Analysis of Hereditary Elliptocytosis with Decreased Binding of Eosin-5-maleimide to Red Blood Cells

Flow cytometric test for analyzing the eosin-5-maleimide (EMA) binding to red blood cells has been believed to be a specific method for diagnosing hereditary spherocytosis (HS). However, it has been reported that diseases other than HS, such as hereditary pyropoikilocytosis (HPP) and Southeast Asian ovalocytosis (SAO), which are forms in the category of hereditary elliptocytosis (HE), show decreased EMA binding to red blood cells. We analyzed EMA binding to red blood cells in 101 healthy control subjects and 42 HS patients and obtained a mean channel fluorescence (MCF) cut-off value of 36.4 (sensitivity 0.97, specificity 0.95). Using this method, we also analyzed 12 HE patients. Among them, four HE patients showed the MCF at or below the cut-off value. It indicates that some HE patients have decreased EMA binding to red blood cells. Two of these four HE patients were classified as common HE, and two were spherocytic HE with reduced spectrin. This study demonstrates that, in addition to patients with HPP or SAO, some HE patients have decreased EMA binding to red blood cells.

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Silent hypoxia: higher NO in red blood cells of COVID-19 patients

BMC Pulmonary Medicine ◽

10.1186/s12890-020-01310-8 ◽

2020 ◽

Vol 20 (1) ◽

Cited By ~ 1

Author(s):

Esmaeil Mortaz ◽

Majid Malkmohammad ◽

Hamidreza Jamaati ◽

Parisa Adimi Naghan ◽

Seyed MohamadReza Hashemian ◽

...

Keyword(s):

Red Blood Cells ◽

Blood Cells ◽

Diffuse Alveolar Damage ◽

Serum Levels ◽

Pulmonary Involvement ◽

Clinical Feature ◽

Control Group ◽

Small Vessels ◽

Control Subjects ◽

Healthy Control

Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes coronavirus disease 2019 (COVID-19) has spread to almost 100 countries, infected over 31 M patients and resulted in 961 K deaths worldwide as of 21st September 2020. The major clinical feature of severe COVID-19 requiring ventilation is acute respiratory distress syndrome (ARDS) with multi-functional failure as a result of a cytokine storm with increased serum levels of cytokines. The pathogenesis of the respiratory failure in COVID-19 is yet unknown, but diffuse alveolar damage with interstitial thickening leading to compromised gas exchange is a plausible mechanism. Hypoxia is seen in the COVID-19 patients, however, patients present with a distinct phenotype. Intracellular levels of nitric oxide (NO) play an important role in the vasodilation of small vessels. To elucidate the intracellular levels of NO inside of RBCs in COVID-19 patients compared with that of healthy control subjects. Methods We recruited 14 COVID-19 infected cases who had pulmonary involvement of their disease, 4 non-COVID-19 healthy controls (without pulmonary involvement and were not hypoxic) and 2 hypoxic non-COVID-19 patients subjects who presented at the Masih Daneshvari Hospital of Tehran, Iran between March–May 2020. Whole blood samples were harvested from patients and intracellular NO levels in 1 × 106 red blood cells (RBC) was measured by DAF staining using flow cytometry (FACS Calibour, BD, CA, USA). Results The Mean florescent of intensity for NO was significantly enhanced in COVID-19 patients compared with healthy control subjects (P ≤ 0.05). As a further control for whether hypoxia induced this higher intracellular NO, we evaluated the levels of NO inside RBC of hypoxic patients. No significant differences in NO levels were seen between the hypoxic and non-hypoxic control group. Conclusions This pilot study demonstrates increased levels of intracellular NO in RBCs from COVID-19 patients. Future multi-centre studies should examine whether this is seen in a larger number of COVID-19 patients and whether NO therapy may be of use in these severe COVID-19 patients.

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Evaluation of a Flow-Cytometric Osmotic Fragility Test for Hereditary Spherocytosis in Chinese Patients

Acta Haematologica ◽

10.1159/000438738 ◽

2015 ◽

Vol 135 (2) ◽

pp. 88-93 ◽

Cited By ~ 5

Author(s):

Yi-Feng Tao ◽

Zeng-Fu Deng ◽

Lin Liao ◽

Yu-Ling Qiu ◽

Xue-Lian Deng ◽

...

