Liposomes composed of a double-chain cationic amphiphile (vectamidine) induce their own encapsulation into human erythrocytes.
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Vectamidine is a liposome-forming double-chain cationic amphiphile. The present work was aimed to microscopically study the interactions of Vectamidine liposomes with the human erythrocyte plasma membrane. Vectamidine rapidly induced stomatocytic shapes. Attachment of Vectamidine liposomes to the erythrocyte induced a strong local invagination of the membrane. This frequently resulted in a complete encapsulation of the liposome. Liposomes composed of phosphatidylcholine (neutral) or phosphatidylserine/phosphatidylcholine (anionic) did not perturb the erythrocyte shape. Our results indicate that besides an attraction of Vectamidine liposomes to the plasma membrane, there is a preference of Vectamidine for the inner bilayer leaflet. We suggest that cationic amphiphiles may transfer from membrane-attached liposomes to the plasma membrane and then translocate to the inner bilayer leaflet where they induce a strong local inward bending of the plasma membrane resulting in an encapsulation of the liposome. (+info)
Static and magic angle spinning (31)P NMR spectroscopy of two natural plasma membranes.
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Static and magic angle spinning (31)P NMR spectroscopy was used for the first time in natural plasma membranes from erythrocytes and skeletal muscle to study phospholipid arrangement and composition. Typical static powder-like spectra were obtained showing that phospholipids were in a bilayer arrangement. Magic angle spinning narrowed spectra into two components. The first one corresponded to phosphatidylcholine and the second one to the other phospholipids with intensities in agreement with the known phospholipid composition. These findings show that NMR data previously acquired using model membranes can be transposed to studies on phospholipids in their natural environment. (+info)
Tn-syndrome.
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The idiopathic Tn-syndrome, formerly called 'permanent mixed-field polyagglutinability', is a rare hematological disorder characterized by the expression of the Tn-antigen on all blood cell lineages. The immunodominant epitope of the Tn-antigen is terminal alpha-N-acetylgalactosamine, O-glycosidically linked to protein. Normally this residue is 3'-substituted by 5-galactose thereby forming the core 1 structure known as the Thomsen-Friedenreich (TF) antigen (Galbeta1 ==> 3GalNAcalpha1 ==> Thr/Ser). The cause of the exposure of the Tn-antigen appears to be due to the silencing of the gene expression of beta1,3galactosyltransferase, since treatment of deficient Tn(+) lymphocyte T clones with 5'azacytidine or Na butyrate leads to reexpression of enzyme activity and the sialylated TF-antigen. The Tn-syndrome is acquired and permanent and affects both sexes at any age. Its origin is unknown. Pluripotent stem cells are affected since all lineages are involved but each one to a variable extent. Therefore, normal cells co-exist with Tn-transformed cells. Clinically, patients suffering from the Tn-syndrome appear healthy. Laboratory findings usually reveal moderate thrombocyto- and leukopenia and some signs of hemolytic anemia not warranting any treatment. (+info)
Biochemical background of paroxysmal nocturnal hemoglobinuria.
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Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired disorder characterized by paroxysms of intravascular hemolysis. A considerable part of erythrocytes in patient blood is susceptible to autologous complement activation because of the deficiency of CD59, which is a glycosylphosphatidylinositol (GPI)-anchored protein and inhibits the formation of the membrane attack complex (MAC) of complement. The deficiency of CD59 is derived from the inability of GPI-anchor synthesis. Although more than 10 proteins are involved in the GPI-anchor synthesis, the mutation of only one protein, PIG-A, causes the defect in about 200 patients with PNH who have been analyzed. The reason why only PIG-A causes the deficiency of GPI anchor is due to the location of its gene on X chromosome. The clonal stem cell mutated with PIG-A gene in the bone marrow loses the capability of the synthesis of GPI-anchor. The mutation of PIG-A gene alone, however, seems to be insufficient to account for the survival of the PIG-A-deficient cells in the bone marrow. Thus, a fraction of the mutant stem cells probably gain a survival advantage by some additional changes, either additional mutations or changes in immunological circumstances. The release of the surviving cells into blood stream results in a clinical syndrome with PNH. (+info)
Erythrocyte membranes undergo cooperative, pH-sensitive state transitions in the physiological temperature range: evidence from Raman spectroscopy.
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We have examined the Raman scattering from erythrocyte ghosts at 2700 to 3000 cm-1 (CH-stretching region). Plots of the intensity (I) of the 2930 cm-1 band relative to the intensity of the thermally stable 2850 cm-1 band, i.e., the [I2930/I2850] ratio, as a function of temperature reveal a sharp discontinuity, which at pH 7.4 has a lower limit of 38 degrees and is irreversible above 42 degrees. [I2930/I2850] is stable between pH 7.0 and pH 7.4, but increases or decreases sharply below pH 7.0 or above pH 7.5, respectively. Reduction of pH to 6.5 lowers the transition temperature by about 16 degrees, and a shift to pH 6.0 drops the transition range to 0 to 7 degrees. The above effects of temperature and pH on Raman scattering closely correspond to those detected by studies on the interaction of membrane protein fluorophores and lipid-soluble fluorescence quenchers [Bieri, V. and Wallach, D.F.H. (1975) Biochim. Biophys. Acta 406, 415-423]. Taken together, these results suggest that the transitions represent concerted process, involving hydrophobic amino acid residues and lipid chains at apolar protein-lipid boundaries. (+info)
Antigen structure and genetic basis of histo-blood groups A, B and O: their changes associated with human cancer.
