scholarly journals Role of Calcium in Phosphatidylserine Externalisation in Red Blood Cells from Sickle Cell Patients

Anemia ◽  
2011 ◽  
Vol 2011 ◽  
pp. 1-8 ◽  
Author(s):  
Erwin Weiss ◽  
David Charles Rees ◽  
John Stanley Gibson
Keyword(s):  
Red Blood Cells ◽  
Sickle Cell ◽  
Blood Cells ◽  
Phosphatidylserine Exposure ◽  
Channel Inhibition ◽  
Cation Conductance ◽  
Gardos Channel ◽  
Cation Permeability

Phosphatidylserine exposure occurs in red blood cells (RBCs) from sickle cell disease (SCD) patients and is increased by deoxygenation. The mechanisms responsible remain unclear. RBCs from SCD patients also have elevated cation permeability, and, in particular, a deoxygenation-induced cation conductance which mediates entry, providing an obvious link with phosphatidylserine exposure. The role of was investigated using FITC-labelled annexin. Results confirmed high phosphatidylserine exposure in RBCs from SCD patients increasing upon deoxygenation. When deoxygenated, phosphatidylserine exposure was further elevated as extracellular [] was increased. This effect was inhibited by dipyridamole, intracellular chelation, and Gardos channel inhibition. Phosphatidylserine exposure was reduced in high saline. levels required to elicit phosphatidylserine exposure were in the low micromolar range. Findings are consistent with entry through the deoxygenation-induced pathway (), activating the Gardos channel. [] required for phosphatidylserine scrambling are in the range achievablein vivo.

Role of Adhesion Molecules and Vascular Endothelium in the Pathogenesis of Sickle Cell Disease

Hematology ◽  
2007 ◽  
Vol 2007 (1) ◽  
pp. 84-90 ◽  
Author(s):  
Marilyn J. Telen
Keyword(s):  
Sickle Cell Disease ◽  
Red Blood Cells ◽  
Adhesion Molecules ◽  
Sickle Cell ◽  
Vascular Endothelium ◽  
Blood Cells ◽  
Cell Disease ◽  
Adhesive Interactions ◽  
Lines Of Evidence

AbstractA number of lines of evidence now support the hypothesis that vaso-occlusion and several of the sequelae of sickle cell disease (SCD) arise, at least in part, from adhesive interactions of sickle red blood cells, leukocytes, and the endothelium. Both experimental and genetic evidence provide support for the importance of these interactions. It is likely that future therapies for SCD might target one or more of these interactions.

scholarly journals Antagonists of the system L neutral amino acid transporter (LAT) promote endothelial adhesivity of human red blood cells

2017 ◽  
Vol 117 (07) ◽  
pp. 1402-1411 ◽  
Author(s):  
Laura Beth Mann Dosier ◽  
Vikram J. Premkumar ◽  
Hongmei Zhu ◽  
Izzet Akosman ◽  
Michael F. Wempe ◽  
...  
Keyword(s):  
Amino Acid ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Amino Acid Transporter ◽  
Neutral Amino Acid ◽  
Neutral Amino Acid Transporter ◽  
System L

SummaryThe system L neutral amino acid transporter (LAT; LAT1, LAT2, LAT3, or LAT4) has multiple functions in human biology, including the cellular import of S-nitrosothiols (SNOs), biologically active derivatives of nitric oxide (NO). SNO formation by haemoglobin within red blood cells (RBC) has been studied, but the conduit whereby a SNO leaves the RBC remains unidentified. Here we hypothesised that SNO export by RBCs may also depend on LAT activity, and investigated the role of RBC LAT in modulating SNO-sensitive RBC-endothelial cell (EC) adhesion. We used multiple pharmacologic inhibitors of LAT in vitro and in vivo to test the role of LAT in SNO export from RBCs and in thereby modulating RBC-EC adhesion. Inhibition of human RBC LAT by type-1-specific or nonspecific LAT antagonists increased RBC-endothelial adhesivity in vitro, and LAT inhibitors tended to increase post-transfusion RBC sequestration in the lung and decreased oxygenation in vivo. A LAT1-specific inhibitor attenuated SNO export from RBCs, and we demonstrated LAT1 in RBC membranes and LAT1 mRNA in reticulocytes. The proadhesive effects of inhibiting LAT1 could be overcome by supplemental L-CSNO (S-nitroso-L-cysteine), but not D-CSNO or L-Cys, and suggest a basal anti-adhesive role for stereospecific intercellular SNO transport. This study reveals for the first time a novel role of LAT1 in the export of SNOs from RBCs to prevent their adhesion to ECs. The findings have implications for the mechanisms of intercellular SNO signalling, and for thrombosis, sickle cell disease, and post-storage RBC transfusion, when RBC adhesivity is increased.

