Membrane Coalescence Research Articles

Superhydrophilic ZSM-5 zeolite-coated membrane for enhancing water coalescence in water-in-oil emulsions

The filtration efficiency of coalescent filter elements used in gasoline and diesel oil dehydration in refineries is unsatisfactory and not sufficient to meet downstream water-content requirements. Thus, it was necessary to improve the internal filter medium of traditional oil-water coalescent filter elements. Zeolite membrane is a kind of material with superhydrophilicity and strong adsorption for and permeability to water. In this study, ZSM-5 zeolite membrane with superhydrophilic properties was successfully prepared by an in situ hydrothermal synthesis. Two experiments with zeolite membranes were performed, including membrane filtration and membrane coalescence tests. Membrane filtration tests showed that it possessed strong adsorption for and permeability to water and that the oil-water filtration efficiency for 32# turbine oil, kerosene, and corn oil were all >95 %. In the second part of this study, composite filtration was carried out using the membrane as well as hydrophilic glass fiber and hydrophobic polypropylene (PP) membranes. Membrane coalescence test demonstrated that, when the oil contained a phenol/water combination, the water rejection rate of the composite filtration with ABACBA (A, PP fiber; B, glass fiber; and C, molecular sieve membrane) was 90.8 %, with the downstream water content controlled within 100 mg/L and the quality factor increased from 0.18 to 0.23, relative to other systems. These experimental results showed potential for improving the current oil-water coalescent filters.

Protein kinase CK2 subunits exert specific and coordinated functions in skeletal muscle differentiation and fusogenic activity.

Casein kinase 2 (CK2) is a tetrameric protein kinase composed of 2 catalytic (α and α') and 2 regulatory β subunits. Our study provides the first molecular and cellular characterization of the different CK2 subunits, highlighting their individual roles in skeletal muscle specification and differentiation. Analysis of C2C12 cell knockout for each CK2 subunit reveals that: 1) CK2β is mandatory for the expression of the muscle master regulator myogenic differentiation 1 in proliferating myoblasts, thus controlling both myogenic commitment and subsequent muscle-specific gene expression and myotube formation; 2) CK2α is involved in the activation of the muscle-specific gene program; and 3) CK2α' activity regulates myoblast fusion by mediating plasma membrane translocation of fusogenic proteins essential for membrane coalescence, like myomixer. Accordingly, CK2α' overexpression in C2C12 cells and in mouse regenerating muscle is sufficient to increase myofiber size and myonuclei content via enhanced satellite cell fusion. Consistent with these results, pharmacological inhibition of CK2 activity substantially blocks the expression of myogenic markers and muscle cell fusion both in vitro in C2C12 and primary myoblasts and in vivo in mouse regenerating muscle and zebrafish development. Overall, our work describes the specific and coordinated functions of CK2 subunits in orchestrating muscle differentiation and fusogenic activity, highlighting CK2 relevance in the physiopathology of skeletal muscle tissue.-Salizzato, V., Zanin, S., Borgo, C., Lidron, E., Salvi, M., Rizzuto, R., Pallafacchina, G., Donella-Deana, A. Protein kinase CK2 subunits exert specific and coordinated functions in skeletal muscle differentiation and fusogenic activity.

Myomaker and Myomerger Work Independently to Control Distinct Steps of Membrane Remodeling during Myoblast Fusion

Classic mechanisms for membrane fusion involve transmembrane proteins that assemble into complexes and dynamically alter their conformation to bend membranes, leading to mixing of membrane lipids (hemifusion) and fusion pore formation. Myomaker and Myomerger govern myoblast fusion and muscle formation but are structurally divergent from traditional fusogenic proteins. Here, we show that Myomaker and Myomerger independently mediate distinct steps in the fusion pathway, where Myomaker is involved in membrane hemifusion and Myomerger is necessary for fusion pore formation. Mechanistically, we demonstrate that Myomerger is required on the cell surface where its ectodomains stress membranes. Moreover, we show that Myomerger drives fusion completion in a heterologous system independentof Myomaker and that a Myomaker-Myomergerphysical interaction is not required for function. Collectively, our data identify a stepwise cell fusion mechanism in myoblasts where different proteins are delegated to perform unique membrane functions essential for membrane coalescence.

