Changes In Fluidity Research Articles

Laurdan in live cell imaging: Effect of acquisition settings, cell culture conditions and data analysis on generalized polarization measurements.

Cell function is highly dependent on membrane structure, organization, and fluidity. Therefore, methods to probe the biophysical properties of biological membranes are required. Determination of generalized polarization (GP) values using Laurdan in fluorescence microscopy studies is one of the most widely-used methods to investigate changes in membrane fluidity in vitro and in vivo. In the last couple of decades, there has been a major increase in the number of studies using Laurdan GP, where several different methodological approaches are used. Such differences interfere with data interpretation inasmuch as it is difficult to validate if Laurdan GP variations actually reflect changes in membrane organization or arise from biased experimental approaches. To address this, we evaluated the influence of different methodological details of experimental data acquisition and analysis on Laurdan GP. Our results showed that absolute GP values are highly dependent on several of the parameters analyzed, showing that incorrect data can result from technical and methodological inconsistencies. Considering these differences, we further analyzed the impact of cell variability on GP determination, focusing on basic cell culture conditions, such as cell confluency, number of passages and media composition. Our results show that GP values can report alterations in the biophysical properties of cell membranes caused by cellular adaptation to the culture conditions. In summary, this study provides thorough analysis of the factors that can lead to Laurdan GP variability and suggests approaches to improve data quality, which would generate more precise interpretation and comparison within individual studies and among the literature on Laurdan GP.

Lipid headgroup and side chain architecture determine manganese-induced dose dependent membrane rigidification and liposome size increase.

Metal ion-membrane interactions have gained appreciable attention over the years resulting in increasing investigations into the mode of action of toxic and essential metals. More work has focused on essential ions like Ca or Mg and toxic metals like Cd and Pb, whereas this study investigates the effects of the abundant essential trace metal manganese with model lipid systems by screening zwitterionic and anionic glycerophospholipids. Despite its essentiality, deleterious impact towards cell survival is known under Mn stress. The fluorescent dyes Laurdan and diphenylhexatriene were used to assess changes in membrane fluidity both in the head group and hydrophobic core region of the membrane, respectively. Mn-rigidified membranes composed of the anionic phospholipids, phosphatidic acid, phosphatidylglycerol, cardiolipin, and phosphatidylserine. Strong binding resulted in large shifts of the phase transition temperature. The increase was in the order phosphatidylserine > phosphatidylglycerol > cardiolipin, and in all cases, saturated analogues > mono-unsaturated forms. Dynamic light scattering measurements revealed that Mn caused extensive aggregation of liposomes composed of saturated analogues of phosphatidic acid and phosphatidylserine, whilst the mono-unsaturated analogue had significant membrane swelling. Increased membrane rigidity may interfere with permeability of ions and small molecules, possibly disrupting cellular homeostasis. Moreover, liposome size changes could indicate fusion, which could also be detrimental to cellular transport. Overall, this study provided further understanding into the effects of Mn with biomembranes, whereby the altered membrane properties are consequential to the proper structural and signalling functions of membrane lipids.

Strikingly Different Roles of SARS-CoV-2 Fusion Peptides Uncovered by Neutron Scattering.

Coronavirus disease-2019 (COVID-19), a potentially lethal respiratory illness caused by the coronavirus SARS-CoV-2, emerged in the end of 2019 and has since spread aggressively across the globe. A thorough understanding of the molecular mechanisms of cellular infection by coronaviruses is therefore of utmost importance. A critical stage in infection is the fusion between viral and host membranes. Here, we present a detailed investigation of the role of selected SARS-CoV-2 Spike fusion peptides, and the influence of calcium and cholesterol, in this fusion process. Structural information from specular neutron reflectometry and small angle neutron scattering, complemented by dynamics information from quasi-elastic and spin–echo neutron spectroscopy, revealed strikingly different functions encoded in the Spike fusion domain. Calcium drives the N-terminal of the Spike fusion domain to fully cross the host plasma membrane. Removing calcium, however, reorients the peptide back to the lipid leaflet closest to the virus, leading to significant changes in lipid fluidity and rigidity. In conjunction with other regions of the fusion domain, which are also positioned to bridge and dehydrate viral and host membranes, the molecular events leading to cell entry by SARS-CoV-2 are proposed.

