Effect of Red Blood Cell Aging In Vivo on Their Aggregation Properties In Vitro: Measurements with Laser Tweezers

Red blood cell (RBC) aggregation highly influences hemorheology and blood microcirculation in the human body. The aggregation properties of RBCs can vary due to numerous factors, including RBC age. The aim of this work was to estimate in vitro the differences in the RBC aggregation properties of different RBC age populations in single-cell experiments using laser tweezers. RBCs from five healthy volunteers were separated into four subpopulations by Percoll density gradient centrifugation. Each subpopulation of the RBC was separately resuspended in autologous plasma or dextran 70 kDa (50 mg/mL). The aggregation force between the single cells was measured with holographic laser tweezers. The obtained data demonstrated an enhancement of RBC aggregation force in doublets with age: the older the cells, the higher the aggregation force. The obtained data revealed the differences between the aggregation and aggregability of RBC in dependence of the RBC in vivo age.

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Vesiculation of Red Blood Cells in the Blood Bank: A Multi-Omics Approach towards Identification of Causes and Consequences

Proteomes ◽

10.3390/proteomes8020006 ◽

2020 ◽

Vol 8 (2) ◽

pp. 6

Author(s):

Joames K. Freitas Leal ◽

Edwin Lasonder ◽

Vikram Sharma ◽

Jürgen Schiller ◽

Giuseppina Fanelli ◽

...

Keyword(s):

Blood Cell ◽

Red Blood Cells ◽

Red Blood Cell ◽

Blood Cells ◽

Blood Bank ◽

Cell Aging ◽

Lipid Organization ◽

Membrane Components

Microvesicle generation is an integral part of the aging process of red blood cells in vivo and in vitro. Extensive vesiculation impairs function and survival of red blood cells after transfusion, and microvesicles contribute to transfusion reactions. The triggers and mechanisms of microvesicle generation are largely unknown. In this study, we combined morphological, immunochemical, proteomic, lipidomic, and metabolomic analyses to obtain an integrated understanding of the mechanisms underlying microvesicle generation during the storage of red blood cell concentrates. Our data indicate that changes in membrane organization, triggered by altered protein conformation, constitute the main mechanism of vesiculation, and precede changes in lipid organization. The resulting selective accumulation of membrane components in microvesicles is accompanied by the recruitment of plasma proteins involved in inflammation and coagulation. Our data may serve as a basis for further dissection of the fundamental mechanisms of red blood cell aging and vesiculation, for identifying the cause-effect relationship between blood bank storage and transfusion complications, and for assessing the role of microvesicles in pathologies affecting red blood cells.

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Effects of increased plasma viscosity and red blood cell aggregation on blood viscosity in vivo

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1981.241.4.h513 ◽

1981 ◽

Vol 241 (4) ◽

pp. H513-H518 ◽

Cited By ~ 9

Author(s):

L. Gustafsson ◽

L. Appelgren ◽

H. E. Myrvold

Keyword(s):

Blood Cell ◽

Red Blood Cell ◽

Apparent Viscosity ◽

Cell Aggregation ◽

Plasma Viscosity ◽

Pressure Flow ◽

Red Blood Cell Aggregation ◽

Rbc Aggregation

The effects of increased plasma viscosity and induced red blood cell (RBC) aggregation on apparent viscosity of blood in vivo in the skeletal muscle of the dog were studied. Apparent viscosity in vivo was determined in the isolated and vasodilated calf muscles of one hindlimb by comparing pressure-flow relationships for RBC suspensions with pressure-flow relationships for a Newtonian solution of known viscosity. RBC suspensions of increased plasma viscosity with and without RBC aggregation were obtained by substituting plasma with isoviscous solutions of high- and low-molecular-weight dextran in saline. Hematocrits of the suspensions were adjusted to either 45 or 60%. The viscosities of the suspensions in vitro were determined in a Wells-Brookfield viscometer. Apparent viscosity of blood in vivo was found to be mainly dependent on the viscosity of plasma. RBC aggregation had no significant influence on the viscosity in vivo.

