellular membrane permeabilization of mitochondria which play key roles in activating apoptosis in mammalian cells. The results show the evidences of nsPEF-induced membrane permeability increase in mitochondria, and suggest that nsPEF is a potential technology for cancer cell ablation without delivery of drug or gene into cells. In Chapter 2, 4 and 6, we study the properties of nsPEF-induced plasma membrane permeabilization. In the beginning, the change of plasma membrane permeability is studied by uptake of YO-PRO-1 and propidium iodide, fluorescent dyes specifically used as indicators of plasma membrane permeabilization. However, the detection is limited by the fluorescent emission efficiency and detector capability. To increase the detection sensitivity, we later develop a method based on cell volume change due to regulation of osmotic balance that causes water and small ions transport through plasma membrane. We find that even a single 10 MV/m pulse of 5 ns duration produces measureable cell swelling. The results demonstrate that cell swelling is susceptible to nsPEF and can detect membrane permeabilization more easily and precisely than fluorescent dyes. We compare the effects of different pulse parameters (pulse duration, pulse number, electric field amplitude and pulse repetition

  16. Cholera toxin activation of adenylate cyclase in cancer cell membrane fragments.

    PubMed Central

    Bitensky, M W; Wheeler, M A; Mehta, H; Miki, N

    1975-01-01

    Activation of adenylate [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] by cholera toxin (84,000 daltons, 5.5 S) is demonstrated in plasma membrane fragments of mouse ascites cancer cells. The activation of adenylate cyclase is mediated by a macromolecular cyclase activating factor (MCAF), which has a sedimentation constant of 2.7 S and a molecular weight of about 26,000. MCAF is derived from, and may be identical to the "A fragment" of cholera toxin. Generation of MCAF depends on prior interaction of cholera toxin with either dithiothreitol, NADH, NAD, or a low-molecular-weight component (less than 700 daltons) present in cytoplasm. Subsequent exposure of this pretreated cholera toxin to cell membranes from a variety of mouse ascites cancer cells is followed rapidly by the appearance of MCAF, which no longer requires dithiothreitol, NADH, or NAD for the activation of adenylate cyclase. Activation of adenylate cyclase by MCAF in ascites cancer cell membrane fragments is not reversed by repeated washing of these membrane fragments. Adenylate cyclase in normal cell membrane fragments fails to respond either to cholera toxin or MCAF in the presence of dithiothreitol. In striking contrast, the adenylate cyclase in membrane fragments from five ascites cancer cells responds to either MCAF or native cholera toxin preincubated with dithiothreitol, NADH, or NAD. PMID:1058474

  17. The role of basal cells in adhesion of columnar epithelium to airway basement membrane.

    PubMed

    Evans, M J; Plopper, C G

    1988-08-01

    In this report, we present a new concept of the role of the basal cell in airway epithelium. Previously, the basal cell was thought to be the progenitor cell for the columnar epithelium. However, several studies have shown that this concept may not be correct. The morphologic aspects of the basal cell suggest that it could play a role in adhesion of the columnar epithelium to the basement membrane. Basal cells form attachments with columnar cells (desmosomes) and with the basement membrane (hemidesmosomes). Columnar cells do not form hemidesmosome attachments with the basement membrane. Basal cells could strengthen the adhesion of columnar cells to the basement membrane by forming hemidesmosome attachments to the basement membrane and desmosome attachments with adjacent columnar cells. Incidental evidence from 2 existing publications concerning airway microanatomy support this concept. As columnar cells grow taller, the proportion of the cell surface in contact with the basement membrane becomes progressively smaller, and thus the cell surface area related to adhesion also becomes smaller. It was found that the number of basal cells per millimeter of basement membrane was closely related to the height of the columnar cell epithelium (r = 0.98), but not to the number of columnar cells (r = 0.42). The consistency of the relationship between increased columnar cell height (and thus decreased surface area for adhesion) and the number of basal cells present (r = 0.98) supports the concept that the basal cell plays a role in adhesion of columnar cells to the basement membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

  18. Isolation and Characterization of Canine Amniotic Membrane-Derived Multipotent Stem Cells

    PubMed Central

    Kim, Hyung-Sik; Kang, Kyung-Sun

    2012-01-01

    Recent studies have shown that amniotic membrane tissue is a rich source of stem cells in humans. In clinical applications, the amniotic membrane tissue had therapeutic effects on wound healing and corneal surface reconstruction. Here, we successfully isolated and identified multipotent stem cells (MSCs) from canine amniotic membrane tissue. We cultured the canine amniotic membrane-derived multipotent stem cells (cAM-MSCs) in low glucose DMEM medium. cAM-MSCs have a fibroblast-like shape and adhere to tissue culture plastic. We characterized the immunophenotype of cAM-MSCs by flow cytometry and measured cell proliferation by the cumulative population doubling level (CPDL). We performed differentiation studies for the detection of trilineage multipotent ability, under the appropriate culture conditions. Taken together, our results show that cAM-MSCs could be a rich source of stem cells in dogs. Furthermore, cAM-MSCs may be useful as a cell therapy application for veterinary regenerative medicine. PMID:23024756

  19. A FRET sensor enables quantitative measurements of membrane charges in live cells.

    PubMed

    Ma, Yuanqing; Yamamoto, Yui; Nicovich, Philip R; Goyette, Jesse; Rossy, Jérémie; Gooding, J Justin; Gaus, Katharina

    2017-04-01

    Membrane charge has a critical role in protein trafficking and signaling. However, quantification of the effective electrostatic potential of cellular membranes has remained challenging. We developed a fluorescence membrane charge sensor (MCS) that reports changes in the membrane charge of live cells via Förster resonance energy transfer (FRET). MCS is permanently attached to the inner leaflet of the plasma membrane and shows a linear, reversible and fast response to changes of the electrostatic potential. The sensor can monitor a wide range of cellular treatments that alter the electrostatic potential, such as incorporation and redistribution of charged lipids and alterations in cytosolic ion concentration. Applying the sensor to T cell biology, we used it to identify charged membrane domains in the immunological synapse. Further, we found that electrostatic interactions prevented spontaneous phosphorylation of the T cell receptor and contributed to the formation of signaling clusters in T cells.