scholarly journals Reconstruction of Sickle Cell Disease with Circulating Sickling Red Blood Cells in Novel Humanized Cytokines and Liver Mistrg Mice

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 29-30
Author(s):  
Yuanbin Song ◽  
Rana Gbyli ◽  
Liang Shan ◽  
Wei Liu ◽  
Yimeng Gao ◽  
...  
Keyword(s):  
Sickle Cell Disease ◽  
Mouse Model ◽  
Red Blood Cells ◽  
Sickle Cell ◽  
Blood Cells ◽  
Red Cell ◽  
Cell Disease ◽  
Ineffective Erythropoiesis ◽  
Cd34 Cells

In vivo models of human erythropoiesis with generation of circulating mature human red blood cells (huRBC) have remained elusive, limiting studies of primary human red cell disorders. In our prior study, we have generated the first combined cytokine-liver humanized immunodeficient mouse model (huHepMISTRG-Fah) with fully mature, circulating huRBC when engrafted with human CD34+ hematopoietic stem and progenitor cells (HSPCs)1. Here we present for the first time a humanized mouse model of human sickle cell disease (SCD) which replicates the hallmark pathophysiologic finding of vaso-occlusion in mice engrafted with primary patient-derived SCD HSPCs. SCD is an inherited blood disorder caused by a single point mutation in the beta-globin gene. Murine models of SCD exclusively express human globins in mouse red blood cells in the background of murine globin knockouts2 which exclusively contain murine erythropoiesis and red cells and thus fail to capture the heterogeneity encountered in patients. To determine whether enhanced erythropoiesis and most importantly circulating huRBC in engrafted huHepMISTRG-Fah mice would be sufficient to replicate the pathophysiology of SCD, we engrafted it with adult SCD BM CD34+ cells as well as age-matched control BM CD34+ cells. Overall huCD45+ and erythroid engraftment in BM (Fig. a, b) and PB (Fig. c, d) were similar between control or SCD. Using multispectral imaging flow cytometry, we observed sickling huRBCs (7-11 sickling huRBCs/ 100 huRBCs) in the PB of SCD (Fig. e) but not in control CD34+ (Fig. f) engrafted mice. To determine whether circulating huRBC would result in vaso-occlusion and associated findings in SCD engrafted huHepMISTRG-Fah mice, we evaluated histological sections of lung, liver, spleen, and kidney from control and SCD CD34+ engrafted mice. SCD CD34+ engrafted mice lungs showed an increase in alveolar macrophages (arrowheads) associated with alveolar hemorrhage and thrombosis (arrows) but not observed control engrafted mice (Fig. g). Spleens of SCD engrafted mice showed erythroid precursor expansion, sickled erythrocytes in the sinusoids (arrowheads), and vascular occlusion and thrombosis (arrows) (Fig. h). Liver architecture was disrupted in SCD engrafted mice with RBCs in sinusoids and microvascular thromboses (Fig. i). Congestion of capillary loops and peritubular capillaries and glomeruli engorged with sickled RBCs was evident in kidneys (Fig. j) of SCD but not control CD34+ engrafted mice. SCD is characterized by ineffective erythropoiesis due to structural abnormalities in erythroid precursors3. As a functional structural unit, erythroblastic islands (EBIs) represent a specialized niche for erythropoiesis, where a central macrophage is surrounded by developing erythroblasts of varying differentiation states4. In our study, both SCD (Fig. k) and control (Fig. l) CD34+ engrafted mice exhibited EBIs with huCD169+ huCD14+ central macrophages surrounded by varying stages of huCD235a+ erythroid progenitors, including enucleated huRBCs (arrows). This implies that huHepMISTRG-Fah mice have the capability to generate human EBIs in vivo and thus represent a valuable tool to not only study the effects of mature RBC but also to elucidate mechanisms of ineffective erythropoiesis in SCD and other red cell disorders. In conclusion, we successfully engrafted adult SCD patient BM derived CD34+ cells in huHepMISTRG-Fah mice and detected circulating, sickling huRBCs in the mouse PB. We observed pathological changes in the lung, spleen, liver and kidney, which are comparable to what is seen in the established SCD mouse models and in patients. In addition, huHepMISTRG-Fah mice offer the opportunity to study the role of the central macrophage in human erythropoiesis in health and disease in an immunologically advantageous context. This novel mouse model could therefore serve to open novel avenues for therapeutic advances in SCD. Reference 1. Song Y, Shan L, Gybli R, et. al. In Vivo reconstruction of Human Erythropoiesis with Circulating Mature Human RBCs in Humanized Liver Mistrg Mice. Blood. 2019;134:338. 2. Ryan TM, Ciavatta DJ, Townes TM. Knockout-transgenic mouse model of sickle cell disease. Science. 1997;278(5339):873-876. 3. Blouin MJ, De Paepe ME, Trudel M. Altered hematopoiesis in murine sickle cell disease. Blood. 1999;94(4):1451-1459. 4. Manwani D, Bieker JJ. The erythroblastic island. Curr Top Dev Biol. 2008;82:23-53. Disclosures Xu: Seattle Genetics: Membership on an entity's Board of Directors or advisory committees. Flavell:Zai labs: Consultancy; GSK: Consultancy.

