scholarly journals Yoda1 and phosphatidylserine exposure in red cells from patients with sickle cell anaemia

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
R. Wadud ◽  
A. Hannemann ◽  
D. C. Rees ◽  
J. N. Brewin ◽  
J. S. Gibson
Keyword(s):  
Sickle Cell ◽  
Sickle Cell Anaemia ◽  
Red Cells ◽  
Mechanosensitive Channel ◽  
Sickle Cells ◽  
Molecular Identity ◽  
Calphostin C ◽  
Intracellular Ca ◽  
Pkc Inhibitors ◽  
Low Levels

AbstractPhosphatidylserine (PS) exposure is increased in red cells from sickle cell anaemia (SCA) patients. Externalised PS is prothrombotic and attractive to phagocytes and activated endothelial cells and thus contributes to the anaemic and ischaemic complications of SCA. The mechanism of PS exposure remains uncertain but it can follow increased intracellular Ca2+ concentration ([Ca2+]i). Normally, [Ca2+]i is maintained at very low levels but in sickle cells, Ca2+ permeability is increased, especially following deoxygenation and sickling, mediated by a pathway sometimes called Psickle. The molecular identity of Psickle is also unclear but recent work has implicated the mechanosensitive channel, PIEZO1. We used Yoda1, an PIEZO1 agonist, to investigate its role in sickle cells. Yoda1 caused an increase in [Ca2+]i and PS exposure, which was inhibited by its antagonist Dooku1 and the PIEZO1 inhibitor GsMTx4, consistent with functional PIEZO1. However, PS exposure did not necessitate an increase in [Ca2+]i. Two PKC inhibitors were also tested, chelerytherine chloride and calphostin C. Both reduced PS exposure whilst chelerytherine chloride also reduced Yoda1-induced increases in [Ca2+]i. Findings are therefore consistent with the presence of PIEZO1 in sickle cells, able to mediate Ca2+ entry but that PKC was also involved in both Ca2+ entry and PS exposure.

scholarly journals Pathophysiological Relevance of Renal Medullary Conditions on the Behaviour of Red Cells From Patients With Sickle Cell Anaemia

2021 ◽  
Vol 12 ◽  
Author(s):  
David C.-Y. Lu ◽  
Rasiqh Wadud ◽  
Anke Hannemann ◽  
David C. Rees ◽  
John N. Brewin ◽  
...  
Keyword(s):  
Sickle Cell ◽  
Sickle Cell Anaemia ◽  
Renal Medulla ◽  
Red Cells ◽  
Red Cell ◽  
Sickle Cells ◽  
Hypoxic Conditions ◽  
Intracellular Ca ◽  
Concentrate Urine ◽  
Abnormal Haemoglobin

Red cells from patients with sickle cell anaemia (SCA) contain the abnormal haemoglobin HbS. Under hypoxic conditions, HbS polymerises and causes red cell sickling, a rise in intracellular Ca2+ and exposure of phosphatidylserine (PS). These changes make sickle cells sticky and liable to lodge in the microvasculature, and so reduce their lifespan. The aim of the present work was to investigate how the peculiar conditions found in the renal medulla – hypoxia, acidosis, lactate, hypertonicity and high levels of urea – affect red cell behaviour. Results show that the first four conditions all increased sickling and PS exposure. The presence of urea at levels found in a healthy medulla during antidiuresis, however, markedly reduced sickling and PS exposure and would therefore protect against red cell adherence. Loss of the ability to concentrate urine, which occurs in sickle cell nephropathy would obviate this protective effect and may therefore contribute to pathogenesis.

scholarly journals The role of WNK in modulation of KCl cotransport activity in red cells from normal individuals and patients with sickle cell anaemia

2019 ◽  
Vol 471 (11-12) ◽  
pp. 1539-1549
Author(s):  
David C.-Y. Lu ◽  
Anke Hannemann ◽  
Rasiqh Wadud ◽  
David C. Rees ◽  
John N. Brewin ◽  
...  
Keyword(s):  
Oxygen Tension ◽  
Sickle Cell ◽  
Sickle Cell Anaemia ◽  
Kinase Inhibitors ◽  
Red Cells ◽  
Red Cell ◽  
Sickle Cells ◽  
Kcl Cotransport ◽  
Pre Treatment