  20. Electrical coupling between A17 cells enhances reciprocal inhibitory feedback to rod bipolar cells.

    PubMed

    Elgueta, Claudio; Leroy, Felix; Vielma, Alex H; Schmachtenberg, Oliver; Palacios, Adrian G

    2018-02-15

    A17 amacrine cells are an important part of the scotopic pathway. Their synaptic varicosities receive glutamatergic inputs from rod bipolar cells (RBC) and release GABA onto the same RBC terminal, forming a reciprocal feedback that shapes RBC depolarization. Here, using patch-clamp recordings, we characterized electrical coupling between A17 cells of the rat retina and report the presence of strongly interconnected and non-coupled A17 cells. In coupled A17 cells, evoked currents preferentially flow out of the cell through GJs and cross-synchronization of presynaptic signals in a pair of A17 cells is correlated to their coupling degree. Moreover, we demonstrate that stimulation of one A17 cell can induce electrical and calcium transients in neighboring A17 cells, thus confirming a functional flow of information through electrical synapses in the A17 coupled network. Finally, blocking GJs caused a strong decrease in the amplitude of the inhibitory feedback onto RBCs. We therefore propose that electrical coupling between A17 cells enhances feedback onto RBCs by synchronizing and facilitating GABA release from inhibitory varicosities surrounding each RBC axon terminal. GJs between A17 cells are therefore critical in shaping the visual flow through the scotopic pathway.

  1. Influence of zinc deficiency on cell-membrane fluidity in Jurkat, 3T3 and IMR-32 cells.

    PubMed Central

    Verstraeten, Sandra V; Zago, M Paola; MacKenzie, Gerardo G; Keen, Carl L; Oteiza, Patricia I

    2004-01-01

    We investigated whether zinc deficiency can affect plasma membrane rheology. Three cell lines, human leukaemia T-cells (Jurkat), rat fibroblasts (3T3) and human neuroblastoma cells (IMR-32), were cultured for 48 h in control medium, in zinc-deficient medium (1.5 microM zinc; 1.5 Zn), or in the zinc-deficient medium supplemented with 15 microM zinc (15 Zn). The number of viable cells was lower in the 1.5 Zn group than in the control and 15 Zn groups. The frequency of apoptosis was higher in the 1.5 Zn group than in the control and 15 Zn groups. Membrane fluidity was evaluated using the 6-(9-anthroyloxy)stearic acid and 16-(9-anthroyloxy)palmitic acid probes. Membrane fluidity was higher in 1.5 Zn cells than in the control cells; no differences were observed between control cells and 15 Zn cells. The effect of zinc deficiency on membrane fluidity at the water/lipid interface was associated with a higher phosphatidylserine externalization. The higher membrane fluidity in the hydrophobic region of the bilayer was correlated with a lower content of arachidonic acid. We suggest that the increased fluidity of the membrane secondary to zinc deficiency is in part due to a decrease in arachidonic acid content and the apoptosis-related changes in phosphatidylserine distribution. PMID:14629198

  2. Molecular dynamics simulations of heterogeneous cell membranes in response to uniaxial membrane stretches at high loading rates.

    PubMed

    Zhang, Lili; Zhang, Zesheng; Jasa, John; Li, Dongli; Cleveland, Robin O; Negahban, Mehrdad; Jérusalem, Antoine

    2017-08-16

    The chemobiomechanical signatures of diseased cells are often distinctively different from that of healthy cells. This mainly arises from cellular structural/compositional alterations induced by disease development or therapeutic molecules. Therapeutic shock waves have the potential to mechanically destroy diseased cells and/or increase cell membrane permeability for drug delivery. However, the biomolecular mechanisms by which shock waves interact with diseased and healthy cellular components remain largely unknown. By integrating atomistic simulations with a novel multiscale numerical framework, this work provides new biomolecular mechanistic perspectives through which many mechanosensitive cellular processes could be quantitatively characterised. Here we examine the biomechanical responses of the chosen representative membrane complexes under rapid mechanical loadings pertinent to therapeutic shock wave conditions. We find that their rupture characteristics do not exhibit significant sensitivity to the applied strain rates. Furthermore, we show that the embedded rigid inclusions markedly facilitate stretch-induced membrane disruptions while mechanically stiffening the associated complexes under the applied membrane stretches. Our results suggest that the presence of rigid molecules in cellular membranes could serve as "mechanical catalysts" to promote the mechanical destructions of the associated complexes, which, in concert with other biochemical/medical considerations, should provide beneficial information for future biomechanical-mediated therapeutics.

  3. Development of composite membrane materials for fuel cells

    NASA Astrophysics Data System (ADS)

    Lebedeva, O. V.; Chesnokova, A. N.; Pozhidaev, Yu N.; Maksimenko, S. D.; Malakhova, E. A.; Raskulova, T. V.

    2018-03-01

    This study is devoted to the development and investigation of composite membrane materials for fuel cells. Proton conductive membranes consisting of silica and various low- and high-molecular organic compounds have been prepared by the sol-gel method. The synthesized membranes are characterized by proton conductivity (up to 10-2 S/cm), ion-exchange capacity (1.84-3.5 meq/g), thermal stability (260 – 400 °C), and tensile modulus (128 - 322 MPa).

  4. Plasma membrane--cortical cytoskeleton interactions: a cell biology approach with biophysical considerations.

    PubMed

    Kapus, András; Janmey, Paul

    2013-07-01

    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.

  5. Binding and Fusion of Extracellular Vesicles to the Plasma Membrane of Their Cell Targets.

    PubMed

    Prada, Ilaria; Meldolesi, Jacopo

    2016-08-09

    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.

  6. Quantifying the abnormal hemodynamics of sickle cell anemia

    NASA Astrophysics Data System (ADS)

    Lei, Huan; Karniadakis, George

    2012-02-01

    Sickle red blood cells (SS-RBC) exhibit heterogeneous morphologies and abnormal hemodynamics in deoxygenated states. A multi-scale model for SS-RBC is developed based on the Dissipative Particle Dynamics (DPD) method. Different cell morphologies (sickle, granular, elongated shapes) typically observed in deoxygenated states are constructed and quantified by the Asphericity and Elliptical shape factors. The hemodynamics of SS-RBC suspensions is studied in both shear and pipe flow systems. The flow resistance obtained from both systems exhibits a larger value than the healthy blood flow due to the abnormal cell properties. Moreover, SS-RBCs exhibit abnormal adhesive interactions with both the vessel endothelium cells and the leukocytes. The effect of the abnormal adhesive interactions on the hemodynamics of sickle blood is investigated using the current model. It is found that both the SS-RBC - endothelium and the SS-RBC - leukocytes interactions, can potentially trigger the vicious ``sickling and entrapment'' cycles, resulting in vaso-occlusion phenomena widely observed in micro-circulation experiments.