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Absence of the Adaptor Protein PEA-15 Is Associated with Altered Pattern of Th Cytokines Production by Activated CD4+ T Lymphocytes In Vitro, and Defective Red Blood Cell Alloimmune Response In Vivo

PLoS ONE ◽

10.1371/journal.pone.0136885 ◽

2015 ◽

Vol 10 (8) ◽

pp. e0136885 ◽

Cited By ~ 6

Author(s):

Stéphane Kerbrat ◽

Benoit Vingert ◽

Marie-Pierre Junier ◽

Flavia Castellano ◽

François Renault-Mihara ◽

...

Keyword(s):

T Lymphocytes ◽

Blood Cell ◽

Red Blood Cell ◽

Adaptor Protein

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Hemoglobin-spectrin complexes: interference with spectrin tetramer assembly as a mechanism for compartmentalization of band 1 and band 2 complexes

Blood ◽

10.1182/blood.v86.1.366.bloodjournal861366 ◽

1995 ◽

Vol 86 (1) ◽

pp. 366-371 ◽

Cited By ~ 15

Author(s):

CR Kiefer ◽

JF Trainor ◽

JB McKenney ◽

CR Valeri ◽

LM Snyder

Keyword(s):

Blood Cell ◽

Western Blot ◽

Red Blood Cell ◽

Blot Analysis ◽

Natural Phenomenon ◽

Cell Aging ◽

Cell Deformability ◽

Self Association

The irreducible complexation of hemoglobin with spectrin is a natural phenomenon of red blood cell aging, positively correlating with increasing cell density and decreasing cell deformability. The current study begins to address the role of these complexes in the disruption of membrane skeletal physiology and structure. The effect of bound hemoglobin on spectrin dimer self-association was investigated in vitro. The extent of conversion of isolated spectrin dimers to tetramers was evaluated as a function of peroxide-induced globin complexation before the conversion incubations. The incremental accumulation of tetramer was observed to decrease with increasing peroxide concentration used in the globin complexation step. The role of oxidized heme in this process was made apparent by the inability of carboxyhemoglobin to inhibit tetramer accumulation. A Western blot analysis of naturally formed globin-spectrin conjugates demonstrated irreducible complexes of globin with both bands 1 and 2. The complexes are tentatively designated “h1” and “h2”. This analysis also demonstrated that h1 is completely extractable from cell ghosts, whereas h2 is only 50% extractable. These findings are incorporated into a hypothesis linking globin-spectrin complexation and the consequent inhibition of spectrin dimer self-association to the clustered band 3 senescence antigen (Low et al, Science 227:531, 1985).

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ASPIRIN MODIFIES RED BLOOD CELL BEHAVIOUR (RBC) IN THE PLATELET-RBC INTERACTION

10.1055/s-0038-1644821 ◽

1987 ◽

Author(s):

M T Santos ◽

J Aznar ◽

J Valles ◽

J L Perez-Reguejo

Keyword(s):

Single Dose ◽

Blood Cell ◽

Platelet Activation ◽

Red Blood Cell ◽

Normal Subjects ◽

Antithrombotic Agent ◽

Cell Behaviour ◽

Insight Into

RBC stimulate the initial stages of platelet activation by collagen as evaluated by the BASIC wave (Perez-Requejo et al. Thromb Haemostas 54:799 1985). In order to get some insight into the mechanisms of platelet-RBC interactions, a BASIC wave was induced by lug/ml of collagen after mixing "in vitro" platelets and RBC obtained both before and two hours after a single dose of 500 mg of ASA from normal subjects. The TXB2 formed was also evaluated. The results show (Table) that non aspirinized RBC (non-ASA-RBC) increase the BASIC wave intensity of aspirinized platelets (ASA-PRP) by a cyclooxygenase-independent pathway since no increase in TXB2 was observed (Exp 1), while both non-ASA-RBC (Exp 2) and ASA-RBC (Exp 3) activate non-ASA platelets with theparticipation of the cyclooxygenase system, since an increase in TXA2 was found.A comparison of the effect of non-ASA-RBC (Exp 1) and ASA-RBC (Exp 4) on aspirinized platelets shows that ASA modifies the RBC behaviour associated with estimulation of platelets by a cyclooxygenase-independent pathway. This effect of ASA on RBC is nottransient and lasts at least 48 hours after ASA ingestion. In addition, when asmall proportion of nonASA platelets (10%) is mixed with aspirinized platelets(90%) and ASA-RBC - a situation that can be encountered "in vivo" inthe hours following ASA ingestion - the intensity of the BASIC wave is 89% of that obtained when all the platelets are non aspirinized. This RBC effect on the mixtureof ASA and nonASA platelets, may help explain the sometimes contradictory effect of ASA as an antithrombotic agent.