AbstractAbnormal activity of red cell KCl cotransport (KCC) is involved in pathogenesis of sickle cell anaemia (SCA). KCC-mediated solute loss causes shrinkage, concentrates HbS, and promotes HbS polymerisation. Red cell KCC also responds to various stimuli including pH, volume, urea, and oxygen tension, and regulation involves protein phosphorylation. The main aim of this study was to investigate the role of the WNK/SPAK/OSR1 pathway in sickle cells. The pan WNK inhibitor WNK463 stimulated KCC with an EC50 of 10.9 ± 1.1 nM and 7.9 ± 1.2 nM in sickle and normal red cells, respectively. SPAK/OSR1 inhibitors had little effect. The action of WNK463 was not additive with other kinase inhibitors (staurosporine and N-ethylmaleimide). Its effects were largely abrogated by pre-treatment with the phosphatase inhibitor calyculin A. WNK463 also reduced the effects of physiological KCC stimuli (pH, volume, urea) and abolished any response of KCC to changes in oxygen tension. Finally, although protein kinases have been implicated in regulation of phosphatidylserine exposure, WNK463 had no effect. Findings indicate a predominant role for WNKs in control of KCC in sickle cells but an apparent absence of downstream involvement of SPAK/OSR1. A more complete understanding of the mechanisms will inform pathogenesis whilst manipulation of WNK activity represents a potential therapeutic approach.

scholarly journals Vaso-occlusion by sickle cells: evidence for selective trapping of dense red cells

Blood ◽  
1986 ◽  
Vol 68 (5) ◽  
pp. 1162-1166 ◽  
Author(s):  
DK Kaul ◽  
ME Fabry ◽  
RL Nagel
Keyword(s):  
Density Gradient ◽  
Sickle Cell ◽  
Morphological Analysis ◽  
Microvascular Obstruction ◽  
Perfusion Pressure ◽  
Peripheral Resistance ◽  
Red Cells ◽  
Arterial Perfusion ◽  
Sickle Cells ◽  
Sickle Cell Crisis

We have characterized the type of red cells from sickle cell patients that were trapped in the course of sickle-cell vaso-occlusion. In addition, the perfusion conditions (arterial perfusion pressure [Pa] and oxygen tension [PO2]) leading to experimentally induced vaso- occlusion in the artificially perfused, innervated mesocecum microvascular preparation were determined. Microvascular obstruction was induced by decrease in Pa; the lower the Pa, the greater the peripheral resistance as well as the extent of obstruction. The cells involved in the obstruction were recovered by vasodilation (secondary to denervation) and increase in Pa. Densitometric analysis of density gradient-separated infused and trapped cells was supplemented with morphological analysis to ascertain the involvement of density classes as well as morphological types seen in oxy and deoxy sickle blood. The trapping phenomenon was sensitive to PO2. Percentage of densest gradient classes, ie, fraction 3 (F3; mainly dense unsicklable SS discocytes [USDs]) and fraction 4 (F4; irreversibly sickled cells [ISCs] and the densest discocytes), showed a significant increase in trapping when perfusion was switched from oxy to deoxy perfusate. Morphological analysis revealed that unsicklable SS discocytes are more effectively trapped when deoxygenated. The deoxygenation of infused cells did not further change the percentage of ISCs trapped, suggesting that ISCs are equally capable of sequestration in the oxy and the deoxy states. The venous effluent showed a selective and significant depletion of dense cells (F4) and ISC counts at all Pa. We conclude that the progressive obstruction of the microcirculation by sickle cells involves selective sequestration of the densest classes of cells and that this mechanism might explain their partial disappearance during painful sickle cell crisis.

scholarly journals Hematological Indices of Sickle Cell Anaemia Patients with Pulmonary Tuberculosis in Northern Nigeria.