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Three areas of research involved in blood group (or histo-blood group) ABO antigens and their genes, developed by our research group, are reviewed: (1) Antigen structures. The structural basis of A and H, A(1) and A(2), i and I antigens expressed in erythrocyte membranes. Major carriers of A and H determinants in erythrocytes are type 2 chain poly-LacNAc, short vs. long and unbranched vs. branched structures termed A(a), A(b), A(c), A(d) and H(1), H(2), H(3), H(4). Regular A (A(1)) and weak A (A(2)) were identified respectively as repetitive A (type 3 chain A) and A-associated H. A(1)- and A(2)-specific type 3 chain A and H, type 1 chain (representing Lewis blood group antigens), and type 4 chain (globo-series antigen; an extremely minor component in erythrocytes) are all glycosphingolipids. A and H determinants in fetal and newborn erythrocytes are carried by unbranched poly-LacNAc, whereas these determinants in adult erythrocytes are carried by branched poly-LacNAc. (2) ABO genes. A few cDNAs encoding A enzyme (UDP-GalNAc: H-a-GalNAc transferase) were cloned based on the amino acid sequence of purified A enzyme and their structures were compared with those of homologous cDNA from blood cells of B and O individuals (genotype BB, OO). Four nucleotide substitutions and four corresponding amino acid sequences essential for expression of A(1) allele and B allele, and differences between A and B enzymes, were identified. Amino acids 266 and 268, i.e. Leu and Gly for A enzyme vs. Met and Ala for B enzyme, were dominant in determining A vs. B activity (presumably recognizing UDP-GalNAc vs. UDP-Gal). The A(2) allele was characterized by deletion of the termination codon, extending nucleotides up to 1128 and thus encoding 21 extra amino acids at the C terminus, which may affect (diminish) the dominant function of amino acids 266 and 268. Typical O allele (O(1)) is characterized by deletion of nucleotide 261 G, causing frame shift and encoding of an entirely different, short polypeptide, due to appearance of early termination codon at nucleotide 354. Structures of other O alleles (O(1 v), O(2)) and other weak A alleles (A(3), A(el)) are also described. The genomic structure of ABO genes consists of seven exons which span approximately 19 kb of genomic DNA on chromosome 9, band q34. Most of the coding sequence is located in exon 7. Analysis of the 5' upstream region revealed the presence of the binding site for transcription factors and enhancer element. (3) Antigens and genes in cancer. A and B phenotypes aberrantly expressed in various types of human cancer, and their genetic basis, have been studied. One widely-occurring change observed in a large variety of human cancers is deletion of A or B epitope, associated with accumulation of their precursor H (Le(y), Le(b)), which causes enhanced malignancy. A less-commonly observed change is expression of incompatible A, identified as real type 1 chain A, in tumors of O or B individuals. A possible molecular genetic mechanism leading to such phenotypic changes is discussed. (+info)
Elasticity of the red cell membrane and its relation to hemolytic disorders: an optical tweezers study.
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We have used optical tweezers to study the elasticity of red cell membranes; force was applied to a bead attached to a permeabilized spherical ghost and the force-extension relation was obtained from the response of a second bead bound at a diametrically opposite position. Interruption of the skeletal network by dissociation of spectrin tetramers or extraction of the actin junctions engendered a fourfold reduction in stiffness at low applied force, but only a twofold change at larger extensions. Proteolytic scission of the ankyrin, which links the membrane skeleton to the integral membrane protein, band 3, induced a similar effect. The modified, unlike the native membranes, showed plastic relaxation under a prolonged stretch. Flaccid giant liposomes showed no measurable elasticity. Our observations indicate that the elastic character is at least as much a consequence of the attachment of spectrin as of a continuous membrane-bound network, and they offer a rationale for formation of elliptocytes in genetic conditions associated with membrane-skeletal perturbations. The theory of Parker and Winlove for elastic deformation of axisymmetric shells (accompanying paper) allows us to determine the function BH(2) for the spherical saponin-permeabilized ghost membranes (where B is the bending modulus and H the shear modulus); taking the literature value of 2 x 10(-19) Nm for B, H then emerges as 2 x 10(-6) Nm(-1). This is an order of magnitude higher than the value reported for intact cells from micropipette aspiration. Reasons for the difference are discussed. (+info)
The deformation of spherical vesicles with permeable, constant-area membranes: application to the red blood cell.
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The deformation of an initially spherical vesicle of radius a with a permeable membrane under extensive forces applied at its poles is calculated as a function of the in-plane shear modulus, H, and the out-of-plane bending modulus, B, using an axisymmetric theory that is valid for large deformations. Suitably nondimensionalized, the results depend upon a single nondimensional parameter, C identical with a(2)H/B. For small deformations, the calculated force-polar strain curves are linear and, under these conditions, the slope of the curve determines only C, not the values of H and B separately. Independent determination of H and B from experimental measurements require deformations that are large enough to produce nonlinear behavior. Simple approximations for large and small C are given, which are applied to experimental measurements on red blood cell ghosts that have been made permeable by treatment with saponin. (+info)