Lattice-Boltzmann flow simulation of an oil-in-water emulsion through a coalescing filter: Effects of filter structure

The permeation of an oil-in-water (O/W) emulsion through a coalescing filter was numerically studied using the lattice Boltzmann method (LBM). A numerical simulation model for the coalescing phenomena was developed based on the free-energy LBM. We investigated the effects of the wettability of fibers, filter porosity, and fiber diameter on the coalescing behaviors by performing two-dimensional permeation simulations for the O/W emulsions through modeled fibrous filters. We mainly focused on hydrophilic filters because they did not generate small secondary droplets during oil droplet detachment from the filter, and this is preferred for precise separation of oil and water. Our simulations demonstrated that filters with larger pore spacings enable formation of larger droplets but allow more droplets to pass without coalescing. To solve this problem, we designed bi-layered filters composed of a small-pore filter to accurately catch the droplets and a large-pore filter to enlarge the droplets; we demonstrated the effectiveness of the bilayer structure for membrane coalescence.

Numerical simulation of coalescence phenomena of oil-in-water emulsions permeating through straight membrane pore

Abstract We numerically investigated the demulsification phenomena of oil-in-water (O/W) emulsions using a membrane coalescence process. We simulated the permeation of O/W emulsions through a membrane with a single straight pore using our numerical framework based on the coupled level set and volume-of-fluid (CLSVOF) method. With a low droplet volume fraction, small droplets, and a large pore size, the oil droplets frequently passed through the membrane without attaching to its surface. After the steady coalescence of the droplets on the membrane surface at the permeation side, large oil droplets could be obtained. With a higher droplet volume fraction, larger oil droplets, and smaller membrane pores, the oil droplets easily blocked the membrane pores and were thus forced away by the permeating flow, which yielded relatively small oil droplets. Our numerical work revealed the fundamental dynamics of the coalescence of oil droplets in an O/W emulsion using a simple membrane model.

Treatment of model oily waste water by microfiltration

Microfiltration membranes made from polytetrafluorethylene (PTFE) have been successfully used for the separation of oily waste waters by membrane coalescence, but filtration of the oily waste waters with high emulsifier content have not yet been investigated. Our aim was to examine the effect of emulsifier content on membrane coalescence on this type of membrane, and to investigate the effect of surfactant addition on oil retention and filtration parameters. It was found that the oil content of the emulsion can be separated with MF PTFE membranes. Efficiency of the retention depends on the concentration of emulsion: increasing concentrations resulted in an increased retention. The addition of coagulant also affected filtration parameters: higher coagulant concentrations resulted in a significant increase of the flux, while the retention marginally decreased.

Endocytic Recycling Proteins EHD1 and EHD2 Interact with Fer-1-like-5 (Fer1L5) and Mediate Myoblast Fusion

The mammalian ferlins are calcium-sensing, C2 domain-containing proteins involved in vesicle trafficking. Myoferlin and dysferlin regulate myoblast fusion and muscle membrane resealing, respectively. Correspondingly, myoferlin is most highly expressed in singly nucleated myoblasts, whereas dysferlin expression is increased in mature, multinucleated myotubes. Myoferlin also mediates endocytic recycling and participates in trafficking the insulin-like growth factor receptor. We have now characterized a novel member of the ferlin family, Fer1L5, because of its high homology to dysferlin and myoferlin. We found that Fer1L5 protein is expressed in small myotubes that contain only two to four nuclei. We also found that Fer1L5 protein binds directly to the endocytic recycling proteins EHD1 and EHD2 and that the second C2 domain in Fer1L5 mediates this interaction. Reduction of EHD1 and/or EHD2 inhibits myoblast fusion, and EHD2 is required for normal translocation of Fer1L5 to the plasma membrane. The characterization of Fer1L5 and its interaction with EHD1 and EHD2 underscores the complex requirement of ferlin proteins and mediators of endocytic recycling for membrane trafficking events during myotube formation.