  7. The hydroxyflavone, fisetin, suppresses mast cell activation induced by interaction with activated T cell membranes

    PubMed Central

    Nagai, K; Takahashi, Y; Mikami, I; Fukusima, T; Oike, H; Kobori, M

    2009-01-01

    Background and purpose: Cell-to-cell interactions between mast cells and activated T cells are increasingly recognized as a possible mechanism in the aetiology of allergic or non-allergic inflammatory disorders. To determine the anti-allergic effect of fisetin, we examined the ability of fisetin to suppress activation of the human mast cell line, HMC-1, induced by activated Jurkat T cell membranes. Experimental approach: HMC-1 cells were incubated with or without fisetin for 15 min and then co-cultured with Jurkat T cell membranes activated by phorbol-12-myristate 13-acetate for 16 h. We determined gene expression in activated HMC-1 cells by DNA microarray and quantitative reverse transcription (RT)-PCR analysis. We also examined activation of the transcription factor NF-κB and MAP kinases (MAPKs) in activated HMC-1 cells. Key results: Fisetin suppresses cell spreading and gene expression in HMC-1 cells stimulated by activated T cell membranes. Additionally, we show that these stimulated HMC-1 cells expressed granzyme B. The stimulatory interaction also induced activation of NF-κB and MAPKs; these activations were suppressed by fisetin. Fisetin also reduced the amount of cell surface antigen CD40 and intercellular adhesion molecule-1 (ICAM-1) on activated HMC-1 cells. Conclusions and implications: Fisetin suppressed activation of HMC-1 cells by activated T cell membranes by interfering with cell-to-cell interaction and inhibiting the activity of NF-κB and MAPKs and thereby suppressing gene expression. Fisetin may protect against the progression of inflammatory diseases by limiting interactions between mast cells and activated T cells. PMID:19702784

  8. Measuring the diffusion coefficient of ganglioside on cell membrane by fluorescence correlation spectroscopy

    NASA Astrophysics Data System (ADS)

    Dong, Shiqing; You, Minghai; Chen, Jianling; Zhou, Jie; Xie, Shusen; Yang, Hongqin

    2017-06-01

    The fluidity of proteins and lipids on cell membrane plays an important role in cell’s physiological functions. Fluorescence correlation spectroscopy (FCS) is an effective technique to detect the rapid dynamic behaviors of proteins and/or lipids in living cells. In this study, we used the rhodamine6G solution to optimize the FCS system. And, cholera toxin B subunit (CT-B) was used to label ganglioside on living Hela cell membranes. The diffusion time and coefficients of ganglioside can be obtained through fitting the autocorrelation curve based on the model of two-dimensional cell membrane. The results showed that the diffusion coefficients of ganglioside distributed within a wide range. It revealed the lateral diffusion of lipids on cell membrane was inhomogeneous, which was due to different microstructures of cytoplasmic membrane. The study provides a helpful method for further studying the dynamic characteristics of proteins and lipids molecules on living cell membrane.

  9. Alkaline fuel cell with nitride membrane

    NASA Astrophysics Data System (ADS)

    Sun, Shen-Huei; Pilaski, Moritz; Wartmann, Jens; Letzkus, Florian; Funke, Benedikt; Dura, Georg; Heinzel, Angelika

    2017-06-01

    The aim of this work is to fabricate patterned nitride membranes with Si-MEMS-technology as a platform to build up new membrane-electrode-assemblies (MEA) for alkaline fuel cell applications. Two 6-inch wafer processes based on chemical vapor deposition (CVD) were developed for the fabrication of separated nitride membranes with a nitride thickness up to 1 μm. The mechanical stability of the perforated nitride membrane has been adjusted in both processes either by embedding of subsequent ion implantation step or by optimizing the deposition process parameters. A nearly 100% yield of separated membranes of each deposition process was achieved with layer thickness from 150 nm to 1 μm and micro-channel pattern width of 1μm at a pitch of 3 μm. The process for membrane coating with electrolyte materials could be verified to build up MEA. Uniform membrane coating with channel filling was achieved after the optimization of speed controlled dip-coating method and the selection of dimethylsulfoxide (DMSO) as electrolyte solvent. Finally, silver as conductive material was defined for printing a conductive layer onto the MEA by Ink-Technology. With the established IR-thermography setup, characterizations of MEAs in terms of catalytic conversion were performed successfully. The results of this work show promise for build up a platform on wafer-level for high throughput experiments.

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

    DOE PAGES

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

    2016-01-21

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

  11. A cell-free translocation system using extracts of cultured insect cells to yield functional membrane proteins.

    PubMed

    Ezure, Toru; Nanatani, Kei; Sato, Yoko; Suzuki, Satomi; Aizawa, Keishi; Souma, Satoshi; Ito, Masaaki; Hohsaka, Takahiro; von Heijine, Gunnar; Utsumi, Toshihiko; Abe, Keietsu; Ando, Eiji; Uozumi, Nobuyuki

    2014-01-01

    Cell-free protein synthesis is a powerful method to explore the structure and function of membrane proteins and to analyze the targeting and translocation of proteins across the ER membrane. Developing a cell-free system based on cultured cells for the synthesis of membrane proteins could provide a highly reproducible alternative to the use of tissues from living animals. We isolated Sf21 microsomes from cultured insect cells by a simplified isolation procedure and evaluated the performance of the translocation system in combination with a cell-free translation system originating from the same source. The isolated microsomes contained the basic translocation machinery for polytopic membrane proteins including SRP-dependent targeting components, translocation channel (translocon)-dependent translocation, and the apparatus for signal peptide cleavage and N-linked glycosylation. A transporter protein synthesized with the cell-free system could be functionally reconstituted into a lipid bilayer. In addition, single and double labeling with non-natural amino acids could be achieved at both the lumen side and the cytosolic side in this system. Moreover, tail-anchored proteins, which are post-translationally integrated by the guided entry of tail-anchored proteins (GET) machinery, were inserted correctly into the microsomes. These results showed that the newly developed cell-free translocation system derived from cultured insect cells is a practical tool for the biogenesis of properly folded polytopic membrane proteins as well as tail-anchored proteins.

  12. Repair of Nerve Cell Membrane Damage by Calcium-Dependent, Membrane-Binding Proteins (Revised)

    DTIC Science & Technology

    2012-09-01

    protein, in model systems can promote a stable repair of broken membranes that could preserve cell viability. Preliminary data obtained using a novel...multilamellar liposomes prepared from bovine brain Folch Fraction I lipids (Sigma-Aldrich) and ion exchange chromatography on Poros Q medium using a Pharmacia...FPLC system [5]. Strategy for establishing the membrane leakage model The strategy employed in these studies was to encapsulate

  13. Additive effect of red blood cell rigidity and adherence to endothelial cells in inducing vascular resistance.

    PubMed

    Kaul, D K; Koshkaryev, A; Artmann, G; Barshtein, G; Yedgar, S

    2008-10-01

    To explore the contribution of red blood cell (RBC) deformability and interaction with endothelial cells (ECs) to circulatory disorders, these RBC properties were modified by treatment with hydrogen peroxide (H(2)O(2)), and their effects on vascular resistance were monitored following their infusion into rat mesocecum vasculature. Treatment with 0.5 mM H(2)O(2) increased RBC/EC adherence without significant alteration of RBC deformability. At 5.0 mM H(2)O(2), RBC deformability was considerably reduced, inducing a threefold increase in the number of undeformable cells, whereas RBC/EC adherence was not further affected by the increased H(2)O(2) concentration. This enabled the selective manipulation of RBC adherence and deformability and the testing of their differential effect on vascular resistance. Perfusion of RBCs with enhanced adherence and unchanged deformability (treatment with 0.5 mM H(2)O(2)) increased vascular resistance by about 35% compared with untreated control RBCs. Perfusion of 5.0 mM H(2)O(2)-treated RBCs, with reduced deformability (without additional increase of adherence), further increased vascular resistance by about 60% compared with untreated control RBCs. These results demonstrate the specific effects of elevated adherence and reduced deformability of oxidized RBCs on vascular resistance. These effects can be additive, depending on the oxidation conditions. The oxidation-induced changes applied in this study are moderate compared with those observed in RBCs in pathological states. Yet, they caused a considerable increase in vascular resistance, thus demonstrating the potency of RBC/EC adherence and RBC deformability in determining resistance to blood flow in vivo.