2010 ◽  
Vol 2 (1) ◽  
pp. e2010014 ◽  
Author(s):  
Sagir G. Ahmed ◽  
Audu A. Bukar ◽  
Bashir Jolayemi
Keyword(s):  
Sickle Cell ◽  
Sickle Cell Anaemia ◽  
Reticulocyte Count ◽  
Hematological Indices ◽  
Sickle Cells ◽  
The World ◽  
Patient Groups ◽  
Wbc Count ◽  
Haematological Indices ◽  
Age And Sex

Nigeria has the fourth highest prevalence of TB and the highest prevalence of Sickle cell anaemia (SCA) in the world. SCA patients have impaired immunity and are vulnerable to TB. Hence, we studied the haematological indices of SCA patients with TB in Nigeria. A total of 23 SCA patients with TB were studied in parallel with equal number of age and sex matched SCA patients without TB. SCA patients with TB had significantly lower haematocrit, higher level of circulating sickle cells (CSCs) and similar level of reticulocyte count in comparison to patients without TB. SCA patients with TB had significantly higher mean WBC count associated with higher frequency of neutrophilia in comparison to those without TB. Monocytosis and eosinopenia were exclusively found in SCA patients with TB at frequencies of 52% and 65% respectively. Lymphocyte and basophil counts were normal in all patients with and without TB. Mean platelet counts were high in both patient groups but the frequency of thrombocytosis was significantly higher in patients with TB. SCA patients with TB had significantly higher mean ESR than their counterparts without the infection. The findings of this study  revealed that TB in SCA patients was associated with rising level of CSCs, falling level of haematocrit, sub-optimal reticulocytosis, neutrophilia, monocytosis, thrombocytosis, eosinopenia and rising level of ESR. Hence, SCA patients presenting with these haematological indices should be investigated for TB.

scholarly journals Vaso-occlusion by sickle cells: evidence for selective trapping of dense red cells

Blood ◽  
1986 ◽  
Vol 68 (5) ◽  
pp. 1162-1166 ◽  
Author(s):  
DK Kaul ◽  
ME Fabry ◽  
RL Nagel
Keyword(s):  
Density Gradient ◽  
Sickle Cell ◽  
Morphological Analysis ◽  
Microvascular Obstruction ◽  
Perfusion Pressure ◽  
Peripheral Resistance ◽  
Red Cells ◽  
Arterial Perfusion ◽  
Sickle Cells ◽  
Sickle Cell Crisis

Abstract We have characterized the type of red cells from sickle cell patients that were trapped in the course of sickle-cell vaso-occlusion. In addition, the perfusion conditions (arterial perfusion pressure [Pa] and oxygen tension [PO2]) leading to experimentally induced vaso- occlusion in the artificially perfused, innervated mesocecum microvascular preparation were determined. Microvascular obstruction was induced by decrease in Pa; the lower the Pa, the greater the peripheral resistance as well as the extent of obstruction. The cells involved in the obstruction were recovered by vasodilation (secondary to denervation) and increase in Pa. Densitometric analysis of density gradient-separated infused and trapped cells was supplemented with morphological analysis to ascertain the involvement of density classes as well as morphological types seen in oxy and deoxy sickle blood. The trapping phenomenon was sensitive to PO2. Percentage of densest gradient classes, ie, fraction 3 (F3; mainly dense unsicklable SS discocytes [USDs]) and fraction 4 (F4; irreversibly sickled cells [ISCs] and the densest discocytes), showed a significant increase in trapping when perfusion was switched from oxy to deoxy perfusate. Morphological analysis revealed that unsicklable SS discocytes are more effectively trapped when deoxygenated. The deoxygenation of infused cells did not further change the percentage of ISCs trapped, suggesting that ISCs are equally capable of sequestration in the oxy and the deoxy states. The venous effluent showed a selective and significant depletion of dense cells (F4) and ISC counts at all Pa. We conclude that the progressive obstruction of the microcirculation by sickle cells involves selective sequestration of the densest classes of cells and that this mechanism might explain their partial disappearance during painful sickle cell crisis.

