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Query: UMLS:C0002895 (sickle cell disease)
 11,747 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The phospholipids of the human red cell are distributed asymmetrically in the bilayer of the red cell membrane. In certain pathologic states, such as sickle cell anemia, phospholipid asymmetry is altered. Although several methods can be used to measure phospholipid organization, small organizational changes have been very difficult to assess. Moreover, these methods fail to identify subpopulations of cells that have lost their normal phospholipid asymmetry. Using fluorescently labeled annexin V in flow cytometry and fluorescent microscopy, we were able to identify and quantify red cells that had lost their phospholipid asymmetry in populations as small as 1 million cells. Moreover, loss of phospholipid organization in subpopulations as small as 0.1% of the total population could be identified, and individual cells could be studied by fluorescent microscopy. An excellent correlation was found between fluorescence-activated cell sorter (FACS) analysis results using annexin V to detect red cells with phosphatidylserine (PS) on their surface and a PS-requiring prothrombinase assay using similar red cells. Cells that bound fluorescein isothiocyanate (FITC)-labeled annexin V could be isolated from the population using magnetic beads covered with an anti-FITC antibody. Evaluation of blood samples from patients with sickle cell anemia under oxygenated conditions demonstrated the presence of subpopulations of cells that had lost phospholipid asymmetry. While only a few red cells were labeled in normal control samples (0.21% +/- 0.12%, n = 8), significantly increased (P < .001) annexin V labeling was observed in samples from patients with sickle cell anemia (2.18% +/- 1.21%, n = 13). We conclude that loss of phospholipid asymmetry may occur in small subpopulations of red cells and that fluorescently labeled annexin V can be used to quantify and isolate these cells.
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PMID:Detection of altered membrane phospholipid asymmetry in subpopulations of human red blood cells using fluorescently labeled annexin V. 856 45

Recombinant human annexin V was bound covalently to 9 nm maghemite (gamma Fe2O3) nanoparticles, yielding annexin-ferrofluid (AnxFF), and used to separate annexin-bound red blood cells (RBC) in a magnetic field and estimate their percentage in various bloods. Annexin binding in normal human RBC increased proportionately with storage from 8% on day 2 to 42% on day 100. Enhanced AnxFF binding was associated with various pathologies. Thus, normal blood contained 10.7 +/- 5.9% AnxFF binding RBC; bloods with normal sedimentation rates (albeit with some disease necessitating analysis) contained 23.5 +/- 6.2%; those with high sedimentation rates contained 51.5 +/- 12.3%; sickle cell anaemia patients' blood contained 50.0 +/- 9.3%, and bloods from patients with other pathologies (deforming rheumatic disease, cancer necessitating chemotherapy, etc.) contained 58.6 +/- 7.6% AnxFF binding RBC. Enhanced Ca+2-dependent annexin binding reflects a loss of the asymmetric distribution of anionic phospholipids in plasma membranes which may constitute a signal for the destruction of the modified cells by the reticuloendothelial system. Once these preliminary results are confirmed, the determination of the fraction of AnxFF bound erythrocytes, following their magnetic separation, could prove a simple and rapid quality test for example in the context of blood transfusion.
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PMID:Use of annexin V-ferrofluid to enumerate erythrocytes damaged in various pathologies or during storage in vitro. 857 91

Increased exposure of phosphatidylserine (PS) in erythrocytes has been postulated to contribute to the pathophysiology of sickle cell disease because of its possible effects on blood coagulation, cell adhesion, and cell clearance. We developed a flow-cytometric assay to measure exposure of PS on the outer face of the erythrocyte membrane based on addition of fluorescein-annexin V to whole-blood specimens. The assay correlated linearly with binding of 125I-annexin V (r2 = .95, n = 125 samples). Normal donors (n = 30) showed virtually no annexin-positive cells (0.34% +/- 0.18% for 24-hour old samples). In contrast, annexin V binding was above the upper limit of normal in 96% of 205 specimens from 17 adult sickle cell and 2 beta-thalassemia patients; the mean percentage of annexin-positive cells was 2.86% +/- 2.00% (range, 0.4% to 12.0%). Values varied substantially over time for some patients, and mean values varied between patients. The percentage of annexin-positive cells always decreased after transfusion (11 events in 6 patients), and out of proportion to the amount of blood transfused. In conclusion, increased exposure of PS on a subpopulation of erythrocytes in vivo is a virtually universal feature of sickle cell disease, and its measurement may be useful to evaluate clinical status and response to therapeutic measures such as blood transfusion.
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PMID:Increased erythrocyte phosphatidylserine exposure in sickle cell disease: flow-cytometric measurement and clinical associations. 878 47