Fluidity is the way to life: lipid phase separation in bacterial membranes.

A hallmark of biological membranes is the dynamic localization of lipids and proteins. Lipids respond to temperature reduction below a critical point with phase separation, and poikilothermic animals and also bacteria adapt their lipid content to prevent gel phase formation in membranes. In a new study, Gohrbandt etal (2022) show that reduced membrane fluidity in bacterial cells causes reversible phase separation without membrane rupture invivo, highlighting the physical robustness of biological membranes.

Changes in fluidity of the E.coli outer membrane in response to temperature, divalent cations and polymyxin-B show two different mechanisms of membrane fluidity adaptation.

The outer membrane (OM) is an essential component of the Gram-negative bacterial cell envelope. Restricted diffusion of integral OM proteins and lipopolysaccharide (LPS) that constitute the outer leaflet of the OM support a model in which the OM is in a semi-crystalline state. The low fluidity of the OM has been suggested to be an important property of this membrane that even contributes to cell rigidity. The LPS characteristics strongly determine the properties of the OM and the LPS layer fluidity has been measured using different techniques that require specific conditions or are technically challenging. Here, we characterize the Escherichia coli LPS fluidity by evaluating the lateral diffusion of the styryl dye FM4-64FX in fluorescence recovery after photobleaching experiments. This technique allowed us to determine the effect of different conditions and genetic backgrounds on the LPS fluidity. Our results show that a fraction of the LPS can slowly diffuse and that the fluidity of the LPS layer adapts by modifying the diffusion of the LPS and the fraction of mobile LPS molecules.

Investigating the effects of alcohols and cholesterol manipulations on bovine aortic endothelial cell membrane properties using phasor analysis of laurdan fluorescence lifetime microscopy and spectral imaging

Endothelial cell membranes are made up of a heterogenous mixture of lipids and proteins and include liquid ordered (Lo) domains called lipid rafts and caveolae. These compartments have higher concentrations of cholesterol and sphingolipids and are concentrated in signaling molecules. Perturbations in membrane fluidity, including changes in cholesterol content can lead to changes in the structure of the plasma membrane and impair signaling mechanisms. In order to study how changing membrane fluidity can lead to changes in cellular signaling, we must be able to characterize changes in fluidity.

Bio-Inspired Proanthocyanidins from Blueberries’ Surface Coating Prevents Red Blood Cell Agglutination on Urinary Silicon-Based Catheters

Thrombosis can cause the occlusion of implantable medical devices, leading to the rejection of the device and subsequent mortality. Thrombosis is primarily induced by red blood aggregation and coagulation. The administration of anticoagulant drugs is generally used as a treatment to avoid these processes. Adverse effects such as bleeding in the event of an anticoagulant overdose, osteoporosis associated with prolonged use, hypersensitivity, and hives have been reported. New strategies such as biomolecule surface functionalization have recently been studied to overcome these problems. In this study, we report a novel coating composed of polydopamine (PDA) and proanthocyanidins (PACs) from blueberry extract to avoid red blood aggregation in short-term use medical devices such as silicone catheters. We showed that PDA formed stable films on silicone surfaces and PACs could be immobilized on PDA layers using laccase as a catalyst. The PDA–PACs films decreased surface hydrophilicity, increased surface roughness, and decreased plasma protein adsorption. The films were stable in phosphate buffer saline (PBS) and cell culture media. Furthermore, red blood cell adsorption and aggregation decreased. These effects are attributed to changes in the membrane fluidity that influences adhesion, the steric hindrance of the layers, and the low adsorption of plasma proteins on the PAC layer.

