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1.
The human erythrocyte membrane skeleton may be an ionic gel 总被引:3,自引:0,他引:3
In the first paper in this series (Stokke et al. Eur Biophys J 1986, 13:203-218) we developed the general theory of the mechanochemical properties and the elastic free energy of the protein gel--lipid bilayer membrane model. Here we report on an extensive numerical analysis of the human erythrocyte shapes and shape transformations predicted by this new cell membrane model. We have calculated the total elastic free energy of deformation of four different cell shape classes: disc-shaped cells, cup-shaped cells, crenated cells, and cells with membrane invaginations. We find that which of these shape classes is favoured depends strongly on the spectrin gel osmotic tension, IIGu, and the surface tensions, IIEu and IIPu, of the extracellular and protoplasmic halves of the membrane lipid bilayer, respectively. For constant ratio IIEu/IIPu greater than O large negative or positive values of IIGu favour respectively the crenated and invaginated cell shape classes. For small absolute values of IIGu, IIEu, and IIPu, biconcave or cup-shaped cells are the stable ones. Our numerical analysis shows that the higher the membrane skeleton compressibility is, the smaller are the values of IIGu needed to induce cell shape transformation. We find that the stable and metastable shapes of discocytes and stomatocytes generally depend both on the shape of the stressfree membrane skeleton and the membrane skeleton compressibility. 相似文献
2.
Biochemical and biophysical observations indicate that the erythrocyte membrane skeleton is composed of a swollen network of long, flexible and ionizable macromolecules located at the cytoplasmic surface of the fluid membrane lipid bilayer. We have analyzed the mechanochemical properties of the erythrocyte membrane assuming that the membrane skeleton constitutes an ionic gel (swollen ionic elastomer). Using recently established statistical thermodynamic theory for such gels, our analysis yields mathematical expressions for the mechanochemical properties of erythrocyte membranes that incorporate membrane molecular parameters to an extent not achieved previously. The erythrocyte membrane elastic shear modulus and maximum elastic extension ratio predicted by our membrane model are in quantitative agreement with reported values for these parameters. The gel theory predicts further that the membrane skeleton modulus of area compression, K
G, may be small as well as large relative to the membrane elastic shear modulus, G, depending on the environmental conditions. Our analysis shows that the ratio between these two parameters affects both the geometry and the stability of the favoured cell shapes. 相似文献
3.
Micropipette aspiration of human erythrocytes induces echinocytes via membrane phospholipid translocation.
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G M Artmann K L Sung T Horn D Whittemore G Norwich S Chien 《Biophysical journal》1997,72(3):1434-1441
When a discocytic erythrocyte (RBC) was partially aspirated into a 1.5-microns glass pipette with a high negative aspiration pressure (delta P = -3.9 kPa), held in the pipette for 30 s (holding time, th), and then released, it underwent a discocyte-echinocyte shape transformation. The degree of shape transformation increased with an increase in th. The echinocytes recovered spontaneously to discocytes in approximately 10 min, and there was no significant difference in recovery time at 20.9 degrees C, 29.5 degrees C, and 37.4 degrees C, respectively. At 11 degrees C the recovery time was significantly elevated to 40.1 +/- 6.7 min. At 20.9 degrees C the shape recovery time varied directly with the isotropic RBC tension induced by the pipetting. Sodium orthovanadate (vanadate, 200 microM), which inhibits the phospholipid translocase, blocks the shape recovery. Chlorpromazine (CP, 25 microM) reversed the pipette-induced echinocytic shape to discocytic in < 2 min, and the RBC became a spherostomatocyte-II after another 30 min. It was hypothesized that the increase in cytosolic pressure during the pipette aspiration induced an isotropic tension in the RBC membrane followed by a net inside-to-outside membrane lipid translocation. After a sudden release of the aspiration pressure the cytosolic pressure and the membrane tension normalized immediately, but the translocated phospholipids remained temporarily "trapped" in the outer layer, causing an area excess and hence the echinocytic shape. The phospholipid translocase activity, when not inhibited by vanadate, caused a gradual return of the translocated phospholipids to the inner layer, and the RBC shape recovered with time. 相似文献
4.