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The Presence of Novel Amino Acids in the Cytoplasmic Domain of Stem Cell Factor Results in Hematopoietic Defects inSteel17H Mice

Blood ◽

10.1182/blood.v94.6.1915 ◽

1999 ◽

Vol 94 (6) ◽

pp. 1915-1925 ◽

Cited By ~ 20

Author(s):

Reuben Kapur ◽

Ryan Cooper ◽

Xingli Xiao ◽

Mitchell J. Weiss ◽

Peter Donovan ◽

...

Keyword(s):

Amino Acids ◽

Bone Marrow ◽

Biological Activity ◽

Stem Cell ◽

Blood Cell ◽

Red Blood Cell ◽

Stem Cell Factor ◽

Cytoplasmic Domain

Abstract Stem cell factor (SCF) is expressed as an integral membrane growth factor that may be differentially processed to produce predominantly soluble (S) (SCF248) or membrane-associated (MA) (SCF220) protein. A critical role for membrane presentation of SCF in the hematopoietic microenvironment (HM) has been suggested from the phenotype of the Steel-dickie(Sld) mice, which lack MA SCF, and by studies performed in our laboratory (and by others) using long-term bone marrow cultures and transgenic mice expressing different SCF isoforms.Steel17H (Sl17H) is an SCF mutant that demonstrates melanocyte defects and sterility in males but not in females. The Sl17H allele contains a intronic mutation resulting in the substitution of 36 amino acids (aa’s) in the SCF cytoplasmic domain with 28 novel aa’s. This mutation, which affects virtually the entire cytoplasmic domain of SCF, could be expected to alter membrane SCF presentation. To investigate this possibility, we examined the biochemical and biologic properties of the Sl17H-encoded protein and its impact in vivo and in vitro on hematopoiesis and on c-Kit signaling. We demonstrate that compound heterozygous Sl/Sl17H mice manifest multiple hematopoietic abnormalities in vivo, including red blood cell deficiency, bone marrow hypoplasia, and defective thymopoiesis. In vitro, both S and MA Sl17H isoforms of SCF exhibit reduced cell surface expression on stromal cells and diminished biological activity in comparison to wild-type (wt) SCF isoforms. These alterations in presentation and biological activity are associated with a significant reduction in the proliferation of an SCF-responsive erythroid progenitor cell line and in the activation of phosphatidylinositol 3-Kinase/Akt and mitogen-activated protein-Kinase signaling pathways. In vivo, transgene expression of the membrane-restricted (MR) (SCFX9/D3) SCF in Sl/Sl17H mutants results in a significant improvement in peripheral red blood cell counts in comparison toSl/Sl17H mice.

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Molecular species of membrane phospholipids containing arachidonic acid and linoleic acid contribute to the interindividual variability of red blood cell Na+-Li+ countertransport: In vivo and in vitro evidence

The Journal of Membrane Biology ◽

10.1007/bf00233791 ◽

1993 ◽

Vol 133 (2) ◽

pp. 99-106 ◽

Cited By ~ 12

Author(s):

Bernd Engelmann ◽

Jochen Duhm ◽

Ulrike M. Schönthier ◽

Sabine Streich ◽

Jos A. F. Op den Kamp ◽

...

Keyword(s):

Arachidonic Acid ◽

Linoleic Acid ◽

Blood Cell ◽

Molecular Species ◽

Red Blood Cell ◽

Interindividual Variability ◽

Membrane Phospholipids

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Pathophysiology of Thrombocytopenia and Anemia in Mice Lacking Transcription Factor NF-E2

Blood ◽

10.1182/blood.v94.9.3037 ◽

1999 ◽

Vol 94 (9) ◽

pp. 3037-3047 ◽

Cited By ~ 69

Author(s):

Jack Levin ◽

Jin-Peng Peng ◽

Georgiann R. Baker ◽

Jean-Luc Villeval ◽

Patrick Lecine ◽

...