Particle uptake driven phagocytosis in macrophages and neutrophils enhances bacterial clearance

Humans are exposed to numerous synthetic foreign particles in the form of drug delivery systems and diagnostic agents. Specialized immune cells (phagocytes) clear these particles by phagocytosing and attempting to degrade them. The process of recognition and internalization of the particles may trigger changes in the function of phagocytes. Some of these changes, especially the ability of a particle-loaded phagocyte to take up and neutralize pathogens, remains poorly studied. Herein, we demonstrate that the uptake of non-stimulatory cargo-free particles enhances the phagocytic ability of monocytes, macrophages and neutrophils. The enhancement in phagocytic ability was independent of particle properties, such as size or the base material constituting the particle. Additionally, we show that the increased phagocytosis was not a result of cellular activation or cellular heterogeneity but was driven by changes in cell membrane fluidity and cellular compliance. A consequence of the enhanced phagocytic activity was that particulate-laden immune cells neutralize Escherichia coli (E. coli) faster in culture. Moreover, when administered in mice as a prophylactic, particulates enable faster clearance of E. coli and Staphylococcus epidermidis. Together, we demonstrate that the process of uptake induces cellular changes that favor additional phagocytic events. This study provides insights into using non-stimulatory cargo-free particles to engineer immune cell functions for applications involving faster clearance of phagocytosable abiotic and biotic material.

Effects of polyethylene microplastics on cell membranes: A combined study of experiments and molecular dynamics simulations

Microplastics (MPs), widely distributed within the environment, can be ingested by humans easily and cause various biological reactions such as oxidative stress, immune response and membrane damage, ultimately representing a threat to health. Cell membranes work as first barrier for MPs entering the cell and playing biological effects. For now, the researches on interactions of MPs on cell membranes lack an in-depth and effective theoretical model to understand molecular details and physicochemical behaviors. In present study, observations of calcein leakage established polyethylene plastic nanoparticles (PE PNPs), especially of high concentrations, harming cell membrane integrity. SYTOX green and lactate dehydrogenase (LDH) assays supported the evidence that the exposure of cells to PE PNPs caused significant cell membrane damage in dose-response. Molecular dynamics (MD) simulations were further applied to determine the effects of PE on the properties of dipalmitoyl phosphatidylcholine (DPPC) bilayer. PE permeated into lipid membranes easily resulting in significant variations in DPPC bilayer with lower density, fluidity changes and membrane thickening. Besides, PE aggregates bound were more likely to cause pore formation and serious damage to the DPPC bilayer. The interaction mechanisms between MPS and cell membrane were explored which provided valuable insights into membrane effect of MPs.

Microstructure and rheological behavior of capillary suspension prepared with plate-shaped particles

Capillary suspension is a unique strategy to prepare high-yield-stress fluids, in which particle-to-particle interaction caused by capillary bridges results in a substantial change in fluidity. The capillary suspension with plate-shaped particles have recently been considered useful in various applications, including Li-battery electrodes and conductive elastomers, due to their large specific surface area, which results in a rigid sample-spanning capillary network. However, there is a poor understanding of the relationship between the dimensions of plate-shaped particles and the fluidity of the capillary suspensions.In this study, the yield stress of the capillary suspension prepared with plate-shaped particles of various diameters and thicknesses was investigated to establish a correlation equation. Additionally, the conformation of the particles in the capillary suspension was also determined by microscopic analysis. As a result, it was discovered that when the secondary fluid is added, plate-shaped particles form a laminated structure supported by capillary bridges generated between their largest faces (pendular state). Furthermore, rheological measurements revealed that the yield stress of the capillary suspension was determined by the number density of particles, cross-sectional area of the particle, and volume fraction of the secondary fluid. Based on these findings, we established a power-law-based correlation equation that accurately predicted (20% error at most) the yield stress of the capillary suspension regardless of the size of the particles.

Metabolomics reveals that CAF-derived lipids promote colorectal cancer peritoneal metastasis by enhancing membrane fluidity.