  14. Infectious rotavirus enters cells by direct cell membrane penetration, not by endocytosis

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kaljot, K.T.; Shaw, R.D.; Greenberg, H.B.

    1988-04-01

    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 ofmore » 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.« less

  15. Externally disposed plasma membrane proteins. I. Enzymatic iodination of mouse L cells

    PubMed Central

    1975-01-01

    The enzymatic iodination technique has been utilized in a study of the externally disposed membrane proteins of the mouse L cell. Iodination of cells in suspension results in lactoperoxidase-specific iodide incorporation with no loss of cell viability under the conditions employed, less than 3% lipid labeling, and more than 90% of the labeled species identifiable as monoiodotyrosine. 90% of the incorporated label is localized to the cell surface by electron microscope autoradiography, with 5-10% in the centrosphere region and postulated to represent pinocytic vesicles. Sodium dodecylsulfate-polyacrylamide gels of solubilized L-cell proteins reveals five to six labeled peaks ranging from 50,000 to 200,000 daltons. Increased resolution by use of gradient slab gels reveals 15-20 radioactive bands. Over 60% of the label resides in approximately nine polypeptides of 80,000 to 150,000 daltons. Various controls indicate that the labeling pattern reflects endogenous membrane proteins, not serum components. The incorporated 125-I, cholesterol, and one plasma membrane enzyme marker, alkaline phosphodiesterase I, are purified in parallel when plasma membranes are isolated from intact, iodinated L cells. The labeled components present in a plasma membrane-rich fraction from iodinated cells are identical to those of the total cell, with a 10- to 20-fold enrichment in specific activity of each radioactive peak in the membrane. PMID:163833

  16. Effects of the consumption of fish meals on the carotid IntimaMedia thickness in patients with hypertension: a prospective study.

    PubMed

    Colussi, GianLuca; Catena, Cristiana; Dialti, Valeria; Mos, Lucio; Sechi, Leonardo A

    2014-01-01

    The composition of dietary fat affects various modifiable cardiovascular risk factors and cardiovascular outcomes in the general population. We investigated the effects of the regular consumption of fish meals on the fatty acid composition of red blood cell (RBC) membranes and the relationship of this parameter with the carotid intima-media thickness (IMT), an early marker of atherosclerosis. In 56 hypertensive patients, we measured the carotid IMT using ultrasound imaging and the RBC membrane fatty acid composition using gas-chromatography and calculated the polyunsaturated to saturated fatty acid (PUFA/SFA) ratio. The patients received intensive nutritional counseling and three weekly meals of fish containing elevated amounts of PUFA, in order to increase the membrane PUFA content. The RBC membrane fatty acid composition and IMT were reassessed after one year. At baseline, the membrane PUFA/SFA ratio was inversely related to the carotid IMT, and the relationship was independent of all major cardiovascular risk factors. At follow-up, the PUFA/SFA ratio increased in the RBC membranes of 25 (45%) of 56 patients. The regular consumption of fish meals resulted in a decreased carotid IMT only in the patients with an increased membrane PUFA/SFA ratio. Changes in the PUFA/SFA ratio induced by the dietary intervention were inversely related to the changes in the IMT, independent of variations in body mass, blood pressure and plasma lipids. In hypertensive patients, a low RBC membrane PUFA/SFA ratio is associated with more prominent vascular damage, and the regular consumption of fish reduces the carotid IMT in patients in whom dietary intervention affects the membrane fatty acid composition.

  17. Membrane fusion-competent virus-like proteoliposomes and proteinaceous supported bilayers made directly from cell plasma membranes.

    PubMed

    Costello, Deirdre A; Hsia, Chih-Yun; Millet, Jean K; Porri, Teresa; Daniel, Susan

    2013-05-28

    Virus-like particles are useful materials for studying virus-host interactions in a safe manner. However, the standard production of pseudovirus based on the vesicular stomatitis virus (VSV) backbone is an intricate procedure that requires trained laboratory personnel. In this work, a new strategy for creating virus-like proteoliposomes (VLPLs) and virus-like supported bilayers (VLSBs) is presented. This strategy uses a cell blebbing technique to induce the formation of nanoscale vesicles from the plasma membrane of BHK cells expressing the hemagglutinin (HA) fusion protein of influenza X-31. These vesicles and supported bilayers contain HA and are used to carry out single particle membrane fusion events, monitored using total internal reflection fluorescence microscopy. The results of these studies show that the VLPLs and VLSBs contain HA proteins that are fully competent to carry out membrane fusion, including the formation of a fusion pore and the release of fluorophores loaded into vesicles. This new strategy for creating spherical and planar geometry virus-like membranes has many potential applications. VLPLs could be used to study fusion proteins of virulent viruses in a safe manner, or they could be used as therapeutic delivery particles to transport beneficial proteins coexpressed in the cells to a target cell. VLSBs could facilitate high throughput screening of antiviral drugs or pathogen-host cell interactions.

  18. Genome-wide association study of red blood cell traits in Hispanics/Latinos: The Hispanic Community Health Study/Study of Latinos

    PubMed Central

    Morrison, Jean V.; Brown, Lisa; Schurmann, Claudia; Chen, Diane D.; Liu, Yong Mei; Auer, Paul L.; Taylor, Kent D.; Papanicolaou, George; Kurita, Ryo; Nakamura, Yukio; Loos, Ruth J. F.; North, Kari E.; Thornton, Timothy A.; Pankratz, Nathan; Bauer, Daniel E.