M J Saxton 《Biophysical journal》1990,57(6):1167-1177
The spectrin network on the cytoplasmic surface of the erythrocyte membrane is modeled as a triangular lattice of spectrin tetramers. This network obstructs lateral diffusion of proteins and provides mechanical reinforcement to the membrane. These effects are treated in a systematic and unified manner in terms of a percolation model. The diffusion coefficient is obtained as a function of the fraction of normal spectrin tetramers for both static and fluctuating barriers. The elasticity of the network is calculated as a function of the fraction of normal spectrin and the ratio of bending to stretching energies. For static barriers, elasticity and lateral diffusion are incompatible: if a network is connected enough to be elastic, it is connected enough to block long-range lateral diffusion. The elasticity and the force required for mechanical breakdown go to zero at the percolation threshold; experimental evidence suggests the existence of a stability threshold at or near the percolation threshold. The model is qualitatively applicable to other cells with membrane skeletons, such as epithelial cells, in which localization of membrane proteins is essential to differentiation. 相似文献
5.
Simulations of the erythrocyte cytoskeleton at large deformation. II. Micropipette aspiration. 总被引:5,自引:2,他引:5
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Coarse-grained molecular models of the erythrocyte membrane's spectrin cytoskeleton are presented in Monte Carlo simulations of whole cells in micropipette aspiration. The nonlinear chain elasticity and sterics revealed in more microscopic cytoskeleton models (developed in a companion paper; Boey et al., 1998. Biophys. J. 75:1573-1583) are faithfully represented here by two- and three-body effective potentials. The number of degrees of freedom of the system are thereby reduced to a range that is computationally tractable. Three effective models for the triangulated cytoskeleton are developed: two models in which the cytoskeleton is stress-free and does or does not have internal attractive interactions, and a third model in which the cytoskeleton is prestressed in situ. These are employed in direct, finite-temperature simulations of erythrocyte deformation in a micropipette. All three models show reasonable agreement with aspiration measurements made on flaccid human erythrocytes, but the prestressed model alone yields optimal agreement with fluorescence imaging experiments. Ensemble-averaging of nonaxisymmetrical, deformed structures exhibiting anisotropic strain are thus shown to provide an answer to the basic question of how a triangulated mesh such as that of the red cell cytoskeleton deforms in experiment. 相似文献
6.
Na2SeO3 supplementation in the ageing medium could protect aged erythrocyte ghosts from decreases in lipid fluidity, Na, K-ATPase activity, and sensitivity to ouabain. Results also showed that Se could obviously prevent the dissociation of spectrin from the erythrocyte membrane. Furthermore, Se could markedly promote the reassociation of spectrin with the spectrin-stripped inside out membrane vesicles(IOVs) of erythrocytes. The protective action of Se on biomembranes is generally interpreted in terms of the activity of Se-containing glutathione peroxidase (GSHPx). However, since GSHPx is mainly distributed in the cytoplasm of erythrocytes, the stabilizing effect of Se on erythrocyte membranes might not be related to the activity of this enzyme. 相似文献
7.
Filamentous skeletons were liberated from isolated human erythrocyte membranes in Triton X-100, spread on fenestrated carbon films, negatively stained, and viewed intact and unfixed in the transmission electron microscope. Two forms of the skeleton were examined: (a) basic skeletons, stripped of accessory proteins with 1.5 M NaCl so that they contain predominantly polypeptide bands 1, 2, 4.1, and 5; and (b) unstripped skeletons, which also bore accessory proteins such as ankyrin and band 3 and small plaques of residual lipid. Freshly prepared skeletons were highly condensed. Incubation at low ionic strength and in the presence of dithiothreitol for an hour or more caused an expansion of the skeletons, which greatly increased the visibility of their elements. The expansion may reflect the opening of spectrin from a compact to an elongated disposition. Expanded skeletons appeared to be organized as networks of short actin filaments joined by multiple (5-8) spectrin tetramers. In unstripped preparations, globular masses were observed near the centers of the spectrin filaments, probably corresponding to complexes of ankyrin with band 3 oligomers. Some of these globules linked pairs of spectrin filaments. Skeletons prepared with a minimum of perturbation had thickened actin protofilaments, presumably reflecting the presence of accessory proteins. The length of these actin filaments was highly uniform, averaging 33 +/- 5 nm. This is the length of nonmuscle tropomyosin. Since there is almost enough tropomyosin present to saturate the F-actin, our data support the hypothesis that tropomyosin may determine the length of actin protofilaments in the red cell membrane. 相似文献
8.