Keyword(s):

Blood Cell ◽

Red Blood Cells ◽

Cell Morphology ◽

Red Blood Cell ◽

Peripheral Blood ◽

Blood Cells ◽

Hereditary Spherocytosis ◽

Osmotic Fragility ◽

Flow Cytometric ◽

Blood Cell Morphology

Background: Osmotic fragility testing based on flow cytometry was recently introduced for the screening of hereditary spherocytosis (HS). This study was undertaken to evaluate the clinical diagnostic value of a flow-cytometric osmotic fragility test for HS. Methods: Peripheral blood was collected from 237 subjects at the First Affiliated Hospital of Guangxi Medical University, including 56 HS patients, 86 thalassemia patients and 95 healthy controls. The samples were examined by flow-cytometric osmotic fragility test and the percentage of residual red blood cells was used to determine HS. Peripheral blood smears were performed to examine the red blood cell morphology. Results: With clinical diagnosis of HS as the gold standard and the percentage of residual red blood cells <23.6% as the diagnostic threshold in the flow-cytometric osmotic fragility test, the sensitivity of the flow-cytometric osmotic fragility test for HS was 85.71% and the specificity was 97.24%. Conclusion: The flow-cytometric osmotic fragility test combined with a red blood cell morphology test by peripheral blood smear could be a simple, practical and accurate laboratory screening method for HS.

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Happy Hypoxia: Higher NO in red blood cells of COVID-19 patients

10.21203/rs.3.rs-49770/v1 ◽

2020 ◽

Author(s):

Esmaeil Moratz ◽

Majid Malekmohammad ◽

Hamidreza Jamaati ◽

Parisa Adimi Naghan ◽

Mohammadreza Hashemian ◽

...

Keyword(s):

Red Blood Cells ◽

Blood Cells ◽

Diffuse Alveolar Damage ◽

Serum Levels ◽

Pulmonary Involvement ◽

Clinical Feature ◽

Control Group ◽

Small Vessels ◽

Control Subjects ◽

Healthy Control

Abstract Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes coronavirus disease 2019 (COVID-19) has spread to almost 100 countries, infected over 10M patients and resulted in 505K deaths worldwide as of 30th June 2020. The major clinical feature of severe COVID-19 requiring ventilation is acute Respiratory Distress Syndrome (ARDS) with multi-functional failure as a result of a cytokine storm with increased serum levels of cytokines. The pathogenesis of the respiratory failure in COVID-19 is yet unknown, but diffuse alveolar damage with interstitial thickening leading to compromised gas exchange is a plausible mechanism. Hypoxia has been seen in the COVID-19 patients however, patients present with a distinct phenotype. Intracellular levels of NO playing important role in the vasodilation of small vessels.Objectives: To elucidate the intracellular levels of NO inside of RBCs in COVID-19 patients compared with that of healthy control subjects.Methods: We recruited 14 COVID-19 infected cases who had pulmonary involvement of their disease, 4 non-COVID-19 healthy controls (without pulmonary involvement and were not hypoxic) and 2 hypoxic non-COVID-19 patients subjects who presented at the Masih Daneshvari Hospital of Tehran, Iran between March-May 2020. Whole blood samples were harvested from patients and intracellular levels of NO in 1 million red blood cells (RBC) was measured by DAF staining using flow cytometry (FACS Calibour, BD, CA, USA).Results: The Mean florescent of intensity for NO was significantly enhanced in COVID-19 patients compared with healthy control subjects (P≤0.05). As a further control for whether hypoxia induced this higher intracellular NO, we evaluated the levels of NO inside RBC of hypoxic patients. No significant differences in NO levels were seen between the hypoxic and non-hypoxic control group.Conclusions: This pilot study demonstrates increased levels of intracellular NO in RBCs from COVID-19 patients. Future studies should examine whether intracellular NO would be increased in large number of COVID-19 patients for usage of possible NO therapy in severe patients.