    2017-01-01

    Prior GWAS have identified loci associated with red blood cell (RBC) traits in populations of European, African, and Asian ancestry. These studies have not included individuals with an Amerindian ancestral background, such as Hispanics/Latinos, nor evaluated the full spectrum of genomic variation beyond single nucleotide variants. Using a custom genotyping array enriched for Amerindian ancestral content and 1000 Genomes imputation, we performed GWAS in 12,502 participants of Hispanic Community Health Study and Study of Latinos (HCHS/SOL) for hematocrit, hemoglobin, RBC count, RBC distribution width (RDW), and RBC indices. Approximately 60% of previously reported RBC trait loci generalized to HCHS/SOL Hispanics/Latinos, including African ancestral alpha- and beta-globin gene variants. In addition to the known 3.8kb alpha-globin copy number variant, we identified an Amerindian ancestral association in an alpha-globin regulatory region on chromosome 16p13.3 for mean corpuscular volume and mean corpuscular hemoglobin. We also discovered and replicated three genome-wide significant variants in previously unreported loci for RDW (SLC12A2 rs17764730, PSMB5 rs941718), and hematocrit (PROX1 rs3754140). Among the proxy variants at the SLC12A2 locus we identified rs3812049, located in a bi-directional promoter between SLC12A2 (which encodes a red cell membrane ion-transport protein) and an upstream anti-sense long-noncoding RNA, LINC01184, as the likely causal variant. We further demonstrate that disruption of the regulatory element harboring rs3812049 affects transcription of SLC12A2 and LINC01184 in human erythroid progenitor cells. Together, these results reinforce the importance of genetic study of diverse ancestral populations, in particular Hispanics/Latinos. PMID:28453575

  19. Cationic nanoparticles induce nanoscale disruption in living cell plasma membranes.

    PubMed

    Chen, Jiumei; Hessler, Jessica A; Putchakayala, Krishna; Panama, Brian K; Khan, Damian P; Hong, Seungpyo; Mullen, Douglas G; Dimaggio, Stassi C; Som, Abhigyan; Tew, Gregory N; Lopatin, Anatoli N; Baker, James R; Holl, Mark M Banaszak; Orr, Bradford G

    2009-08-13

    It has long been recognized that cationic nanoparticles induce cell membrane permeability. Recently, it has been found that cationic nanoparticles induce the formation and/or growth of nanoscale holes in supported lipid bilayers. In this paper, we show that noncytotoxic concentrations of cationic nanoparticles induce 30-2000 pA currents in 293A (human embryonic kidney) and KB (human epidermoid carcinoma) cells, consistent with a nanoscale defect such as a single hole or group of holes in the cell membrane ranging from 1 to 350 nm(2) in total area. Other forms of nanoscale defects, including the nanoparticle porating agents adsorbing onto or intercalating into the lipid bilayer, are also consistent; although the size of the defect must increase to account for any reduction in ion conduction, as compared to a water channel. An individual defect forming event takes 1-100 ms, while membrane resealing may occur over tens of seconds. Patch-clamp data provide direct evidence for the formation of nanoscale defects in living cell membranes. The cationic polymer data are compared and contrasted with patch-clamp data obtained for an amphiphilic phenylene ethynylene antimicrobial oligomer (AMO-3), a small molecule that is proposed to make well-defined 3.4 nm holes in lipid bilayers. Here, we observe data that are consistent with AMO-3 making approximately 3 nm holes in living cell membranes.

  20. Membrane tension: A challenging but universal physical parameter in cell biology.

    PubMed

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

    2017-11-01

    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.

  1. Block copolymers for alkaline fuel cell membrane materials

    NASA Astrophysics Data System (ADS)

    Li, Yifan

    Alkaline fuel cells (AFCs) using anion exchange membranes (AEMs) as electrolyte have recently received considerable attention. AFCs offer some advantages over proton exchange membrane fuel cells, including the potential of non-noble metal (e.g. nickel, silver) catalyst on the cathode, which can dramatically lower the fuel cell cost. The main drawback of traditional AFCs is the use of liquid electrolyte (e.g. aqueous potassium hydroxide), which can result in the formation of carbonate precipitates by reaction with carbon dioxide. AEMs with tethered cations can overcome the precipitates formed in traditional AFCs. Our current research focuses on developing different polymer systems (blend, block, grafted, and crosslinked polymers) in order to understand alkaline fuel cell membrane in many aspects and design optimized anion exchange membranes with better alkaline stability, mechanical integrity and ionic conductivity. A number of distinct materials have been produced and characterized. A polymer blend system comprised of poly(vinylbenzyl chloride)-b-polystyrene (PVBC-b-PS) diblock copolymer, prepared by nitroxide mediated polymerization (NMP), with poly(2,6-dimethyl-1,4-phenylene oxide) (PPO) or brominated PPO was studied for conversion into a blend membrane for AEM. The formation of a miscible blend matrix improved mechanical properties while maintaining high ionic conductivity through formation of phase separated ionic domains. Using anionic polymerization, a polyethylene based block copolymer was designed where the polyethylene-based block copolymer formed bicontinuous morphological structures to enhance the hydroxide conductivity (up to 94 mS/cm at 80 °C) while excellent mechanical properties (strain up to 205%) of the polyethylene block copolymer membrane was observed. A polymer system was designed and characterized with monomethoxy polyethylene glycol (mPEG) as a hydrophilic polymer grafted through substitution of pendent benzyl chloride groups of a PVBC

  2. Effect of RBC concentrate transfusions on serum ferritin content in children with acute leukaemia.

    PubMed

    Bebeshko, V G; Bruslova, E M; Tsvietkova, N M; Iatsemirskii, S M; Puchkareva, T I; Gonchar, L A; Krukovska, V V; Zelinska, A V; Mishchenko, L P

    2013-01-01

    To study the serum ferritin levels in children with acute leukemia, depending on the number of transfusions of RBC concentrate and period of disease. We studied the red blood count, serum iron and ferritin levels in 54 patients with acute leukemia before chemotherapy, at the time of a standardized treatment protocol, and after transfusions of RBC concentrates. In the debute of acute leukemia just before treatment lauch the serum ferritin in 81.5% of children was 2.3-2.5 higher than normal. The need for transfusion of RBC concentrates was higher under serum ferritin level exceeding 500 ng/mL. The association was established between ferritin content and age of the children, variant of acute leukemia and period of the disease. The level of serum ferritin can be used as a marker of ferrokinetic status for timely diagnosis of iron overload in children with acute leukemias and for application of treatment-and-prophylactic actions. Bebeshko V. G., Bruslova K. M., Cvjetkova N. M., Jacemyrskyj S. M., Pushkarova T. I., Gonchar L. O., Krukovska V. V., Zelinska A. V., Mishhenko L. P., 2013.