High resolution two-dimensional gel electrophoresis of human erythrocyte membrane proteins. 总被引:1,自引:0,他引:1
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Three different two-dimensional (2-D) gel electrophoretic techniques have been modified to provide high resolution of human erythrocyte membrane proteins. The resulting gels were referenced to the established one-dimensional (1-D) sodium dodecylsulfate (SDS) gel electrophoretic profile, and the effects of endogenous proteolysis and cytosolic contamination were studied. It is concluded that in vitro proteolysis and cytosolic contamination do not contribute significantly to the patterns observed on the 2-D gels, under the conditions used for erythrocyte ghost preparation. The procedures require only small quantities of blood; as many as twenty 2-D gel profiles can be obtained from 5 ml of blood. The combination of nonequilibrium isoelectric focusing (IEF) in the first dimension, SDS electrophoresis in the second dimension, and very sensitive silver staining techniques resolves more than 250 individual protein spots. This appears to be the most useful single procedure for the analysis of red cell membrane proteins. Membrane protein profiles from patients with Duchenne muscular dystrophy, Wernicke-Korsakoff syndrome, and acanthocytosis with degeneration of the basal ganglia were compared with normal controls. The patterns for Duchenne muscular dystrophy and Wernicke-Korsakoff syndrome were not different from normal patterns. The pattern for the patient with acanthocytosis and degeneration of the basal ganglia consistently showed a high level for one protein in the 100,000 mol. wt. range. 相似文献
9.
In the erythrocyte membrane, the mobility of band 3 protein, the receptor for concanavalin A (Con A), is drastically reduced by the membrane skeleton. Yet, the vesicles free of membrane skeletal proteins, isolated from the highly agglutinable proteinase-treated cells, are found to be devoid of Con A agglutinability. The vesicles bind Con A in normal amounts, and remain agglutinable with the wheat germ and Ricinus agglutinins. Intracellular entrapment of monospecific antibodies to spectrin and 4.1 protein (two of the major skeletal components of the membrane) is also found to inhibit agglutination by 30-50%. Thus the membrane skeleton appears to play a positive role in the agglutination of the cells with Con A. The anti-ankyrin antibodies are found to be without any effect. The anti-band 3 (cytoplasmic domain) antibodies are also inhibitory to agglutination. Since Con A binding to cells alters the shape responses and deformability of the cells, and the cells resist fragmentation at 49 degrees C, the properties of the whole skeleton, especially spectrin, appear to be changed. The Con A-bound membranes also do not release the complex of spectrin-band 4.1-actin when extracted with a hypotonic medium. It appears that Con A binding leads to interaction of the cytoplasmic domain of the receptor with a skeletal component, possibly spectrin. Subsequent to this, the receptor molecules and the skeletal proteins undergo aggregation in the membrane, which is detected by their crosslinking by an 8.6-A span bifunctional reagent. The contractility believed to be associated with the membrane skeleton may be responsible for the aggregation. 相似文献
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11.
A sample of 12 anencephalic fetuses with gestational ages ranging from 26 to 40 weeks and exhibiting varying degrees of severity of the dorsal cranial defect was compared to three normal fetuses of comparable gestational ages with regard to the morphology and positional relationships of the maxillofacial skeletal complex. Gross dissection, alizarin red S staining, radiographs, cephalometric tracings, and histologic techniques were utilized. It was found that some facial bones were severely affected in morphology, size, spatial and angular relationships. The manner in which these were altered suggests that their morphogenesis is an adaptation to the primary defect of the neurocranium. 相似文献
12.
13.
We have examined fragments of the filamentous network underlying the human erythrocyte membrane by high-resolution electron microscopy. Networks were released from ghosts by extraction with Triton X-100, freed of extraneous proteins in 1.5 M NaCl, and collected by centrifugation onto a sucrose cushion. These preparations contained primarily protein bands 1 + 2 (spectrin), band 4.1 and band 5 (actin). The networks were partially disassembled by incubation at 37 degrees C in 2 mM NaPi (pH 7), which caused the preferential dissociation of spectrin tetramers to dimers. The fragments so generated were fractionated by gel filtration chromatography and visualized by negative staining with uranyl acetate on fenestrated carbon films. Unit complexes, which sedimented at approximately 40S, contained linear filaments approximately 7-8 nm diam from which several slender and convoluted filaments projected. The linear filaments had a mean length of 52 +/- 17 nm and a serrated profile reminiscent of F-actin. They could be decorated in an arrowhead pattern with S1 fragments of muscle heavy meromyosin which, incidentally, displaced the convoluted filaments. Furthermore, the linear filaments nucleated the polymerization of rabbit muscle G-actin, predominantly but not exclusively from the fast-growing ends. On this basis, we have identified the linear filaments as F-actin; we infer that the convoluted filaments are spectrin. Spectrin molecules were usually attached to actin filaments in clusters that showed a preference for the ends of the F-actin. We also observed free globules up to 15 nm diam, usually associated with three spectrin molecules, which also nucleated actin polymerization; these may be simple junctional complexes of spectrin, actin, and band 4.1. In larger ensembles, spectrin tetramers linked actin filaments and/or globules into irregular arrays. Intact networks were an elaboration of the basic pattern manifested by the fragments. Thus, we have provided ultrastructural evidence that the submembrane skeleton is organized, as widely inferred from less direct information, into short actin filaments linked by multiple tetramers of spectrin clustered at sites of association with band 4.1. 相似文献
14.