It has recently been proved that, in vitro, red blood cells (RBCs) from patients with homozygous beta-thalassemia behave as procoagulant cells. The procoagulant activity of beta-thalassemia RBCs might be the result of an increased exposure of procoagulant phospholipids (i.e. phosphatidylserine) in the outer leaflet of the membrane. In order to test this hypothesis, we compared the catalytic properties of RBCs of patients with beta-thalassemia and homozygous sickle cell disease (SS-RBCs) with that of controls. The catalytic parameters (Km, kcat) of prothrombin activation by factor Xa were determined both in the absence and in the presence of RBCs. The turn-over number (kcat) of the reaction was not modified by normal, SS- or beta-thalassemia RBCs. The Km was lower in the presence of normal RBCs (mean value: 9.1 microM) than in the absence of cells (26 microM). The Km measured in the presence of either SS-RBCs (mean value: 1.6 microM) or beta-thalassemia RBCs (mean value: 1.5 microM) was significantly lower compared to normal RBCs (p < 0.001). No significant difference was observed between SS-RBCs and beta-thalassemia RBCs. Annexin V, a protein with high affinity and specificity for anionic phospholipids, inhibited the procoagulant activity of both SS-RBCs and beta-thalassemia RBCs, in a dose-dependent manner. More than 95% inhibition was achieved at nanomolar concentrations of annexin V. These results indicate that the procoagulant activity of both beta-thalassemia RBCs and SS-RBCs may be fully ascribed to an abnormal exposure of phosphatidylserine at the outer surface of the red cells.
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PMID:Increased procoagulant activity of red blood cells from patients with homozygous sickle cell disease and beta-thalassemia. 888 64

Oxidant-induced damage has been proposed to be the underlying mechanism for loss of membrane phospholipid asymmetry in the erythrocyte membrane. In sickle cell disease, thalassemia, and diabetes as well as in senescent erythrocytes, an apparent correlation between oxidative damage and loss of phosphatidylserine asymmetry has been reported. In the present study, erythrocytes were subjected to various levels of oxidative stress and/or sulfhydryl modifying agents. The transmembrane location of phosphatidylserine (PS) was assessed by FITC-conjugated annexin V labeling and the PS-dependent prothrombinase assay. Transbilayer movement of spin-labeled PS was used to determine aminophospholipid translocase activity. Our data show that cells did not expose PS as the result of oxidative stress induced by phenylhydrazine, hydrogen peroxide, tert-butyl hydroperoxide, cumene hydroperoxide, or sulfhydryl modification by N-ethylmaleimide (NEM) and diamide, even under conditions that led to severe cellular damage and impairment of aminophospholipid translocase activity. In contrast, the increase of intracellular calcium induced by treatment with calcium and ionophore A23187 leads to a rapid scrambling of the lipid bilayer and the exposure of PS, which can be exacerbated by the inhibition of aminophospholipid translocase activity. Oxidation of the cells with hydrogen peroxide or phenylhydrazine did not affect A23187-induced uptake of calcium, but partly inhibited calcium-induced membrane scrambling. In conclusion, oxidative damage of erythrocytes does not induce exposure of phosphatidylserine on the membrane surface, but can interfere with both aminophospholipid translocase activity and calcium-induced randomization of membrane phospholipids.
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PMID:Oxidative damage does not alter membrane phospholipid asymmetry in human erythrocytes. 918 59

Phosphatidylserine (PS) normally localizes to the inner leaflet of cell membranes but becomes exposed in abnormal or apoptotic cells, signaling macrophages to ingest them. Along similar lines, it seemed possible that the removal of red cells from circulation because of normal aging or in hemolytic anemias might be triggered by PS exposure. To investigate the role of PS exposure in normal red cell aging, we used N-hydroxysuccinimide-biotin to tag rabbit red cells in vivo, then used phycoerythrin-streptavidin to label the biotinylated cells, and annexin V-fluorescein isothiocyanate (FITC) to detect the exposed PS. Flow cytometric analysis of these cells drawn at 10-day intervals up to 70 days after biotinylation indicated that older, biotinylated cells expose more PS. Furthermore, our data match a simple model of red cell senescence that assumes both an age-dependent destruction of senescent red cells preceded by several hours of PS exposure and a random destruction of red cells without PS exposure. By using this model, we demonstrated that the exposure of PS parallels the rate at which biotinylated red cells are removed from circulation. On the other hand, using an annexin V-FITC label and flow cytometry demonstrates that exposed PS does not cause the reduced red cell life span of patients with hemolytic anemia, with the possible exception of those with unstable hemoglobins or sickle cell anemia. Thus, in some cases PS exposure on the cell surface may signal the removal of red cells from circulation, but in other cases some other signal must trigger the sequestration of cells.
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PMID:Phosphatidylserine exposure and red cell viability in red cell aging and in hemolytic anemia. 950 Dec 18