Patients with peritoneal metastasis (PM) of colorectal cancer (CRC) have poorer overall survival outcomes than those without PM. Cancer-associated fibroblasts (CAFs) are a major component of the tumor microenvironment and mediate CRC progression and PM. It is imperative to identify and develop novel therapeutic targets for PM-CRC driven by CAFs. Using lipidomics, we reveal that the abundance of phosphatidylcholine (PC) with unsaturated acyl chains was increased in clinical PM-CRC specimens. Additionally, we found that CAFs were present at a higher relative abundance in primary PM-CRC tumors and that membrane fluidity in CRC cells was increased after incubation with CAF-conditioned medium (CM) through three independent methods: lipidomics, fluorescence recovery after photobleaching (FRAP), and generalized polarization. Then, we found that increased membrane fluidity can enhance glucose uptake and metabolism, as supported by real-time bioenergetics analysis and U-13C glucose labeling. Interestingly, stearoyl-CoA desaturase 1 (SCD), the rate-limiting enzyme in the biosynthesis of unsaturated fatty acids (uS-FAs), was expressed at low levels in PM and associated with poor prognosis in CRC patients. Importantly, by untargeted metabolomics analysis and fatty acid ([U-13C]-stearic acid) tracing analyses, we found that CRC cells take up lipids and lipid-like metabolites secreted from CAFs, which may compensate for low SCD expression. Both in vitro and in vivo experiments demonstrated that sodium palmitate (C16:0) treatment could decrease the CAF-induced change in cell membrane fluidity, limit glucose metabolism, suppress cell invasiveness, and impair tumor growth and intraperitoneal dissemination. An increased C16:0 concentration was shown to induce apoptosis linked to lipotoxicity. Furthermore, C16:0 effectively enhanced the antitumor activity of 5-fluorouracil (5-FU) in vitro and was well tolerated in vivo. Taken together, these findings suggest that adding the saturated fatty acid (S-FA) C16:0 to neoadjuvant chemotherapy may open new opportunities for treating PM-CRC in the future.

Influence of Nanomodified Waterproofing Agent on the Performance of Rigid Waterproof Concrete in Underground Engineering

Waterproof and impermeable problems seriously affect the safe use, normal structure, and working life of concrete projects. The waterproof performance of concrete can be improved by mixing with a proper amount of waterproofing agent; nevertheless, the early strength of concrete will be seriously affected. With the development of complex structures such as larg e spans in soil and water engineering and the increasingly complex engineering environment, people have put forward higher requirements for the working performance, strength, impermeability, durability, and intelligence of cement‐based materials required. This article compares the changes in compressive strength, fluidity, and water permeability of mixed concrete by controlling the two variables of waterproofing agent and nanosilica. Combining the results of XRD microstructure analysis, the influence of nanosilica and waterproofing agent on the performance of concrete is explored. Moreover, it is hoped that other excellent properties of concrete can be improved, and a waterproof material with good opacity can be found. The test results show that after adding waterproofing agent alone, the 3 d compressive strength of concrete is decreased by 14.81%, the water permeability is reduced by 71.6%, and the depth of carbonic acid molecules at 28 d is also decreased by 37.3%. On the other hand, compared with concrete using waterproof coating alone, after adding in nanosilica, the 3 d compressive strength is increased by about 30%, and the water immersion height is decreased. The results show that the addition of polymer has a great influence on the compressive strength and carbonic acid resistance of concrete.

Hemodynamic Forces, Endothelial Mechanotransduction, and Vascular Diseases.

Cells in the tissues and organs of a living body are subjected to mechanical forces, such as pressure, friction, and tension from their surrounding environment. Cells are equipped with a mechanotransduction mechanism by which they perceive mechanical forces and transmit information into the cell interior, thereby causing physiological or pathogenetic mechano-responses. Endothelial cells (ECs) lining the inner surface of blood vessels are constantly exposed to shear stress caused by blood flow and a cyclic strain caused by intravascular pressure. A number of studies have shown that ECs are sensitive to changes in these hemodynamic forces and alter their morphology and function, sometimes by modifying gene expression. The mechanism of endothelial mechanotransduction has been elucidated, and the plasma membrane has recently been shown to act as a mechanosensor. The lipid order and cholesterol content of plasma membranes change immediately upon the exposure of ECs to hemodynamic forces, resulting in a change in membrane fluidity. These changes in a plasma membrane's physical properties affect the conformation and function of various ion channels, receptors, and microdomains (such as caveolae and primary cilia), thereby activating a wide variety of downstream signaling pathways. Such endothelial mechanotransduction works to maintain circulatory homeostasis; however, errors in endothelial mechanotransduction can cause abnormalities in vascular physiological function, leading to the initiation and progression of various vascular diseases, such as hypertension, thrombosis, aneurysms, and atherosclerosis. Recent advances in detailed imaging technology and computational fluid dynamics analysis have enabled us to evaluate the hemodynamic forces acting on vascular tissue accurately, contributing greatly to our understanding of vascular mechanotransduction and the pathogenesis of vascular diseases, as well as the development of new therapies for vascular diseases.

