ABSTRACT
Coagulation and optical (based on chromogenic substrate) methods were employed to examine antithrombin activity of erythrocytes and erythrocyte-derived microvesicles isolated days 7, 14, 21, and 28 on erythrocyte storage. The erythrocyte-derived microvesicles decelerated fibrin clot formation from fibrinogen in the presence of exogenous thrombin both with and without heparin. Microvesicles reduced optical density of chromogenic substrate. These data suggest that erythrocyte-derived microvesicles display a prominent antithrombin activity, which significantly increases during erythrocyte storage.
Subject(s)
Antithrombin Proteins/chemistry , Blood Preservation/methods , Cell-Derived Microparticles/chemistry , Erythrocytes/chemistry , Fibrinogen/chemistry , Thrombin/chemistry , Adenine/chemistry , Blood Coagulation , Blood Coagulation Tests , Cells, Cultured , Chromogenic Compounds/analysis , Citrates/chemistry , Erythrocytes/cytology , Fibrin/chemistry , Glucose/chemistry , Heparin/chemistry , Humans , Phosphates/chemistry , Refrigeration/methods , SpectrophotometryABSTRACT
An aggregation of erythrocytes in autologous plasma after blood storage for 14 days at 4 °C was studied using photometry and light microscopy. The decrease of ATP content, the formation of echinocytes and spheroechinocytes, the decrease of rouleaux form of erythrocyte aggregation were observed during the storage. On the other hand the aggregates of echinocytes were formed in the stored blood. The addition of plasma from the fresh blood didn't restore the normal discocytic shape and aggregation of erythrocytes in the stored blood. The possible mechanisms of erythrocytes and echinocytes aggregation are discussed.
Subject(s)
Adenosine Triphosphate/deficiency , Erythrocyte Aggregation , Erythrocytes/cytology , Erythrocyte Count , Fixatives , Glutaral , Humans , Microscopy , Primary Cell CultureABSTRACT
We studied the influence of erythrocyte microvesicles, isolated after 4 weeks incubation the washed and the unwashed erythrocytes healthy donors from preservative, on spontaneous (flow-induced) aggregation of platelets. It was found out that microvesicles of the unwashed erythrocytes enhanced the spontaneous aggregation of platelets and also formation of fibrin. Microvesicles from the washed erythrocytes reduce the platelet aggregation. Possible mechanisms of the effect of erythrocyte microvesicles on platelet aggregation were investigated.
Subject(s)
Blood Platelets/metabolism , Cell-Derived Microparticles/metabolism , Erythrocytes/metabolism , Fibrin/metabolism , Platelet Aggregation , Adult , Blood Platelets/cytology , Erythrocytes/cytology , Humans , MaleABSTRACT
INTRODUCTION: Hemorheological disorders play an important part in pathogenesis of acute period of burn injury. This mechanism remains practically unstudied. Thus, unknown is the role of hemoglobin glycation and erythrocyte microvesiculation in the decrease in erythrocyte deformability after thermal trauma. METHODS: Research was performed on 30 blood samples of burn patients in the acute period and 40 blood samples of healthy donors. The number of erythrocyte-derived microvesicles was determined by flow cytometry and then standardized in the samples; the microvesicles were preliminarily separated by ultracentrifugation at 100,000 g, for 60 minutes. Electrophoretic mobility of erythrocytes was measured in a processing chamber of the optical cuvette under the light microscope. Deformability of erythrocytes was assessed by the level of their extension in the artificial shear flow. RESULTS: It was found that the amount of HbA1c in red blood cells of burn patients demonstrated a 2-fold increase compared to healthy donors. In the experiments in vitro it was proved that deformability of erythrocytes correlates with the level of hemoglobin glycation. Hb glycation leads to the increased rigidity of erythrocytes also by increasing their microvesiculation. The number of microvesicles derived from red blood cells of burn patients demonstrated a 3.47-fold increase compared to healthy donors. An important reason for microvesiculation is the destabilization of lipid complex of erythrocyte membrane, which is accompanied by the increase in the erythrocyte negative charge. It can be concluded that Hb glycation and redistribution of erythrocyte membrane phospholipids are he important reasons for the increase erythrocyte microvesiculation and are accompanied by the decrease in erythrocyte deformability after thermal trauma.
Subject(s)
Burns/blood , Cell-Derived Microparticles/metabolism , Erythrocyte Deformability , Erythrocytes/metabolism , Hematologic Diseases/blood , Hemoglobins/metabolism , Adolescent , Adult , Aged , Burns/complications , Cell-Derived Microparticles/pathology , Erythrocytes/pathology , Female , Hematologic Diseases/etiology , Humans , Male , Middle AgedABSTRACT
Burn Injury is accompanied by a significant homeostasis disorder, including the disorder of primary homeostasis, associated with aggregation of platelets. The role of erythrocyte-derived microvesicles in this process has not undergone thorough research. Microvesicles were isolated from washed erythrocytes after one day of storage by ultracentrifugation at 100000 g. The number of MVs was determined by flow cytometry and was standardized in the samples. Heparin-dependent and heparin-independent antithrombin activity in erythrocyte microvesicles was studied by coagulation method. We studied platelet aggregation induced and not induced by ADP under the conditions of artificial shear flow. It was shown that at the early stage of bum injury the number of erythrocyte-derived microvesicles in blood demonstrated a 4.2-fold ncrease. We determined that microvesicles, derived from the erythrocytes of burn patients displayed a significantly less aggregation activity than the microvesicles from donors. The main reason is a considerably lower antithrombin activity in the erythrocyte microvesicles of bum patients. Thus, we can conclude that the decrease of antiaggregation and antithrombin activity of erythrocyte microvesicles associated with the increase in their concentration in blood contributes to thrombophilia of bum patients. Keywords: erythrocytes, microvesicles, bum injury, platelet aggregation, antithrombin activity
Subject(s)
Blood Platelets/metabolism , Burns/metabolism , Cell-Derived Microparticles/metabolism , Erythrocytes/metabolism , Platelet Aggregation , Adolescent , Adult , Blood Platelets/pathology , Burns/pathology , Cell-Derived Microparticles/pathology , Erythrocytes/pathology , Female , Humans , Male , Middle AgedABSTRACT
We studied the effect of erythrocyte microvesicles on various stages of platelet-free autoplasma coagulation process. The erythrocyte microvesicles not only exhibited procoagulant activity, but also produced a pronounced anticoagulant effect by inhibiting fibrin formation process. The effects of microvesicles isolated from erythrocytes after 24- and 48-hour cell storage on plasma coagulation were different. The hemocoagulation effects of the "light" and "heavy" microvesicles were studied.