Short survival of phosphatidylserine-exposing red blood cells in murine sickle cell anemia

Blood ◽  
2001 ◽  
Vol 98 (5) ◽  
pp. 1577-1584 ◽  
Author(s):  
Kitty de Jong ◽  
Renee K. Emerson ◽  
James Butler ◽  
Jacob Bastacky ◽  
Narla Mohandas ◽  
...  
Keyword(s):  
Red Blood Cells ◽  
Sickle Cell ◽  
Sickle Cell Anemia ◽  
Blood Cells ◽  
Cell Model ◽  
Red Cell ◽  
Sickle Cells ◽  
Red Cell Survival ◽  
Transgenic Murine Models

Several transgenic murine models for sickle cell anemia have been developed that closely reproduce the biochemical and physiological disorders in the human disease. A comprehensive characterization is described of hematologic parameters of mature red blood cells, reticulocytes, and red cell precursors in the bone marrow and spleen of a murine sickle cell model in which erythroid cells expressed exclusively human α, γ, and βS globin. Red cell survival was dramatically decreased in these anemic animals, partially compensated by considerable enhancement in erythropoietic activity. As in humans, these murine sickle cells contain a subpopulation of phosphatidylserine-exposing cells that may play a role in their premature removal. Continuous in vivo generation of this phosphatidylserine-exposing subset may have a significant impact on the pathophysiology of sickle cell disease.

scholarly journals Persistence of Chronic Inflammation Despite Years of Transfusion Program in SCD Patients: Changing Red Blood Cells Is Not Sufficient to Treat Sickle Cell Disease

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 764-764
Author(s):  
Abdoul Karim Dembele ◽  
Patricia Hermand-Tournamille ◽  
Florence Missud ◽  
Emmanuelle Lesprit ◽  
Malika Benkerrou ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Sickle Cell Disease ◽  
Red Blood Cells ◽  
Chronic Inflammation ◽  
Sickle Cell ◽  
Blood Cells ◽  
Healthy Controls ◽  
Cell Disease ◽  
The Impact

Abstract Sickle cell disease (SCD) is a severe hemoglobinopathy due to abnormal hemoglobin S (HbS). Although red blood cell dysfunction is at the core of the SCD pathophysiology, several studies have highlighted the important role of inflammatory cells like neutrophils. One of the most serious complications of SCD is cerebral vasculopathy (CV), due to the occlusion of one or more intracranial or cervical arteries. In 1998, the STOP study demonstrated that monthly blood transfusions could reduce the risk of stroke by 90% in children with CV. However, there is large heterogeneity in the evolution of CV under chronic transfusion, sometimes requiring exchange transfusion (ET) program for years without succeeding in healing the CV. The aim of the study is to investigate the impact of long-term transfusion program on neutrophil dysfunction, in order to understand if persistent inflammation could contribute to the non-healing of CV despite HbS permanently below 40%. In SCD children undergoing ET program for at least 1 year, we analysed i)the phenotype of neutrophils with 8 markers of activation/adhesion/ageing, ii)the plasmatic levels of elastase, witnessing the NETose activity of neutrophils, and iii)the ex-vivo adhesion of neutrophils on activated endothelial cells. One hundred and two SCD children with an ET transfusion program for at least 6 months because of CV were included in the study. ET session, carried out every 5 weeks and most of the time by erythrapheresis, reached their biological objectives with a mean HbS rate after ET session of 14.1%, and 35.4% before the next ET session, which means that these patients globally live at an average HbS level of 24% for at least 1 year. We managed to limit iron overload with a mean ferritinemia of 207 µg/L in the whole cohort. Despite these satisfactory results in terms of HbS reduction, the efficiency in curing the CV was modest in accordance with the previously described efficiency of ET program in SCD children: after a mean ET program duration of 4.4 years only 22% of them had an improvement of their CV since the beginning of the ET program, while 60% of them had a stagnation of their CV, and 18% of them worsened their vascular lesions. Considering inflammatory parameters, the patients had persistence of high leukocytosis and high neutrophils count (respective mean of 9810 G/L and 5742 G/L), significantly not different of neutrophils count before inclusion in the ET program. In a random subgroup of 20 patients, we analysed neutrophils phenotype, NETose and endothelial adhesion and compared them to healthy controls and SCD children without ET, treated or not with Hydroxyurea (HU). Overall, we observed as expected an activated, aged and adherent profile of neutrophils from untreated SCD children compared to healthy controls, characterized by an overexpression of CD18/CD11b (p=0,03), CD18/CD11a (p=0,02), CD162 (p=0,01), CD66a (p=0,01) and the ageing markers CD184 high/CD62Llow (p=0,04) as well as a higher plasmatic level of elastase (p=0. 01) and higher adhesion of neutrophils to endothelial cells. All these parameters were alleviated in SCD patients treated with HU. In SCD patient undergoing ET program, we found a similar profile of activated neutrophils to that of untreated SCD patients with a similar expression of activation molecules, high level of elastase and the same increase of neutrophils adhesion to endothelial cells compared to controls, witnessing a persistence of chronic inflammation despites years of ET. Overall, our study highlights that the replacement of sickle red blood cells, even for years, is not sufficient to reverse the deleterious inflammatory phenotype of neutrophils. Given the major role of inflammation in endothelial dysfunction, these could contribute to the persistence of CV in a majority of patients despite efficient ET programs. This raises the question of systematically combining ET program with anti-inflammatory treatment such as HU or P-selectin inhibitors in children with CV. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals Evolving treatment paradigms in sickle cell disease