Correction: Hyperthermia and associated changes in membrane fluidity potentiate P2X7 activation to promote tumor cell death.

Correction: Hyperthermia and associated changes in membrane fluidity potentiate P2X7 activation to promote tumor cell death.

Controlled rate slow freezing with lyoprotective agent to retain the integrity of lipid nanovesicles during lyophilization

We designed a novel lyophilization method using controlled rate slow freezing (CSF) with lyoprotective agent (LPA) to achieve intact lipid nanovesicles after lyophilization. During the freezing step, LPA prevented water supercooling, and the freezing rate was controlled by CSF. Regulating the freezing rate by various liquid media was a crucial determinant of membrane disruption, and isopropanol (freezing rate of 0.933 °C/min) was the optimal medium for the CSF system. Lyophilized lipid nanovesicle using both CSF and LPA retained 92.9% of the core material and had uniform size distributions (Z-average diameter = 133.4 nm, polydispersity index = 0.144), similar to intact vesicles (120.7 nm and 0.159, respectively), after rehydration. Only lyophilized lipid nanovesicle using both CSF and LPA showed no changes in membrane fluidity and polarity. This lyophilization method can be applied to improve storage stability of lipid nanocarriers encapsulating drugs while retaining their original activity.

Phase-separated protein droplets of amyotrophic lateral sclerosis-associated p62/SQSTM1 mutants show reduced inner fluidity

Several amyotrophic lateral sclerosis (ALS)-related proteins such as FUS, TDP-43, and hnRNPA1 demonstrate liquid–liquid phase separation, and their disease-related mutations correlate with a transition of their liquid droplet form into aggregates. Missense mutations in SQSTM1/p62, which have been identified throughout the gene, are associated with ALS, frontotemporal degeneration (FTD), and Paget’s disease of bone. SQSTM1/p62 protein forms liquid droplets through interaction with ubiquitinated proteins, and these droplets serve as a platform for autophagosome formation and the antioxidative stress response via the LC3-interacting region (LIR) and KEAP1-interacting region (KIR) of p62, respectively. However, it remains unclear whether ALS/FTD-related p62 mutations in the LIR and KIR disrupt liquid droplet formation leading to defects in autophagy, the stress response, or both. To evaluate the effects of ALS/FTD-related p62 mutations in the LIR and KIR on a major oxidative stress system, the Keap1-Nrf2 pathway, as well as on autophagic turnover, we developed systems to monitor each of these with high sensitivity. These methods such as intracellular protein–protein interaction assay, doxycycline-inducible gene expression system, and gene expression into primary cultured cells with recombinant adenovirus revealed that some mutants, but not all, caused reduced NRF2 activation and delayed autophagic cargo turnover. In contrast, while all p62 mutants demonstrated sufficient ability to form liquid droplets, all of these droplets also exhibited reduced inner fluidity. These results indicate that like other ALS-related mutant proteins, p62 missense mutations result in a primary defect in ALS/FTD via a qualitative change in p62 liquid droplet fluidity.

Plasma Membrane Fluidity: An Environment Thermal Detector in Plants.

The lipid matrix in cell membranes is a dynamic, bidimensional array of amphipathic molecules exhibiting mesomorphism, which contributes to the membrane fluidity changes in response to temperature fluctuation. As sessile organisms, plants must rapidly and accurately respond to environmental thermal variations. However, mechanisms underlying temperature perception in plants are poorly understood. We studied the thermal plasticity of membrane fluidity using three fluorescent probes across a temperature range of −5 to 41 °C in isolated microsomal fraction (MF), vacuolar membrane (VM), and plasma membrane (PM) vesicles from Arabidopsis plants. Results showed that PM were highly fluid and exhibited more phase transitions and hysteresis, while VM and MF lacked such attributes. These findings suggest that PM is an important cell hub with the capacity to rapidly undergo fluidity modifications in response to small changes of temperatures in ranges spanning those experienced in natural habitats. PM fluidity behaves as an ideal temperature detector: it is always present, covers the whole cell, responds quickly and with sensitivity to temperature variations, functions with a cell free-energy cost, and it is physically connected with potential thermal signal transducers to elicit a cell response. It is an optimal alternative for temperature detection selected for the plant kingdom.