Estimating the risk of coalescence in membrane emulsification

In order to optimise the membrane emulsification process, there is a need for a model that can be used to keep the production time of en emulsion to a minimum, without impairing its quality. This was the aim of our work. For that, we have studied the influence of transmembrane pressure on the fraction of active pores and on the drop diameter and the drop diameter distribution. We have found that an increase in transmembrane pressure makes it possible to increase the dispersed phase flux by a rise in the number of active pores (from 1 to 13% for the 0.2 μm membrane and from 2 to 45 and 49% for the 0.5 and 0.8 μm membranes, respectively). Furthermore, the increase in transmembrane pressure leads to an increase in the diameter of the droplets produced. For the 0.5 and 0.8 μm pore diameter, we have even found an increase in the polydispersity of the emulsion produced. A thorough study allowed us to locate the origin of this polydispersity: we ruled out post-production coalescence both in the bulk and in the laminar sub-layer. We showed the possibility of the expansion of the three-phase contact line over more than one pore, phenomenon we referred to as membrane coalescence. Based on these results, we built a model which allows to evaluate the risk of membrane coalescence under experimental conditions of dispersed phase flux and transmembrane pressure: the number of active pores at a distance d d/2 from the centre of a given active pore must be <2. For the three pore diameter membranes studied, our model matched the experimental results and could explain the polydispersity observed.

Role of Spike Protein Endodomains in Regulating Coronavirus Entry

Enveloped viruses enter cells by viral glycoprotein-mediated binding to host cells and subsequent fusion of virus and host cell membranes. For the coronaviruses, viral spike (S) proteins execute these cell entry functions. The S proteins are set apart from other viral and cellular membrane fusion proteins by their extensively palmitoylated membrane-associated tails. Palmitate adducts are generally required for protein-mediated fusions, but their precise roles in the process are unclear. To obtain additional insights into the S-mediated membrane fusion process, we focused on these acylated carboxyl-terminal intravirion tails. Substituting alanines for the cysteines that are subject to palmitoylation had effects on both S incorporation into virions and S-mediated membrane fusions. In specifically dissecting the effects of endodomain mutations on the fusion process, we used antiviral heptad repeat peptides that bind only to folding intermediates in the S-mediated fusion process and found that mutants lacking three palmitoylated cysteines remained in transitional folding states nearly 10 times longer than native S proteins. This slower refolding was also reflected in the paucity of postfusion six-helix bundle configurations among the mutant S proteins. Viruses with fewer palmitoylated S protein cysteines entered cells slowly and had reduced specific infectivities. These findings indicate that lipid adducts anchoring S proteins into virus membranes are necessary for the rapid, productive S protein refolding events that culminate in membrane fusions. These studies reveal a previously unappreciated role for covalently attached lipids on the endodomains of viral proteins eliciting membrane fusion reactions.

Phosphocreatine as an energy source for actin cytoskeletal rearrangements during myoblast fusion

Myoblast fusion is essential for muscle development, postnatal growth and muscle repair after injury. Recent studies have demonstrated roles for actin polymerization during myoblast fusion. Dynamic cytoskeletal assemblies directing cell-cell contact, membrane coalescence and ultimately fusion require substantial cellular energy demands. Various energy generating systems exist in cells but the partitioning of energy sources during myoblast fusion is unknown. Here, we demonstrate a novel role for phosphocreatine (PCr) as a spatiotemporal energy buffer during primary mouse myoblast fusion with nascent myotubes. Creatine treatment enhanced cell fusion in a creatine kinase (CK)-dependent manner suggesting that ATP-consuming reactions are replenished through the PCr/CK system. Furthermore, selective inhibition of actin polymerization prevented myonuclear addition following creatine treatment. As myotube formation is dependent on cytoskeletal reorganization, our findings suggest that PCr hydrolysis is coupled to actin dynamics during myoblast fusion. We conclude that myoblast fusion is a high-energy process, and can be enhanced by PCr buffering of energy demands during actin cytoskeletal rearrangements in myoblast fusion. These findings implicate roles for PCr as a high-energy phosphate buffer in the fusion of multiple cell types including sperm/oocyte, trophoblasts and macrophages. Furthermore, our results suggest the observed beneficial effects of oral creatine supplementation in humans may result in part from enhanced myoblast fusion.