The membrane change in En(a-) human erythrocytes. Absence of the major erythrocyte sialoglycoprotein. 总被引:5,自引:0,他引:5
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We investigated the membrane of En(a-) human erythrocytes as part of a study of the structure and biochemical function of the surface glycoproteins of the mammalian cell. 2. En(a-) erythrocytes were selected because they have more extensive changes at the cell surface than any other known erythrocyte variant. 3. Our results show that in En(a-) erythrocytes: (a) the major membrane sialoglycoprotein is lacking; (b) the other major membrane-penetrating glycoprotein (band 3) has an altered electrophoretic mobility. 4. The apparent clinical normality of En(a-) cells suggests that the change in band 3 may compensate for the loss of the membrane sialoglycoproteins. It is clear that a viable erythrocyte can exist despite the absence of one of its major surface components. 相似文献
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18.
S Fujikawa 《Journal of ultrastructure research》1983,84(3):289-298
The effect of fixatives on the membrane skeleton underlying the human erythrocyte membrane was examined by freeze-etching. An anastomosing fibrillar network was readily observed on the protoplasmic surface of the erythrocyte membrane treated with tannic acid. Such structure was much less defined in unfixed membrane or membrane fixed with glutaraldehyde or glutaraldehyde followed by osmium tetraoxide. Tannic acid caused a marked increase in diameter of the fibrillar components of the membrane skeleton and of the protoplasmic surface particles of inside-out vesicles prepared by alkali treatment but did not affect the size of intramembranous particles seen on fracture faces nor the appearance of exoplasmic surfaces. The improved visualization of the membrane skeleton after treatment with tannic acid resulted from interactions between tannic acid and exposed membrane proteins. 相似文献
19.
R Fiorini E Bertoli A Kantar P L Giorgi G Curatola 《Bollettino della Società italiana di biologia sperimentale》1989,65(8):703-710
The process of red blood cell senescence in the blood stream results in many changes in their physical and biochemical properties. In this work we have studied the physico-chemical state of erythrocyte membranes prepared from 5 subpopulations of erythrocytes of different age by using the fluorescence technique. Membrane fluidity has been evaluated by the fluorescence polarization of 1,6-diphenyl-1,3,5-hexatriene (DPH) and a further study of the fluorescence decay of this probe has been performed by multifrequency phase and modulation fluorometry. DPH fluorescence polarization is significantly increased in the membranes prepared from the youngest fraction of erythrocytes, indicating a decreased fluidity without any significant change in DPH fluorescence decay. 相似文献
20.
Concanavalin A binding to human erythrocytes leads to alterations in properties of the membrane skeleton.
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Three properties related to the erythrocyte membrane skeleton are found to be altered after the binding of concanavalin A (Con A) to erythrocytes or their isolated membranes. Con A binding to normal erythrocytes imparts resistance to heat (49 degrees C)-induced fragmentation of the cells. The fragmentation, due to denaturation of spectrin at 49 degrees C, is prevented by Con A in a dose-dependent manner, but levels off at concentrations of Con A in excess of 100 micrograms/ml. The binding of Con A to ghosts isolated from normal, trypsin- or Pronase-treated cells prevents (completely or substantially) the elution of the skeletal protein complex when the membranes are extracted under low-ionic-strength conditions in the cold. The Con A-agglutinated membranes of trypsin- and Pronase-treated, but not normal, cells show cross-linking of skeletal proteins and band 3 with dimethyl adipimidate, a 0.86 nm (8.6 A)-span bifunctional reagent. The extent of cross-linking is greater in the Pronase-treated membrane than in the less-agglutinable trypsin-treated membranes. The results show that, after Con A has bound, rearrangements occur in the membrane that alter properties of the skeletal proteins. Additionally, redistribution of the skeletal proteins and the Con A receptor occurs in the lectin-agglutinated membranes. 相似文献