Electron Spin Resonance Evaluation of Buccal Membrane Fluidity Alterations by Sodium Caprylate and L-Menthol.

The ability of sodium caprylate and l-menthol to fluidize phospholipid bilayers composed of lipids simulating the buccal epithelium was investigated using electron spin resonance (ESR) to evaluate the action of these agents as permeation enhancers. 5-Doxyl stearic acid (5-DSA) and 16-doxyl stearic acid (16-DSA) were used as spin labels to identify alterations in membrane fluidity near the polar head groups or inner acyl regions of the lipid bilayer, respectively. The molecular motion of both 5-DSA and 16-DSA showed increased disorder near the polar and inner hydrophobic regions of the bilayer in the presence of sodium caprylate suggesting fluidization in both the regions, which contributes to its permeation enhancing effects. L-menthol decreased the order parameter for 16-DSA, showing membrane fluidization only in the inner acyl regions of the bilayer, which also corresponded to its weaker permeation enhancing effects. The rapid evaluation of changes in fluidity of the bilayer in the presence of potential permeation enhancers using ESR enables improved selection of effective permeation enhancers and enhancer combinations based on their effect on membrane fluidization.

Using the negative hydration of inorganic salt ions to improve the properties of Alkali-activated slag cement in plastic stage

Alkali-activated slag (AAS) cement is a low-carbon environmentally friendly cementitious material. Water glass (WG) activated slag cement has excellent physical and mechanical properties, but it is not conducive to the construction due to its rapid setting and hardening, easy loss of fluidity. Negative hydration ions can reduce the viscosity of aqueous solution. So, in the present paper, inorganic salt composed of negative hydration ions were used to adjust the fluidity and setting time of AAS paste. The results show that with the increase of NaCl, KCl, KBr and KNO3, the initial fluidity of AAS paste increases slightly, and the 30 min fluidity increases a lot, while the fluidity at 60 min is greatly affected by the amount of inorganic salt. KNO3 shows the greatest initial fluidity increase. When 0.2% Ba(NO3)2 is added with other four inorganic salts, the fluidity retention is significantly improved, especially the fluidity of 60 min. When 5% NaCl, KCl, KBr and KNO3 respectively mixed with 0.2% Ba(NO3)2 are added, the fluidity retention of AAS paste at 60 min are 97.4%, 92.7%, 100%, 82.5% respectively. Compared with the AAS paste without inorganic salt, when 5% NaCl, KCl, KBr and KNO3 respectively blended with 0.2% Ba(NO3)2 are added, the initial setting time increase by 228.6%, 285.7% 307.1% and 239.3%, while the final setting time increase by 175.6%, 222.0%, 231.7% and 178.0% respectively. With the inorganic salt, the ζ-potential of the originally negative AAS cement suspension become more negative, both yield stress and plastic viscosity coefficient decease, especially the plastic viscosity coefficient greatly reduces. This may be the reason for the change of fluidity and setting time of AAS paste.

PsbS-зависимые изменения в содержании липидов и фотосинтетических пигментов в растениях риса

Plants acclimate to fluctuations in light conditions by adjusting their photosynthetic apparatus. When the light intensity exceeds, an unbalanced excitation of the two photosystems occurs. It results in reduced photosynthetic efficiency. Photosystem II (PSII) is the most susceptible and dynamically regulated part of the light reactions in the thylakoid membrane. Non-photochemical quenching of chlorophyll fluorescence (NPQ) is one of the short-term photoprotective mechanisms, which consist of the number of components. The strongest NPQ component — qE is localized in the PSII antenna and induced in plants by lumen acidification, the activation of the pH sensor PsbS, and the conversion of the violaxanthin to zeaxanthin within the xanthophyll cycle. Here, I present data that characterizes the role of the PsbS protein in organization of PSII structural components in isolated PSII-enriched membranes. The preparations were isolated from wild-type (WT) and PsbS-less (PsbS-KO) mutant rice plant. Based on the obtained results, the PSII-enriched membranes from WT and PsbS-KO differ as in the level of lipids, also in carotenoids. I conclude that the PsbS-dependent changes in membrane fluidity in PsbS-KO mutant plants compensated with increased lipid level in mutant plants.