Research Trends

Properties of microporous structures prepared from polyethylene/ polymethacrylate IPN Pervaporation membranes based on imide-containing poly(amic acid) and poly(phenylene oxide) Morphology and gas permeation performance of polyimide membranes Factors affecting membrane coalescence of stable oil-in-water emulsions Preparation and characterization of polyacrylonitrile UF membranes Ion-exchange membrane electrodialytic salt production Synthesis and characterization of porous ferrimagnetic membranes Dehydration of water — alcohol mixtures by vapour permeation Recovery of clindamycin from fermentation wastewater using NF Proton-exchange — membrane materials

Factors affecting membrane coalescence of stable oil-in-water emulsions

This paper presents results on membrane coalescence of stable oil-in-water emulsions. The method involves a two-stage process in which the membranes are used to enlarge oil droplets such that a traditional gravity settler can be used for oil phase separation. Several factors that affect the performance of the membrane coalescer have been studied. In particular, the effect of membrane pore size and the imposed in-pore shear rates on coalescence of stable micron and submicron oil droplets present in oil-in-water emulsions were investigated. Hydrophobic membranes made of Teflon (PTFE) with nominal average pore sizes of 0.22, 0.45, 1.2 and 5.0 μm were tested to assess their ability to coalesce oil emulsions with an average d 50 1.5 μm droplet diameter (volume based). Experiments were carried out in a batch dead-end filtration cell and the influence of operating conditions such as transmembrane pressure, membrane orientation, and emulsion concentration were investigated. The coalesced filtrate was allowed to settle for 1 h, and the percentage oil removal achieved for 0.22, 0.45, 1.2 and 5.0 μm was up to 86, 81, 66 and 45%, respectively. The results showed that membrane pore size had a large influence on the coalescence of oil droplets and that membrane orientation is important. Also, imposed shear rates inside the membrane pores play a key role during membrane coalescence, where sufficient in-pore shear rates are required to coalesce oil droplets, although higher shear rates may reverse the process.

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Cells of the innate immune system, notably macrophages, neutrophils and dendritic cells, perform essential antimicrobial and homeostatic functions. These functions rely on the dynamic surveillance of the environment supported by the formation of elaborate membrane protrusions. Such protrusions — pseudopodia, lamellipodia and filopodia — facilitate the sampling of the surrounding fluid by macropinocytosis, as well as the engulfment of particulates by phagocytosis. Both processes entail extreme plasma membrane deformations that require the coordinated rearrangement of cytoskeletal polymers, which exert protrusive force and drive membrane coalescence and scission. The resulting vacuolar compartments undergo pronounced remodeling and ultimate resolution by mechanisms that also involve the cytoskeleton. Here, we describe the regulation and functions of cytoskeletal assembly and remodeling during macropinocytosis and phagocytosis.

The effects of intermittent permeate flow and crossflow on membrane coalescence of oil-in-water emulsions

Membrane coalescence is a chemical-free technique for breaking stable oil-in-water emulsions. In this work the effect of oil droplet residence time within the membrane porous matrix was investigated and a novel process is proposed based on intermittent permeate flow (flow or no flow) at 0.01 Hz. Polytetrafluoroethylene membranes with 0.45 and 1.20 μm nominal average pore size were used to coalesce emulsions with an average 1.5 μm droplet diameter. Experiments were carried out in a crossflow filtration cell and crossflow velocities were varied from 0.07 to 0.28 ms −1. Fluxes were found to increase with crossflow although oil rejection was zero, implying no concentration polarization. Results also demonstrated an improvement in coalescence performance during intermittent operation at low crossflow velocities. Under some conditions intermittent permeate flow operation produced higher permeate fluxes than without intermittent operation and is a potential novel process for membrane coalescence.