High Calcium Requirement for Phosphatidylserine Exposure in Sickle Cell Disease.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3788-3788 ◽  
Author(s):  
Kitty DeJong ◽  
Annabeth M. Miller ◽  
Renee K. Emerson ◽  
Frans A. Kuypers
Keyword(s):  
Sickle Cell ◽  
Annexin V ◽  
Membrane Bilayer ◽  
Sickle Cells ◽  
Calcium Requirement ◽  
Entire Duration ◽  
High Calcium ◽  
Intracellular Ca ◽  
Sulfhydryl Modification ◽  
Phospholipid Scrambling

Abstract While the adverse effects of red blood cells (RBC) that expose phosphatidylserine (PS) in the sickle cell circulation are well recognized, the mechanism that underlies the formation of these cells is not well understood. Diminished flippase activity is found in a subset of sickle cells, and while this explains how PS persists in the outer RBC monolayer, it does not explain how the phospholipids are randomized across the membrane bilayer. Rapid scrambling of phospholipids, resulting in PS exposure, requires elevation of intracellular free Ca++ levels by Ca++ influx into the RBC. In this study we assessed how much Ca++ is required for PS exposure, and how this relates to previous findings of elevated Ca++ influx during deoxygenation of sickle cells. We loaded RBC for 30 minutes at 37°C with accurately established levels of intracellular Ca++ by balancing CaCl2 solutions with the chelator BAPTA, followed by Annexin V labeling under conditions that scrambling did not continue. In both normal and sickle RBC from either human or mouse, PS exposure was only found at or above 10 microM Ca++. Flippase inhibition with vanadate did not change this, indicating that phospholipid scrambling had not been initiated under these conditions. Scramblase sulfhydryl modification with NEM reduced the Ca++ requirement to lower than 1 microM, as did treatment with the scramblase stimulator oligomycin. Further experiments showed that Ca++ needs to be present during the entire duration of the scrambling process, and removal of Ca++ arrests the scrambling process. With 1 microM Ca++ in oligomycin-treated (scramblase stimulated) RBC and in absence of flippase activity (using vanadate), it took 5 minutes at 37°C to obtain a significant (>2%) population of PS-exposing cells. These conditions are unlikely met in all sickle cells. In addition, these experiments showed that the flippase is not inhibited by 1 or 100 microM Ca++, despite previous reports suggesting that Ca++ loading inhibits the flippase. Taken together, our data indicate that PS scrambling requires more Ca++ influx than has been reported for deoxygenated sickle cells (50–100 nM). We suggest that a small percentage of individual sickle cells contain Ca++ levels of 1 microM or above, which would lead to PS exposure in those RBC that have sustained flippase inhibition and scramblase modifications similar to those achieved with NEM or oligomycin. We have previously reported evidence for such scramblase modifications, as well as reduced flippase activity in a subset of sickle cells. Our data indicate that the degree of Ca++ influx may be the determining parameter for the extent of PS exposure in sickle cells.

scholarly journals The percentage of dense red cells does not predict incidence of sickle cell painful crisis

Blood ◽  
1986 ◽  
Vol 68 (1) ◽  
pp. 301-303 ◽  
Author(s):  
HH Billett ◽  
K Kim ◽  
ME Fabry ◽  
RL Nagel
Keyword(s):  
Sickle Cell ◽  
Gradient Centrifugation ◽  
Red Cells ◽  
Small Subset ◽  
Sickle Cells ◽  
Alpha Thalassemia ◽  
Cellular Factors ◽  
Alpha Gene ◽  
Gene Status ◽  
Painful Crisis