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

    PubMed

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

    2015-10-14

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

  4. Red blood cell-derived microparticles: An overview.

    PubMed

    Westerman, Maxwell; Porter, John B

    2016-07-01

    The red blood cell (RBC) is historically the original parent cell of microparticles (MPs). In this overview, we describe the discovery and the early history of red cell-derived microparticles (RMPs) and present an overview of the evolution of RMP. We report the formation, characteristics, effects of RMP and factors which may affect RMP evaluation. The review examines RMP derived from both normal and pathologic RBC. The pathologic RBC studies include sickle cell anemia (SCA), sickle cell trait (STr), thalassemia intermedia (TI), hereditary spherocytosis (HS), hereditary elliptocytosis (HE), hereditary stomatocytosis (HSt) and glucose-6-phosphate dehydrogenase deficiency (G6PD). Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Single-Molecule Imaging of Wnt3A Protein Diffusion on Living Cell Membranes.

    PubMed

    Lippert, Anna; Janeczek, Agnieszka A; Fürstenberg, Alexandre; Ponjavic, Aleks; Moerner, W E; Nusse, Roel; Helms, Jill A; Evans, Nicholas D; Lee, Steven F

    2017-12-19

    Wnt proteins are secreted, hydrophobic, lipidated proteins found in all animals that play essential roles in development and disease. Lipid modification is thought to facilitate the interaction of the protein with its receptor, Frizzled, but may also regulate the transport of Wnt protein and its localization at the cell membrane. Here, by employing single-molecule fluorescence techniques, we show that Wnt proteins associate with and diffuse on the plasma membranes of living cells in the absence of any receptor binding. We find that labeled Wnt3A transiently and dynamically associates with the membranes of Drosophila Schneider 2 cells, diffuses with Brownian kinetics on flattened membranes and on cellular protrusions, and does not transfer between cells in close contact. In S2 receptor-plus (S2R+) cells, which express Frizzled receptors, membrane diffusion rate is reduced and membrane residency time is increased. These results provide direct evidence of Wnt3A interaction with living cell membranes, and represent, to our knowledge, a new system for investigating the dynamics of Wnt transport. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  6. Do mechanical forces contribute to nanoscale membrane organisation in T cells?

    PubMed

    Klotzsch, Enrico; Stiegler, Johannes; Ben-Ishay, Eldad; Gaus, Katharina

    2015-04-01

    Mechanotransduction describes how a cell senses and interacts with its environment. The concept originated in adhesion biology where adhesion receptors, integrins, facilitate force transmission between the extracellular matrix and the intracellular actin cytoskeleton. Indeed, during any adhesive contacts, cells do exert mechanical force. Hence, the probing of the local environment by cells results in mechanical cues that contribute to cellular functions and cell fate decisions such as migration, proliferation, differentiation and apoptosis. On the molecular level, mechanical forces can rearrange proteins laterally within the membrane, regulate their activity by inducing conformational changes and probe the mechanical properties and bond strength of receptor-ligands. From this point of view, it appears surprising that molecular forces have been largely overlooked in membrane organisation and ligand discrimination processes in lymphocytes. During T cell activation, the T cell receptor recognises and distinguishes antigenic from benign endogenous peptides to initiate the reorganisation of membrane proteins into signalling clusters within the immunological synapse. In this review, we asked whether characteristics of fibroblast force sensing could be applied to immune cell antigen recognition and signalling, and outline state-of-the-art experimental strategies for studying forces in the context of membrane organisation. This article is part of a Special Issue entitled: Nanoscale membrane orgainisation and signalling. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. Hydrogen ion dynamics in human red blood cells

    PubMed Central

    Swietach, Pawel; Tiffert, Teresa; Mauritz, Jakob M A; Seear, Rachel; Esposito, Alessandro; Kaminski, Clemens F; Lew, Virgilio L; Vaughan-Jones, Richard D

    2010-01-01

    Our understanding of pH regulation within red blood cells (RBCs) has been inferred mainly from indirect experiments rather than from in situ measurements of intracellular pH (pHi). The present work shows that carboxy-SNARF-1, a pH fluorophore, when used with confocal imaging or flow cytometry, reliably reports pHi in individual, human RBCs, provided intracellular fluorescence is calibrated using a ‘null-point’ procedure. Mean pHi was 7.25 in CO2/HCO3−-buffered medium and 7.15 in Hepes-buffered medium, and varied linearly with extracellular pH (slope of 0.77). Intrinsic (non-CO2/HCO3−-dependent) buffering power, estimated in the intact cell (85 mmol (l cell)−1 (pH unit)−1 at resting pHi), was somewhat higher than previous estimates from cell lysates (50–70 mmol (l cell)−1 (pH unit)−1). Acute displacement of pHi (superfusion of weak acids/bases) triggered rapid pHi recovery. This was mediated via membrane Cl−/HCO3− exchange (the AE1 gene product), irrespective of whether recovery was from an intracellular acid or base load, and with no evident contribution from other transporters such as Na+/H+ exchange. H+-equivalent flux through AE1 was a linear function of [H+]i and reversed at resting pHi, indicating that its activity is not allosterically regulated by pHi, in contrast to other AE isoforms. By simultaneously monitoring pHi and markers of cell volume, a functional link between membrane ion transport, volume and pHi was demonstrated. RBC pHi is therefore tightly regulated via AE1 activity, but modulated during changes of cell volume. A comparable volume–pHi link may also be important in other cell types expressing anion exchangers. Direct measurement of pHi should be useful in future investigations of RBC physiology and pathology. PMID:20962000

  8. The effect of abnormal hemoglobins on the membrane regulation of cell hydration.

    PubMed

    Clark, M R; Shohet, S B

    Several hemoglobinopathies are associated with abnormalities in the permeability of the red cell membrane, in some cases leading to permanent alterations of the intracellular milieu. Homozygous sickle cell disease is the most thoroughly studied example. Deoxygenation of sickle cells causes a transient increase in the permeability to monovalent cations and Ca; prolonged deoxygenation can lead to a permanent accumulation of Ca and loss of total cations and water. Although the mechanisms for the permeability changes are not yet defined, mechanical stress on the membrane, with subsequent damages by excess Ca or membrane-associated hemoglobin have been suggested to play a role. Loss of cell water and increase in mean cell hemoglobin concentration causes massive reduction of cell deformability in the oxygenated state and makes the hemoglobin more likely to undergo sickling because of the strong concentration dependence of the sickling process. Limited evidence suggests the occurrence of permeability defects in other hemoglobinopathies and the thalassemias. The suggested alterations range from a slight increase in K permeability of incubated thalassemia cells to substantial dehydration of cells from patients with homozygous hemoglobin C disease. Oxidative damage to the membrane, involving an abnormal hemoglobin-membrane association, may underly the permeability changes in these cells.

  9. [The Role of Membrane-Bound Heat Shock Proteins Hsp90 in Migration of Tumor Cells in vitro and Involvement of Cell Surface Heparan Sulfate Proteoglycans in Protein Binding to Plasma Membrane].