Nuclear fusion leads to chromosome doubling during mannitol pretreatment of barley (Hordeum vulgare L.) microspores

A cytological study of barley microspores during pretreatment of the uninucleate stage to the early culture stage was conducted utilizing six genotypes. Among the three main pretreatments investigated, microspores completed the first mitotic division during 28 d cold pretreatment of spikes, with or without leaf sheath attached, and during 0.3 M mannitol pretreatment of anthers at 25 degrees C. However, during a 4 d pretreatment in 0.3 M mannitol at 4 degrees C this first mitotic division was blocked or delayed and subsequently most often occurred during the first day on culture medium. The first mitotic division of most microspores pretreated in 0.3 M mannitol was mostly symmetrical (55-60%), whereas it was asymmetric (94%) during the 28 d cold pretreatment of spikes. Following the first mitotic division during the mannitol pretreatment at 25 degrees C, closely associated daughter nuclei often appeared to fuse via membrane coalescence, leading to a high frequency of large uninucleate microspores. Based upon nuclear size, the frequencies of fused uninucleate microspores in genotypes GBC 778, GBC 777 and Igri were estimated to be 87%, 54% and 75%, respectively, after a 4 d mannitol pretreatment at 25 degrees C. Chromosome numbers in dividing nuclei and relative densitometry measurements of nuclear DNA in microspores from cv. Igri confirmed the apparent fused nature of large nuclei in uninucleate microspores. The high frequency of fused nuclei indicates that nuclear fusion occurred between both symmetric and asymmetric nuclei. Microspores of cv. Igri cultured on filter paper following three different pretreatments provided an average of about 12 000 embryo-like structures (ELS) per plate. In samples, 85-97% of these ELS regenerated green shoots. The frequency of doubled haploids (74-83%) following all pretreatments was similar to the frequencies of fused nuclei. The pretreatment of spikes in 0.3 M mannitol at 4 degrees C for 4 d is preferred as it appears to provide genotype independent induction and suspension of nuclear division, as well as regenerating green plants in a shorter time than cold alone.

Membrane Coalescence for Phase Separation of Oil-in-Water Emulsions Stabilized by Surfactants and Dispersed into Smallest Droplets

In the field of chemical engineering separation processes, such as extraction or distillation of heteroazeotropes, severe problems exist in phase separation of oil/water dispersions containing very small droplets. This is especially true if the bulk phases contain surfactants stabilizing the interface and thus inhibit coalescence. This work was aimed at investigating a novel technique of phase separation due to membrane coalescence for various emulsions stabilized by diverse surfactants in order to clarify the performance as well as the operational limitations.

Electrofusion: A biophysical modification of cell membrane and a mechanism in exocytosis

The molecular bases of the exocytosis process remain poorly known. Many proteins have been recognized to play key roles in the machinery. Their functions are well characterized in the specificity of the docking processes. Forces involved in the merging of the two partners must take into account the physics of membrane interfaces. The target membrane and the vesicle are both electrically charged interfaces. Strong electrostatic fields are triggered when they are brought in close neighborhood. These fields are high enough to induce an electropermeabilisation process. It is now well known that when applied on a cell, an external field induces a modulation of the transmembrane potential difference. When high enough the transmembrane potential may induce a membrane destabilisation. This results in a free exchange of polar molecules across well defined parts of the cell surface. Furthermore, when permeabilization is present on two cells, if those parts of the cell surfaces are brought in close contact, membrane merging occurs spontaneously. Cell fusion results from this membrane coalescence. The similarity with what is taking place in exocytosis is striking. The present review describes the state-of-the-art in the knowledge on electrofusion. It is emphasized that it results from electropermeabilisation and not from a direct effect of the external field. A local destabilisation of the vesicle membrane results from electrostatic interactions while keeping unaffected its viability. Such processes appear relevant for what takes place during exocytosis.