Keyword(s):

Transcription Factor ◽

Blood Cell ◽

Red Blood Cell ◽

Fetal Life ◽

Wild Type ◽

Cell Lineages ◽

Cell Fragments ◽

Proliferation Potential

Abstract Expression of the p45 subunit of transcription factor NF-E2 is restricted to selected blood cell lineages, including megakaryocytes and developing erythrocytes. Mice lacking p45 NF-E2 show profound thrombocytopenia, resulting from a late arrest in megakaryocyte differentiation, and a number of red blood cell defects, including anisocytosis and hypochromia. Here we report results of studies aimed to explore the pathophysiology of these abnormalities. Mice lacking NF-E2 produce very few platelet-like particles that display highly disorganized ultrastructure and respond poorly to platelet agonists, features consistent with the usually lethal hemorrhage in these animals. Thrombocytopenia was evident during fetal life and was not corrected by splenectomy in adults. Surprisingly, fetal NF-E2–deficient megakaryocyte progenitors showed reduced proliferation potential in vitro. Thus, NF-E2 is required for regulated megakaryocyte growth as well as for differentiation into platelets. All the erythroid abnormalities were reproduced in lethally irradiated wild-type recipients of hematopoietic cells derived from NF-E2-null fetuses. Whole blood from mice lacking p45 NF-E2 showed numerous small red blood cell fragments; however, survival of intact erythrocytes in vivo was indistinguishable from control mice. Considered together, these observations indicate a requirement for NF-E2 in generating normal erythrocytes. Despite impressive splenomegaly at baseline, mice lacking p45 NF-E2 survived splenectomy, which resulted in increased reticulocyte numbers. This reveals considerable erythroid reserve within extra-splenic sites of hematopoiesis and suggests a role for the spleen in clearing abnormal erythrocytes. Our findings address distinct aspects of the requirements for NF-E2 in blood cell homeostasis and establish its roles in proper differentiation of megakaryocytes and erythrocytes.

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Abnormal Erythropoiesis in Microgravity.

Blood ◽

10.1182/blood.v104.11.3701.3701 ◽

2004 ◽

Vol 104 (11) ◽

pp. 3701-3701 ◽

Cited By ~ 1

Author(s):

Kun Xu ◽

Keith V. Holubec ◽

John E. Love ◽

Thomas J. Goodwin ◽

Arthur J. Sytkowski

Keyword(s):

Blood Cell ◽

Cell Line ◽

Red Blood Cell ◽

Bone Marrow Cells ◽

Cell Mass ◽

Erythroid Cell ◽

Specific Marker ◽

Murine Erythroleukemia

Abstract Humans and experimental animals subjected to microgravity, such as experienced during space flight, exhibit alterations in erythropoiesis, including changes in red blood cell morphology, survival and a reduction in red blood cell mass. Some of these alterations have been attributed to a disruption of normal in vivo erythropoietin physiology. However, human bone marrow cells grown on orbit showed a profound reduction in the number of erythroid cells, suggesting a cellular component. We now report the results of a study carried out on orbit on the International Space Station (ISS UF-1) in which an erythroid cell line was induced to differentiate. Rauscher murine erythroleukemia cells, a continuous cell line that can undergo erythropoietin (Epo)- or chemical-induced differentiation similar to normal erythropoiesis, were cultured for 6 days either in microgravity on board the ISS or on earth and then for 3 days in the absence or presence of 50 U Epo/ml or 0.7% dimethyl sulfoxide (DMSO). The cells were fixed, stored on orbit and returned to earth for study. Compared to ground-based controls, cells cultured in microgravity exhibited a greater degree of differentiation (hemoglobinization) (p<0.01). However, TER-119 antigen, a specific marker of the late stages of murine erythroid differentiation, was not detected on the surface of cells grown in microgravity. A significantly higher percentage (p<0.05) of cell clusters formed on orbit, whereas actin content appeared reduced. Furthermore, there was a more profound loss of actin stress fibers in microgravity following Epo or DMSO treatment. These results demonstrate abnormal erythropoiesis in vitro in microgravity and are consistent with the hypothesis that erythropoiesis is affected by gravitational forces at the cellular level.(Supported by NASA Grants NAG9-1368 and NAG2-1592 to AJS)

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