    PubMed

    Snigireva, A V; Vrublevskaya, V V; Skarga, Y Y; Morenkov, O S

    2016-01-01

    Heat shock protein Hsp90, detected in the extracellular space and on the membrane of cells, plays an important role in cell motility, migration, invasion and metastasis of tumor cells. At present, the functional role and molecular mechanisms of Hsp90 binding to plasma membrane are not elucidated. Using isoform-specific antibodies against Hsp90, Hsp9α and Hsp90β, we showed that membrane-bound Hsp90α and Hsp90β play a significant role in migration of human fibrosarcoma (HT1080) and glioblastoma (A-172) cells in vitro. Disorders of sulfonation of cell heparan sulfates, cleavage of cell heparan. sulfates by heparinase I/III as well as treatment of cells with heparin lead to an abrupt reduction in the expression level of Hsp90 isoforms. Furthermore, heparin significantly inhibits tumor cell migration. The results obtained demonstrate that two isoforms of membrane-bound Hsp90 are involved in migration of tumor cells in vitro and that cell surface heparan sulfate proteoglycans play a pivotal role in the "anchoring" of Hsp90α and Hsp90β to the plasma membrane.

  10. Advances in the high performance polymer electrolyte membranes for fuel cells.

    PubMed

    Zhang, Hongwei; Shen, Pei Kang

    2012-03-21

    This critical review tersely and concisely reviews the recent development of the polymer electrolyte membranes and the relationship between their properties and affecting factors like operation temperature. In the first section, the advantages and shortcomings of the corresponding polymer electrolyte membrane fuel cells are analyzed. Then, the limitations of Nafion membranes and their alternatives to large-scale commercial applications are discussed. Secondly, the concepts and approaches of the alternative proton exchange membranes for low temperature and high temperature fuel cells are described. The highlights of the current scientific achievements are given for various aspects of approaches. Thirdly, the progress of anion exchange membranes is presented. Finally, the perspectives of future trends on polymer electrolyte membranes for different applications are commented on (400 references).

  11. Phosphatidylinositol 3-phosphates-at the interface between cell signalling and membrane traffic.

    PubMed

    Marat, Andrea L; Haucke, Volker

    2016-03-15

    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.

  12. Proton-conductive nanochannel membrane for fuel-cell applications.

    PubMed

    Oleksandrov, Sergiy; Lee, Jeong-Woo; Jang, Joo-Hee; Haam, Seungjoo; Chung, Chan-Hwa

    2009-02-01

    Novel design of proton conductive membrane for direct methanol fuel cells is based on proton conductivity of nanochannels, which is acquired due to the electric double layer overlap. Proton conductivity and methanol permeability of an array of nanochannels were studied. Anodic aluminum oxide with pore diameter of 20 nm was used as nanochannel matrix. Channel surfaces of an AAO template were functionalized with sulfonic groups to increase proton conductivity of nanochannels. This was done in two steps; at first -SH groups were attached to walls of nanochannels using (3-Mercaptopropyl)-trimethyloxysilane and then they were converted to -SO3H groups using hydrogen peroxide. Treatment steps were analyzed by Fourier Transform Infrared spectroscopy and X-ray Photoelectron Spectroscopy. Proton conductivity and methanol permeability were measured. The data show methanol permeability of membrane to be an order of magnitude lower, than that measured of Nafion. Ion conductivity of functionalized AAO membrane was measured by an impedance analyzer at frequencies ranging from 1 Hz to 100 kHz and voltage 50 mV to be 0.15 Scm(-1). Measured ion conductivity of Nafion membrane was 0.05 Scm(-1). Obtained data show better results in comparison with commonly used commercial available proton conductive membrane Nafion, thus making nanochannel membrane very promising for use in fuel cell applications.

  13. Polymer-mediated immunocamouflage of red blood cells: effects of polymer size on antigenic and immunogenic recognition of allogeneic donor blood cells.

    PubMed

    Wang, DunCheng; Kyluik, Dana L; Murad, Kari L; Toyofuku, Wendy M; Scott, Mark D

    2011-07-01

    Developing a practical means of reducing alloimmunization in chronically transfused patients would be of significant clinical benefit. Immunocamouflaging red blood cells (RBCs) by membrane grafting of methoxypoly(ethylene glycol) (mPEG) may reduce the risk of allo-immunization. The results of this study showed that antibody recognition of non-ABO antigens was significantly reduced in an mPEG-dose- and polymer size-dependent manner, with higher molecular weight mPEGs providing better immunoprotection. Furthermore, in vivo immunogenicity was significantly reduced in mice serially transfused with mPEG-modified xenogeneic (sheep; sRBCs), allogeneic (C57Bl/6), or syngeneic (Balb/c) RBCs. Following a primary transfusion of sRBCs, mice receiving mPEG-sRBCs showed a >90% reduction in anti-sRBC IgG antibody levels. After two transfusions, mice receiving mPEG-sRBCs showed reductions of >80% in anti-sRBC IgG levels. Importantly, mPEG-modified autologous cells did not induce neoantigens or an immune (IgG or IgM) response. These data suggest that the global immunocamouflage of RBCs by polymer grafting may provide a safe and cost-effective means of reducing the risk of alloimmunization.

  14. Cell-autonomous defense, re-organization and trafficking of membranes in plant-microbe interactions.

    PubMed

    Dörmann, Peter; Kim, Hyeran; Ott, Thomas; Schulze-Lefert, Paul; Trujillo, Marco; Wewer, Vera; Hückelhoven, Ralph

    2014-12-01

    Plant cells dynamically change their architecture and molecular composition following encounters with beneficial or parasitic microbes, a process referred to as host cell reprogramming. Cell-autonomous defense reactions are typically polarized to the plant cell periphery underneath microbial contact sites, including de novo cell wall biosynthesis. Alternatively, host cell reprogramming converges in the biogenesis of membrane-enveloped compartments for accommodation of beneficial bacteria or invasive infection structures of filamentous microbes. Recent advances have revealed that, in response to microbial encounters, plasma membrane symmetry is broken, membrane tethering and SNARE complexes are recruited, lipid composition changes and plasma membrane-to-cytoskeleton signaling is activated, either for pre-invasive defense or for microbial entry. We provide a critical appraisal on recent studies with a focus on how plant cells re-structure membranes and the associated cytoskeleton in interactions with microbial pathogens, nitrogen-fixing rhizobia and mycorrhiza fungi. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

  15. [Observation of the L929 cell membrane after infrasound exposure with atomic force microscope].

    PubMed

    Wang, Bing-shui; Chen, Jing-zao; Liu, Bin; Li, Ling; Yi, Nan; Liu, Jing; Zhang, Sa

    2005-12-01

    To observe the changes of L929 cell membrane with atomic force microscope (AFM) after infrasound exposure and to explore the mechanisms of effect of infrasound on cell membrane. After primary culture, the L929 cells were exposed to infrasound with intensity output of 130 dB and frequency of 16 Hz 2 hours each day for 3 days. The subsequent changes in the membrane of the control cells and the cells exposed to the infrasound were determined by nano-scale scanning with AFM. After infrasound exposure, the normal prominence of the membrane became short and the dent became shallow in the 7.5 microm x 7.5 microm and 4.0 microm x 4.0 microm photographs. The prominence appeared as cobblestones. The surface of the membrane became smooth. The membrane structure of the L929 cells can be changed by infrasound exposure with intensity of 130 dB and frequency of 16 Hz. The change might be one of the characteristics of effect of infrasound on cell membrane.

