Changes in the cytoskeletal structure of human platelets following thrombin activation.

Jennings, Lisa K.; Fox, Joan E.B.; Edwards, Harold H.; Phillips, David R.

doi:10.1016/s0021-9258(19)69080-0

Cited by 220 publications

(41 citation statements)

References 31 publications

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“…For the preparation of unstimulated discoid platelets we used stabilizing buffer containing 10 ng/ml prostacyclin (gift of Dr. Y. Shibata), again based on Fox et al (11). For activation, platelets were incubated for 3 min with 1 U/m1 human plasma thrombin (Sigma Chemical Co., St. Louis, MO) in stabilizing buffer at room temperature (23). Shapes of the platelets in these samples were examined by Nomarski differential interference contrast microscopy before further processing.…”

Section: Preparation Of Plateletsmentioning

confidence: 99%

“…We saponized the samples without fixation, just as in the preparation for quick-freezing, to see the influence of the permeabilization and found that many myosin molecules existed in saponin-extracted activated cytoskeletons, as shown biochemically in Triton-extracted cytoskeleton (23,30).…”

Section: Localization Of Myosinmentioning

confidence: 99%

“…The results suggest that myosin forms a gel with actin filaments in activated platelets. Close associations between actin filaments and organelles in activated platelets suggests that contraction of this actomyosin gel could bring about the observed centralization of organelles.p LATELETS, which play an important role in the maintenance of hemostasis have been a favorite subject for the study of cytoskeletal reorganization of actin filaments and their related proteins in nonmuscle cells because platelets undergo dynamic morphological changes from disks to irregular forms with several pseudopods and centralize their organelles after mechanical or chemical activation (23,26). Platelets contain not only actin, myosin, and tubulin, but also actin-binding protein (ABP), ~ r vinculin, talin, and other actin-related proteins (8,31,32,36,39,40).…”

mentioning

confidence: 99%

“…p LATELETS, which play an important role in the maintenance of hemostasis have been a favorite subject for the study of cytoskeletal reorganization of actin filaments and their related proteins in nonmuscle cells because platelets undergo dynamic morphological changes from disks to irregular forms with several pseudopods and centralize their organelles after mechanical or chemical activation (23,26). Platelets contain not only actin, myosin, and tubulin, but also actin-binding protein (ABP), ~ r vinculin, talin, and other actin-related proteins (8,31,32,36,39,40).…”

mentioning

confidence: 99%

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Cytoskeletal reorganization of human platelets after stimulation revealed by the quick-freeze deep-etch technique.

Nakata

Hirokawa

1987

The Journal of Cell Biology

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Abstract. We studied the cytoskeletal reorganization of saponized human platelets after stimulation by using the quick-freeze deep-etch technique, and examined the localization of myosin in thrombin-treated platelets by immunocytochemistry at the electron microscopic level.In unstimulated saponized platelets we observed cross-bridges between: adjoining microtubules, adjoining actin filaments, microtubules and actin filaments, and actin filaments and plasma membranes.After activation with 1 U/ml thrombin for 3 min, massive arrays of actin filaments with mixed polarity were found in the cytoplasm. Two types of crossbridges between actin filaments were observed: short cross-bridges (11 5:2 nm), just like those observed in the resting platelets, and longer ones (22 + 3 nm). Actin filaments were linked with the plasma membrane via fine short filaments and sometimes ended on the membrane. Actin filaments and microtubules frequently ran close to the membrane organelles. We also found that actin filaments were associated by end-on attachments with some organelles. Decoration with subfragment 1 of myosin revealed that all the actin filaments associated end-on with the membrane pointed away in their polarity.Immunocytochemical study revealed that myosin was present in the saponin-extracted cytoskeleton after activation and that myosin was localized on the filamentous network. The results suggest that myosin forms a gel with actin filaments in activated platelets. Close associations between actin filaments and organelles in activated platelets suggests that contraction of this actomyosin gel could bring about the observed centralization of organelles. p LATELETS, which play an important role in the maintenance of hemostasis have been a favorite subject for the study of cytoskeletal reorganization of actin filaments and their related proteins in nonmuscle cells because platelets undergo dynamic morphological changes from disks to irregular forms with several pseudopods and centralize their organelles after mechanical or chemical activation (23,26). Platelets contain not only actin, myosin, and tubulin, but also actin-binding protein (ABP), ~ r vinculin, talin, and other actin-related proteins (8,31,32,36,39,40). Recent studies also demonstrated the existence of microtubule-associated proteins (24,38). Biochemical studies have demonstrated changes in some cytoskeletal components with increase of actin, myosin, and other proteins in the Tritoninsoluble cytoskeletons after activation (2, 4-6, 23, 30), and immunofluorescence studies of redistribution of these contractile proteins have supported these ideas (8,9,36,39,40). Redistribution of several membrane proteins was studied at the electron microscopic level (27,33,41,42).A detailed observation of three-dimensional architecture in situ in both resting and activated platelets at the molecular 1. Abbreviations used in this paper: ABE actin-binding protein; S1, myosin subfragment 1.level is necessary for a more complete understanding of the reorganization of the platelet cytoske...