The role of CD4 in HIV binding and entry

The primary cellular receptor for the human and simian immunodeficiency viruses HIV-1, HIV-2 and SIV is the CD4 antigen (Sattentau et al. 1988; Sattentau & Weiss 1988). HIV infection of CD4+ cells is initiated by binding of the virus to the cell surface, via a high-affinity interaction between the first domain of CD4 and the HIV outer envelope glycoprotein, gp120. The use of a soluble recombinant form of CD4 (sCD4) as a receptor mimic has simplified the analysis of receptor binding and post-binding events which result in virus-cell membrane fusion. With cell-line adapted isolates of HIV-1, sCD4 binding induces conformational changes in gp120, leading to the complete dissociation of gp120 from the transmembrane glycoprotein, gp41, and exposing cryptic epitopes of gp41. Similar observations have been made with cell-anchored CD4: recruitment of CD4 molecules leads to exposure of cryptic gp41 epitopes at the fusion interface between clusters of CD4 expressing and HIV-infected cells. It has therefore been proposed that CD4 binding induces exposure of fusogenic components of gp41 which mediate virus-cell membrane coalescence, a process termed receptor-mediated activation of fusion. With the related lentiviruses HIV-2 and SIV, the CD4 induced molecular rearrangements in gp120 are more subtle, implying that there is a spectrum of responses to sCD4 binding.

CD4 activation of HIV fusion.

The primary cellular receptor for the human immunodeficiency viruses type 1 (HIV-1) and type 2 (HIV-2) is the CD4 antigen. HIV infection of CD4+ cells is initiated by binding of the virus to the cell surface, via a high affinity interaction between CD4 and the HIV outer envelope glycoprotein, gp120. The development of model systems using soluble recombinant forms of CD4 (sCD4) has allowed kinetic and thermodynamic analyses of CD4 binding to gp120, and study of the post-binding events leading to virus-cell membrane fusion. It has thus been demonstrated that the affinity of sCD4 for gp120 on virions or HIV-infected cells depends on both the primary sequence and the tertiary structure of gp120 in the membrane. With cell-line adapted isolates of HIV-1, sCD4 binding induces conformational changes in gp120, leading to the complete dissociation of gp120 from the transmembrane glycoprotein, gp41, and exposing cryptic epitopes of gp41. Similar observations have been made with cell-anchored CD4; exposure of cryptic gp41 epitopes occurs at the fusion interface between clusters of CD4-expressing and HIV-infected cells. Thus, for HIV-1, CD4 induces exposure of fusogenic components of gp41 which triggers virus-cell membrane coalescence. This is termed receptor-mediated activation of fusion. With primary isolates of HIV-1 and the related lentiviruses, HIV-2 and simian immunodeficiency virus (SIV), the CD4-induced molecular rearrangements in gp120 are more subtle, implying that there is a spectrum of responses to sCD4 binding. The high-affinity binding site on CD4 for gp120 is necessary and probably sufficient for activation of HIV fusion, although other regions of CD4 may indirectly influence viral entry. There are two regions on the envelope glycoproteins which are recognized as playing a role in HIV entry: the N-terminus of gp41 and the gp120 V3 loop. The roles of these domains are discussed.

Electron microscope study on the regenerative process of peripheral nerves of mice.

An electron microscope study of the reactive process taking place in the proximal segment of sectioned dorso-spinal nerves of white mice was carried out in about 50 animals. Attention was mainly directed to the study of the earliest changes (30′) observable in the neuroplasm but the series of experiments included the study of nerves fixed up to 20 days after section. Most of the facts found (existence of microvesicles, multiplication of mitochondria, and presence of multivesicular bodies) confirmed the observation done in a preceding investigation, plus some non-described new findings. Early after section (30′) the nerve membrane is seen, in some cases, masked by the presence of microvesicles which seem to take origine from the membrane which also shows deep infoldings. These coalesce in many places taking thus the appearance of rows of vesicles. At later stages (60′, 80′) this reactive process is more general in the nerve stumps and vast areas of microvesicles are seen in the fiber neuroplasm. This kind of reaction (membrane infolding and coalescence) decrease later on and is rarely observable at 100′. Microvesicles are nevertheless increasing in number and many of them change their round profile for an elongated shape. Elongation leads to the formation of tubuli of progressively thinner diameter. This process is described as a possible way of neurofilament formation.