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Impact of storage, leukofiltration, and ascorbic acid fortification on red cell-derived microparticles in stored packed red blood cells: A flow cytometric and procoagulant study

Journal of Applied Hematology ◽

10.4103/joah.joah_76_19 ◽

2020 ◽

Vol 11 (2) ◽

pp. 51

Author(s):

Raghda Fouda ◽

AzzaA Aboul Enein ◽

NermeenA El-Desoukey ◽

RandaM Abo Elfetouh ◽

AhmedM. A Abdel Hafez

Keyword(s):

Ascorbic Acid ◽

Red Blood Cells ◽

Blood Cells ◽

Red Cell ◽

Packed Red Blood Cells ◽

Flow Cytometric

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Fine Structure of the Red Pulp of the Spleen in Hereditary Spherocytosis

Blood ◽

10.1182/blood.v39.1.81.81 ◽

1972 ◽

Vol 39 (1) ◽

pp. 81-98 ◽

Cited By ~ 28

Author(s):

ZELMA MOLNAR ◽

HENRY RAPPAPORT

Keyword(s):

Endothelial Cells ◽

Red Blood Cells ◽

Blood Cells ◽

Hereditary Spherocytosis ◽

Probable Mechanism ◽

Hemoglobin Content ◽

Sinus Endothelial Cells ◽

In Transit ◽

Conventional Light Microscopy ◽

Red Pulp

Abstract The spleens from two children and one adult with hereditary spherocytosis were studied in the electron microscope. Stagnation of the erythrocytes within the splenic cords is attributable to their lack of plasticity as evidenced by the absence of bilobed, tailed, or squeezed forms in transit through the walls of the sinuses. In contrast to the sections studied by conventional light microscopy, the splenic sinuses in hereditary spherocytosis were not "empty," but contained red blood cells, the majority of which had lost their hemoglobin content. Cordal macrophages were increased in all three cases and were abundant in the splenic cords of the adult patient, causing a further impediment to the rapid passage of erythrocytes. Macrophages, and, to a lesser degree, sinus endothelial cells contained the products of hemoglobin breakdown. The macrophages showed active erythrophagocytosis. Sinus endothelial cells rarely contained intact red blood cells, but showed pronounced pinocytotic activity, a probable mechanism of hemoglobin incorporation. Platelets within the endothelial cells of the sinuses were much more frequently seen in the three cases of hereditary spherocytosis than in control spleens. The presence of ferritin in platelets suggests that they too may play a role in clearing the end products of hemolysis from the spleen.

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Ability of Plasmodium falciparum to invade Southeast Asian ovalocytes varies between parasite lines

Blood ◽

10.1182/blood-2004-06-2136 ◽

2004 ◽

Vol 104 (9) ◽

pp. 2961-2966 ◽

Cited By ~ 44

Author(s):

Alfred Cortés ◽

Ariadna Benet ◽

Brian M. Cooke ◽

John W. Barnwell ◽

John C. Reeder

Keyword(s):

Plasmodium Falciparum ◽

Red Blood Cells ◽

Blood Cells ◽

Clinical Observation ◽

Southeast Asian ◽

Parasite Line ◽

Invasion Pathways ◽

Receptor Interactions ◽

Multiple Ligand

Abstract Plasmodium falciparum, the causative agent of the most lethal form of human malaria, uses multiple ligand-receptor interactions to invade host red blood cells (RBCs). We studied the invasion of P falciparum into abnormal RBCs from humans carrying the Southeast Asian ovalocytosis (SAO) trait. One particular parasite line, 3D7-A, invaded these cells efficiently, whereas all other lines studied invaded SAO RBCs to only about 20% of the extent of normal (non-SAO) cells. This result is consistent with the clinical observation that SAO individuals can experience high-density P falciparum infections and provides an explanation for previous discrepant results on invasion of SAO RBCs. Characterization of the invasion phenotype of 3D7-A revealed that efficient invasion of SAO RBCs was paralleled by relatively efficient invasion of normal RBCs treated with either neuraminidase, trypsin, or chymotrypsin and a novel capacity to invade normal RBCs treated sequentially with both neuraminidase and trypsin. Our results suggest that only parasites able to use some particular invasion pathways can invade SAO RBCs efficiently in culture. A similar situation might occur in the field.

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