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Section: Preparation Of Plateletsmentioning

confidence: 99%

Section: Localization Of Myosinmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Cytoskeletal reorganization of human platelets after stimulation revealed by the quick-freeze deep-etch technique.

Nakata

Hirokawa

1987

The Journal of Cell Biology

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“…As a result, our simulation results could underestimate the number of platelets adhered to the injured vessel wall. In addition, platelets become more deformable upon activation through reordering of actin network in their membrane and undergo drastic morphological changes [62,63]. In this work, we focus on the initial steps of thrombus formation triggered by coagulation cascade and thus did not consider the shape change of activated platelets to reduce the complexity of the model.…”

Section: Discussionmentioning

confidence: 99%

Integrating blood cell mechanics, platelet adhesive dynamics and coagulation cascade for modelling thrombus formation in normal and diabetic blood

Yazdani

Deng

et al. 2021

J. R. Soc. Interface.

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Normal haemostasis is an important physiological mechanism that prevents excessive bleeding during trauma, whereas the pathological thrombosis especially in diabetics leads to increased incidence of heart attacks and strokes as well as peripheral vascular events. In this work, we propose a new multiscale framework that integrates seamlessly four key components of blood clotting, namely transport of coagulation factors, coagulation kinetics, blood cell mechanics and platelet adhesive dynamics, to model the development of thrombi under physiological and pathological conditions. We implement this framework to simulate platelet adhesion due to the exposure of tissue factor in a three-dimensional microchannel. Our results show that our model can simulate thrombin-mediated platelet activation in the flowing blood, resulting in platelet adhesion to the injury site of the channel wall. Furthermore, we simulate platelet adhesion in diabetic blood, and our results show that both the pathological alterations in the biomechanics of blood cells and changes in the amount of coagulation factors contribute to the excessive platelet adhesion and aggregation in diabetic blood. Taken together, this new framework can be used to probe synergistic mechanisms of thrombus formation under physiological and pathological conditions, and open new directions in modelling complex biological problems that involve several multiscale processes.

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Myosin translocation in retinal pericytes during free-radical induced apoptosis

Shojaee

Patton²,

Hechtman

et al. 1999

J. Cell. Biochem.

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Vascular pathologies induced by ischemia/reperfusion involve the production of reactive oxygen species (ROS) that in part cause tissue injury. The production of ROS that occurs upon reperfusion activates specific second messenger pathways. In diabetic retinopathy there is a characteristic loss of the microvascular pericyte. Pericytes are more sensitive than endothelial cells to low concentrations of ROS, such as hydrogen peroxide (H(2)O(2)) when tested in vitro. Whether the pericyte loss is due to toxic cell death triggered by the noxious H(2)O(2) or apoptosis, due to activation of specific second messenger pathways, is unknown. During apoptosis, a cell's nucleus and cytoplasm condense, the cell becomes fragmented, and ultimately forms apoptotic bodies. It is generally assumed that apoptosis depends on nuclear signaling, but cytoplasmic morphological processes are not well described. We find that exposing cultured retinal pericytes to 100 microM H(2)O(2) for 30 min leads to myosin heavy chain translocation from the cytosol to the cytoskeleton and a significant decrease in cell surface area. Pericyte death follows within 60-120 min. Exposing cells to 150 mJ/cm(2) ultraviolet radiation, an alternate free radical generating system, also causes pericyte myosin translocation and apoptosis. Proteolytic cleavage of actin is not observed in pericyte apoptosis. 3-aminobenzamide, a pharmacological inhibitor of the cleavage and activation of the DNA-repairing enzyme poly (ADP-ribose) polymerase (PARP) inhibits pericyte apoptosis, and prevents myosin translocation. Deferoxamine, an iron chelator known to interfere with free radical generation, also inhibits pericyte myosin translocation, contractility, and cell death. Myosin translocation to the cytoskeleton may be an early step in assembly of a competent contractile apparatus, which is involved in apoptotic cell condensation. These results suggest that pericyte loss associated with increased free radical production in diabetic retina may be by an apoptotic phenomenon.

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Changes in the cytoskeletal structure of human platelets following thrombin activation.

Cited by 220 publications

References 31 publications

Cytoskeletal reorganization of human platelets after stimulation revealed by the quick-freeze deep-etch technique.

Cytoskeletal reorganization of human platelets after stimulation revealed by the quick-freeze deep-etch technique.

Integrating blood cell mechanics, platelet adhesive dynamics and coagulation cascade for modelling thrombus formation in normal and diabetic blood

Myosin translocation in retinal pericytes during free-radical induced apoptosis

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