Spherocyte shape transformation and release of tubular nanovesicles in human erythrocytes (original) (raw)
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2000
We have studied dodecylmaltoside-induced echinocyte-spheroechincyte-spherocyte shape transformation and membrane vesiculation using transmission electron microscopy (TEM) on freeze-fracture replicas. It is indicated that spherical erythrocyte shape at higher dodecylmaltoside concentration is formed due to loss of membrane in the process where small, mostly tubular nanovesicles are released predominantly from the top of echinocyte and spheroechinocyte spicules. D 2003 Elsevier B.V. All
Microscopy Research and Technique, 2006
For analyses of dynamic ultrastructures of erythrocyte intramembranous particles (IMPs) in situ, a quick-freezing method was used to stabilize the flow behavior of erythrocytes embedded in vitreous ice. Fresh human blood was jetted at various pressures through artificial tubes, in which the flowing erythrocytes were elongated from biconcave discoid shapes to elliptical ones, and quickly frozen in liquid isopentane-propane cryogen (À1938C). They were freeze-fractured using a scalpel in liquid nitrogen, and routinely prepared for replica membranes. Many IMPs were observed on the protoplasmic freeze-fracture face (P-face) of the erythrocyte membranes. Some control erythrocytes under nonflowing or stationary conditions showed IMPs with their random distribution. However, other jetted erythrocytes under flowing conditions showed variously sized IMPs with much closer distribution. They were also arranged into parallel rows in some parts, and aggregated together. This quick-freezing method enabled for the first time the visualization of time-dependent topology and the molecular alteration of IMPs in dynamically flowing erythrocytes. Microsc. Res. Tech. 69: 291-295,
Nature Communications
Extracellular vesicles (EVs) are widely studied regarding their role in cell-to-cell communication and disease, as well as for applications as biomarkers or drug delivery vehicles. EVs contain membrane and intraluminal proteins, affecting their structure and thereby likely their functioning. Here, we use atomic force microscopy for mechanical characterization of erythrocyte, or red blood cell (RBC), EVs from healthy individuals and from patients with hereditary spherocytosis (HS) due to ankyrin deficiency. While these EVs are packed with proteins, their response to indentation resembles that of fluid liposomes lacking proteins. The bending modulus of RBC EVs of healthy donors is~15 k b T, similar to the RBC membrane. Surprisingly, whereas RBCs become more rigid in HS, patient EVs have a significantly (~40%) lower bending modulus than donor EVs. These results shed light on the mechanism and effects of EV budding and might explain the reported increase in vesiculation of RBCs in HS patients.
Colloids and Surfaces B: Biointerfaces, 2015
Colloidal stabile nanoerythrosomes with 200 nm average diameter were formed from hemoglobin-free erythrocyte ghost membrane via sonication and membrane extrusion. The incorporation of extra lipid (1,2-dipalmitoyl-sn-glycero-3-phosphocholine, DPPC), added to the sonicated ghosts, caused significant changes in the thermotropic character of the original membranes. As a result of the increased DPPC ratio the chain melting of the hydrated DPPC system and the characteristic small angle X-ray scattering (SAXS) of the lipid bilayers appeared. Significant morphological changes were followed by transmission electron microscopy combined with freeze fracture method (FF-TEM). After the ultrasonic treatment the large entities of erythrocyte ghosts transformed into nearly spherical nanoerythrosomes with diameters between 100-300 nm and at the same time a great number of 10-30 nm large membrane proteins or protein clusters were dispersed in the aqueous medium. The infrared spectroscopy (FT-IR) pointed out, that the sonication did not cause changes in the secondary structures of the membrane proteins under our preparation conditions. About fivefold of extra lipidcompared to the lipid content of the original membranecaused homogeneous dispersion of nanoerythrosomes however the shape of the vesicles was not uniform. After the addition of about tenfold of DPPC, monoform and monodisperse nanoerythrosomes became typical. The outer surfaces of these roughly spherical objects were frequently polygonal, consisting of a net of pentagons and hexagons..
Characterization of Microvesicles Released from Human Red Blood Cells
Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology, 2016
Extracellular vesicles (EVs) are spherical fragments of cell membrane released from various cell types under physiological as well as pathological conditions. Based on their size and origin, EVs are classified as exosome, microvesicles (MVs) and apoptotic bodies. Recently, the release of MVs from human red blood cells (RBCs) under different conditions has been reported. MVs are released by outward budding and fission of the plasma membrane. However, the outward budding process itself, the release of MVs and the physical properties of these MVs have not been well investigated. The aim of this study is to investigate the formation process, isolation and characterization of MVs released from RBCs under conditions of stimulating Ca2+ uptake and activation of protein kinase C. Experiments were performed based on single cell fluorescence imaging, fluorescence activated cell sorter/flow cytometer (FACS), scanning electron microscopy (SEM), atomic force microscopy (AFM) and dynamic light sc...
Sample preparation and imaging of erythrocyte cytoskeleton with the atomic force microscopy
Cell Biochemistry and Biophysics, 2003
A novel method for the covalent attachment of erythrocytes to glass microscope coverslips that can be used to image intact cells and the cytoplasmic side of the cell membrane with either solid or liquid mode atomic force microscopy (AFM) is described. The strong binding of cells to the glass surface is achieved by the interaction of cell membrane carbohydrates to lectin, which is bound to N-5-azido-2-nitrobenzoyloxysuccinimide (ANBNOS)-coated coverslips (1). The effectiveness of this method is compared with the other commonly used methods of immobilizing intact erythrocytes on glass coverslips for AFM observations. Experimental conditions of AFM imaging of biologic tissue are discussed, and typical topographies of the extracellular and the cytoplasmic surfaces of the plasma membrane in the dry state and in the liquid state are presented. Comparison of the spectrin network of cell age-separated erythrocytes has demonstrated significant loss in the network order in older erythrocytes. The changes are quantitatively described using the pixel height histogram and window size grain analysis.
Lipid nanoparticles with erythrocyte cell-membrane proteins
Journal of Molecular Liquids, 2023
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Morphology and Formation Mechanisms of Cellular Vesicles Harvested from Blood
Physiology, 2022
Theoretical and experimental evidence on cellular vesicles (CVs) isolated from blood is presented. It is suggested that comparison of the observed shapes with theoretical shapes obtained by minimization of membrane-free energy in combination with electron microscopy is key in the assessment of CV identity. We found that shapes of CVs isolated from blood by repetitive centrifugation (up to 20.000 g) and washing, and observed by scanning electron microscopy (SEM) agreed well with theoretically observed shapes. It is indicated that these CVs are colloids deriving from residual blood cells, mostly platelets. SEM images of washed erythrocytes undergoing budding and transmission electron microscopy (TEM) images of isolated erythrocyte microvesicles likewise showed smooth shapes that we described as characteristic for colloidal CVs. Besides these, the CV isolates may contain other small particles, such as exosomes and viruses, as observed in isolates from tomato homogenate, however, we cou...