Hematology ◽  
2017 ◽  
Vol 2017 (1) ◽  
pp. 440-446 ◽  
Author(s):  
Ramasamy Jagadeeswaran ◽  
Angela Rivers
Keyword(s):  
Sickle Cell Disease ◽  
Red Blood Cells ◽  
Sickle Cell ◽  
Blood Cells ◽  
Cellular Adhesion ◽  
Ros Generation ◽  
Cell Disease ◽  
Therapeutic Approaches ◽  
Chronic Hemolytic Anemia

AbstractSickle cell disease (SCD) is an inheritable hemoglobinopathy characterized by polymerization of hemoglobin S in red blood cells resulting in chronic hemolytic anemia, vaso-occlusive painful crisis, and multiorgan damage. In SCD, an increased reactive oxygen species (ROS) generation occurs both inside the red blood cells and inside the vascular lumen, which augment hemolysis and cellular adhesion. This review discusses the evolving body of literature on the role of ROS in the pathophysiology of SCD as well as some emerging therapeutic approaches to SCD with a focus on the reduction of ROS.

The Role of Lactadherin in the Phagocytosis of Phosphatidylserine-Expressing Sickle Red Blood Cell by Macrophages.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3773-3773
Author(s):  
Swapan K. Dasgupta ◽  
Perumal Thiagarajan
Keyword(s):  
Flow Cytometry ◽  
Red Blood Cells ◽  
Sickle Cell ◽  
Blood Cells ◽  
Milk Fat ◽  
Integrin Αvβ3 ◽  
Fold Increase ◽  
Anionic Phospholipid ◽  
Sickle Red Blood Cells

Abstract Sickle cell anemia, the most common serious hemoglobinopathy, is associated with a markedly reduced life span of red blood cells due to their preferential clearance by macrophages. During polymerization of sickle hemoglobin, phosphatidylserine, an anionic phospholipid normally present exclusively on the inner leaflet of the membrane bilayer is exteriorized to outer leaflet. This exposure of phosphatidylserine is thought to be a tag for macrophage recognition. Lactadherin, also known as milk fat globule-EGF factor 8, is a phosphatidylserine-binding glycoprotein secreted by macrophages that promotes the engulfment of apoptotic cells. Here, we investigated the role of lactadherin in the phagocytosis of sickle red blood cells. The binding of fluorescein-lactadherin to normal and sickle red blood cells was studied by flow cytometry. We quantified the effect of lactadherin on phagocytosis of red blood cells by monocyte-derived macrophage. In normal individuals, less than 0.5% of red blood cells showed any binding to lactadherin when analyzed by flow cytometry. However, in sickle cell patients, circulating red blood cells showed 2 to 10- fold increase in lactadherin binding (P<0.0002). Lactadherin stimulated the phagocytosis of resting sickle red blood cells by macrophages but had no significant effect on the phagocytosis of normal red blood cells. Deoxygenation of sickle red blood cells further increased the lactadherin binding and phagocytosis. Antibodies to integrin αVβ3 also inhibited macrophage binding and phagocytosis. These results show lactadherin may play a major role in sickle red cell clearance by anchoring the phosphatidylserine-expressing sickle red blood cells to integrins on tissue macrophages.

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