  16. Alkaline polymer electrolyte membranes for fuel cell applications.

    PubMed

    Wang, Yan-Jie; Qiao, Jinli; Baker, Ryan; Zhang, Jiujun

    2013-07-07

    In this review, we examine the most recent progress and research trends in the area of alkaline polymer electrolyte membrane (PEM) development in terms of material selection, synthesis, characterization, and theoretical approach, as well as their fabrication into alkaline PEM-based membrane electrode assemblies (MEAs) and the corresponding performance/durability in alkaline polymer electrolyte membrane fuel cells (PEMFCs). Respective advantages and challenges are also reviewed. To overcome challenges hindering alkaline PEM technology advancement and commercialization, several research directions are then proposed.

  17. Hemoglobin-driven pathophysiology is an in vivo consequence of the red blood cell storage lesion that can be attenuated in guinea pigs by haptoglobin therapy.

    PubMed

    Baek, Jin Hyen; D'Agnillo, Felice; Vallelian, Florence; Pereira, Claudia P; Williams, Matthew C; Jia, Yiping; Schaer, Dominik J; Buehler, Paul W

    2012-04-01

    Massive transfusion of blood can lead to clinical complications, including multiorgan dysfunction and even death. Such severe clinical outcomes have been associated with longer red blood cell (rbc) storage times. Collectively referred to as the rbc storage lesion, rbc storage results in multiple biochemical changes that impact intracellular processes as well as membrane and cytoskeletal properties, resulting in cellular injury in vitro. However, how the rbc storage lesion triggers pathophysiology in vivo remains poorly defined. In this study, we developed a guinea pig transfusion model with blood stored under standard blood banking conditions for 2 (new), 21 (intermediate), or 28 days (old blood). Transfusion with old but not new blood led to intravascular hemolysis, acute hypertension, vascular injury, and kidney dysfunction associated with pathophysiology driven by hemoglobin (Hb). These adverse effects were dramatically attenuated when the high-affinity Hb scavenger haptoglobin (Hp) was administered at the time of transfusion with old blood. Pathologies observed after transfusion with old blood, together with the favorable response to Hp supplementation, allowed us to define the in vivo consequences of the rbc storage lesion as storage-related posttransfusion hemolysis producing Hb-driven pathophysiology. Hb sequestration by Hp might therefore be a therapeutic modality for enhancing transfusion safety in severely ill or massively transfused patients.

  18. Membrane Composition Tunes the Outer Hair Cell Motor

    NASA Astrophysics Data System (ADS)

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

    2009-02-01

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

  19. Chimera proteins with affinity for membranes and microtubule tips polarize in the membrane of fission yeast cells.

    PubMed

    Recouvreux, Pierre; Sokolowski, Thomas R; Grammoustianou, Aristea; ten Wolde, Pieter Rein; Dogterom, Marileen

    2016-02-16

    Cell polarity refers to a functional spatial organization of proteins that is crucial for the control of essential cellular processes such as growth and division. To establish polarity, cells rely on elaborate regulation networks that control the distribution of proteins at the cell membrane. In fission yeast cells, a microtubule-dependent network has been identified that polarizes the distribution of signaling proteins that restricts growth to cell ends and targets the cytokinetic machinery to the middle of the cell. Although many molecular components have been shown to play a role in this network, it remains unknown which molecular functionalities are minimally required to establish a polarized protein distribution in this system. Here we show that a membrane-binding protein fragment, which distributes homogeneously in wild-type fission yeast cells, can be made to concentrate at cell ends by attaching it to a cytoplasmic microtubule end-binding protein. This concentration results in a polarized pattern of chimera proteins with a spatial extension that is very reminiscent of natural polarity patterns in fission yeast. However, chimera levels fluctuate in response to microtubule dynamics, and disruption of microtubules leads to disappearance of the pattern. Numerical simulations confirm that the combined functionality of membrane anchoring and microtubule tip affinity is in principle sufficient to create polarized patterns. Our chimera protein may thus represent a simple molecular functionality that is able to polarize the membrane, onto which additional layers of molecular complexity may be built to provide the temporal robustness that is typical of natural polarity patterns.

  20. C8-glycosphingolipids preferentially insert into tumor cell membranes and promote chemotherapeutic drug uptake.

    PubMed

    Cordeiro Pedrosa, Lília R; van Cappellen, Wiggert A; Steurer, Barbara; Ciceri, Dalila; ten Hagen, Timo L M; Eggermont, Alexander M M; Verheij, Marcel; Goñi, Felix María; Koning, Gerben A; Contreras, F-Xabier

    2015-08-01

    Insufficient drug delivery into tumor cells limits the therapeutic efficacy of chemotherapy. Co-delivery of liposome-encapsulated drug and synthetic short-chain glycosphingolipids (SC-GSLs) significantly improved drug bioavailability by enhancing intracellular drug uptake. Investigating the mechanisms underlying this SC-GSL-mediated drug uptake enhancement is the aim of this study. Fluorescence microscopy was used to visualize the cell membrane lipid transfer intracellular fate of fluorescently labeled C6-NBD-GalCer incorporated in liposomes in tumor and non-tumor cells. Additionally click chemistry was applied to image and quantify native SC-GSLs in tumor and non-tumor cell membranes. SC-GSL-mediated flip-flop was investigated in model membranes to confirm membrane-incorporation of SC-GSL and its effect on membrane remodeling. SC-GSL enriched liposomes containing doxorubicin (Dox) were incubated at 4°C and 37°C and intracellular drug uptake was studied in comparison to standard liposomes and free Dox. SC-GSL transfer to the cell membrane was independent of liposomal uptake and the majority of the transferred lipid remained in the plasma membrane. The transfer of SC-GSL was tumor cell-specific and induced membrane rearrangement as evidenced by a transbilayer flip-flop of pyrene-SM. However, pore formation was measured, as leakage of hydrophilic fluorescent probes was not observed. Moreover, drug uptake appeared to be mediated by SC-GSLs. SC-GSLs enhanced the interaction of doxorubicin (Dox) with the outer leaflet of the plasma membrane of tumor cells at 4°C. Our results demonstrate that SC-GSLs preferentially insert into tumor cell plasma membranes enhancing cell intrinsic capacity to translocate amphiphilic drugs such as Dox across the membrane via a biophysical process. Copyright © 2015 Elsevier B.V. All rights reserved.