Abstract To test the hypothesis that the tendency of hemoglobin S (HbS) to polymerize within cells is the major determinant of the incidence of vaso-occlusive episodes, we have examined the effect of the percentage of dense cells (as measured by Percoll-Stractan continuous density gradient centrifugation) on the frequency of painful crises in a group of 36 patients with sickle cell disease. No correlation was found between the percentage of dense cells and admissions for crisis. Among the patients with known alpha-gene status (n = 25), the strong correlation between decreased dense cells and alpha-thalassemia (- alpha/alpha alpha) reported previously was confirmed (P less than .001). In addition, in this small subset, patients with alpha- thalassemia (-alpha/alpha alpha) appeared to have a marginally increased number of admissions for sickle cell crisis (t = 2.1910, P less than .05), which was independent of the percentage of dense cells. We conclude that the percentage of dense sickle cells cannot predict the incidence of painful crisis, suggesting that other factors (microcirculatory regulation or other humoral and cellular factors) are more important in the generation maintenance of painful crises than the necessary, but not sufficient, tendency of HbS-containing red cells to sickle.

scholarly journals The percentage of dense red cells does not predict incidence of sickle cell painful crisis

Blood ◽  
1986 ◽  
Vol 68 (1) ◽  
pp. 301-303 ◽  
Author(s):  
HH Billett ◽  
K Kim ◽  
ME Fabry ◽  
RL Nagel
Keyword(s):  
Sickle Cell ◽  
Gradient Centrifugation ◽  
Red Cells ◽  
Small Subset ◽  
Sickle Cells ◽  
Alpha Thalassemia ◽  
Cellular Factors ◽  
Alpha Gene ◽  
Gene Status ◽  
Painful Crisis

To test the hypothesis that the tendency of hemoglobin S (HbS) to polymerize within cells is the major determinant of the incidence of vaso-occlusive episodes, we have examined the effect of the percentage of dense cells (as measured by Percoll-Stractan continuous density gradient centrifugation) on the frequency of painful crises in a group of 36 patients with sickle cell disease. No correlation was found between the percentage of dense cells and admissions for crisis. Among the patients with known alpha-gene status (n = 25), the strong correlation between decreased dense cells and alpha-thalassemia (- alpha/alpha alpha) reported previously was confirmed (P less than .001). In addition, in this small subset, patients with alpha- thalassemia (-alpha/alpha alpha) appeared to have a marginally increased number of admissions for sickle cell crisis (t = 2.1910, P less than .05), which was independent of the percentage of dense cells. We conclude that the percentage of dense sickle cells cannot predict the incidence of painful crisis, suggesting that other factors (microcirculatory regulation or other humoral and cellular factors) are more important in the generation maintenance of painful crises than the necessary, but not sufficient, tendency of HbS-containing red cells to sickle.

scholarly journals Role of Emergency Automated Red Cell Exchange in Sickle Cell Crisis: A Case Report

2020 ◽  
Vol 13 ◽  
pp. 117954762097020
Author(s):  
Anubhav Gupta ◽  
Kiran Chaudhary ◽  
Rajnish Kaushik
Keyword(s):  
Sickle Cell ◽  
Sickle Cell Anaemia ◽  
Symptomatic Improvement ◽  
Red Cells ◽  
Acute Chest Syndrome ◽  
Compound Heterozygous ◽  
Red Cell ◽  
Transfusion Therapy ◽  
Sickle Cell Crisis ◽  
Patient At Risk

For many years main stay of treatment for sickle cell anaemia was transfusion therapy. But repeated transfusions put the patient at risk of iron overload. Automated red cell exchange is an evolving and newer technique which rapidly removes the sickle cells and has benefit in decreasing sickle cell load and related complications. Red cell exchange is a therapeutic procedure in which the patient’s whole blood is processed centrifugally in cell separator. Patient’s red cells are separated from other blood components and removed and replaced with donor red cells and colloids. We report our first experience of automated red cell exchange in 24-year-old female diagnosed case of sickle cell anaemia presented to us with acute chest syndrome with septic shock. Red cell exchange was planned to tide over the acute sickle cell crisis and provide symptomatic improvement. We also highlight that compound heterozygous thalassaemia could be associated with sickle cell disease which could make the diagnosis difficult. New generation automated Apheresis equipment’s provides better monitoring of the procedure that can be useful in severely ill patients also.

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