The use of cell-targeted ferrofluid in the characterization of modifications of cell membranes is reviewed. Maghemite ferrofluid was synthesized by the Massart method, complexed with dimercaptosuccinic acid (FF). Cell targeting by FF was developed by coupling FF to various biological effectors such as antibodies, lectins, etc, which enabled magnetic cell sorting. Modifications in erythrocyte membranes were studied using FF bound to recombinant human annexin V (AnxFF) which is very sensitive, compared to other Anx-based reagents, in the early detection of phosphatidylserine (PS) exposition on the outer leaflet of the plasma membrane. Thus PS exposition on mouse RBC was detected already after a 24-h storage at 4 degrees C and, transiently, 24 h after their infection by Plasmodium parasites, at which time the parasites are still confined to the liver, thus leading to the recruitment of young RBC and the accumulation of a species, intermediate between reticulocytes and erythrocytes, and the actual RBC target of plasmodial invasion. AnxFF revealed PS exposition on RBC from sickle cell anemia patients, following various inflammations and already after 20 days of human blood storage under blood bank conditions. Such a sensitive detection should be similar to that of macrophages which recognize exposed PS on cells and bring about the latter's elimination from the circulation. AnxFF binding determination was combined with that of cell electrophoretic mobility, glycerol resistance and filterability to characterize RBC membrane modifications in Alzheimer's disease patients which suggested a continuous damage and regeneration in RBC of these patients. A logistic analysis suggested that several three-parameter combinations could permit diagnosis of Alzheimer's disease with up to 95% accuracy. THP1 cells and macrophages, derived themselves by incubation with retinoic acid, were bound to FF and placed in a radio frequency alternating magnetic field. Magnetocytolysis was associated with FF attachment to the cells without damage to non-bound cells and without heating of the surrounding solution.
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PMID:Biomedical applications of maghemite ferrofluid. 978 79

In pathological conditions such as sickle cell disease, falciparum malaria and diabetes, an abnormal adherence of erythrocytes to endothelium is concomitant with loss of phospholipid asymmetry resulting in phosphatidylserine (PS) exposure. We have investigated the involvement of PS in this interaction by studying adhesion of human erythrocytes, treated with Ca2+-ionophore A23187 in combination with N-ethylmaleimide, to human umbilical vein endothelial cells in a flow-based assay. Results showed that erythrocytes which exposed PS, massively adhered to HUVEC in a Ca2+-dependent manner. This adhesion was inhibited by PS liposomes and by annexin V, giving clear evidence of the PS dependence of these interactions.
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PMID:Phosphatidylserine-related adhesion of human erythrocytes to vascular endothelium. 1058 15

Phospholipid asymmetry is well maintained in erythrocyte (RBC) membranes with phosphatidylserine (PS) exclusively present in the inner leaflet. The appearance of PS on the surface of the cell can have major physiologic consequences, including increased cell-cell interactions. Because increased adherence of PS-exposing RBCs to endothelial cells (ECs) may be pathologically important in hemoglobinopathies such as sickle cell disease and thalassemia, we studied the role of PS exposure in calcium ionophore-treated normal RBC adherence to human umbilical vein endothelial cell (HUVEC) monolayers. When HUVEC monolayers were incubated with these PS-exposing RBCs, the ECs retracted and the RBCs adhered primarily in the gaps opened between the ECs. A linear correlation was found between the number of PS-exposing RBCs in the population and the number of adhering RBCs to the monolayer. Pretreatment of RBCs with annexin V significantly decreased adherence by shielding PS on the RBCs. Similarly, PS-containing lipid vesicles decreased RBC binding by competing for the PS binding sites in the monolayer. PS-exposing RBCs and PS-containing lipid vesicles adhered to immobilized thrombospondin (TSP) and matrix TSP, respectively, and adherence of PS-exposing RBCs to EC monolayers was reduced by antibodies to TSP and to its EC receptor, alpha(v)beta(3). Together, these results indicate a role for PS and matrix TSP in the adherence of PS-exposing RBCs to EC monolayers, and suggest an important contribution of PS-exposing RBCs in pathologies with reported vascular damage, such as sickle cell anemia. (Blood. 2000;95:1293-1300)
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PMID:Adherence of phosphatidylserine-exposing erythrocytes to endothelial matrix thrombospondin. 1066 2

Thrombosis is a major complication of human hemolytic anemias such as sickle cell disease, thalassemia, and severe hereditary spherocytosis (HS). Mice with severe HS and severe hereditary elliptocytosis (HE) also suffer from thrombosis, with incidences ranging from 15 and 22% in beta-spectrin- and ankyrin-deficient mice, respectively, to 85 to 100% in alpha-spectrin-deficient and band 3 knockout mice. A contributing factor to thrombosis could be loss of phospholipid asymmetry of the mutant red blood cells (RBCs), with concomitant exposure of the aminophospholipid phosphatidylserine (PS). Increased PS exposure occurs in RBCs from sickle cell and thalassemia patients and in RBCs from band 3-deficient mice. To determine if increased PS exposure correlates with thrombotic risk in HS and HE mice with ankyrin, beta-spectrin, and alpha-spectrin deficiencies, measurements of FITC-labeled annexin V binding to externalized PS on RBCs were performed. PS exposure is elevated in all mice with HS and HE, but the percentage of RBCs with exposed PS does not correlate with thrombotic risk in these mice.
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PMID:Erythroid phosphatidyl serine exposure is not predictive of thrombotic risk in mice with hemolytic anemia. 1077 78


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