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Biochem J. 2016 Feb 15;473(4):435-48. doi: 10.1042/BJ20150779. Epub 2015 Nov 25.

Calpain-controlled detachment of major glycoproteins from the cytoskeleton regulates adhesive properties of activated phosphatidylserine-positive platelets.

Author information

1
Laboratory of Molecular Mechanisms of Haemostasis, Centre for Theoretical Problems of Physicochemical Pharmacology, 4 Kosygina Str., Moscow 119334, Russia Cellular Haemostasis and Thrombosis, Federal Research and Clinical Centre of Paediatric Haematology, Oncology and Immunology, Moscow, 1 Samory Mashela Str., Moscow 117198, Russia.
2
Laboratory of Immunity Stimulation, Institute of Immunology FMBA of Russia, 24 Kashirskoye sh., Moscow 115478, Russia.
3
Cellular Haemostasis and Thrombosis, Federal Research and Clinical Centre of Paediatric Haematology, Oncology and Immunology, Moscow, 1 Samory Mashela Str., Moscow 117198, Russia Laboratory of Cell Biophysics, Centre for Theoretical Problems of Physicochemical Pharmacology, 4 Kosygina Str., Moscow 119334, Russia Faculty of Physics, Moscow State University, Vorobievy Gory, GSP-1, Moscow 119991, Russia Faculty of Biological and Medical Physics, Moscow Institute of Physics and Technology, Dolgoprudny, Russia.
4
Laboratory of Molecular Mechanisms of Haemostasis, Centre for Theoretical Problems of Physicochemical Pharmacology, 4 Kosygina Str., Moscow 119334, Russia Cellular Haemostasis and Thrombosis, Federal Research and Clinical Centre of Paediatric Haematology, Oncology and Immunology, Moscow, 1 Samory Mashela Str., Moscow 117198, Russia Faculty of Physics, Moscow State University, Vorobievy Gory, GSP-1, Moscow 119991, Russia Faculty of Biological and Medical Physics, Moscow Institute of Physics and Technology, Dolgoprudny, Russia Therapeutic Faculty, Pirogov Russian National Research Medical University, Moscow 117997, Russia mapanteleev@yandex.ru.

Abstract

In resting platelets, adhesive membrane glycoproteins are attached to the cytoskeleton. On strong activation, phosphatidylserine(PS)-positive and -negative platelet subpopulations are formed. Platelet activation is accompanied by cytoskeletal rearrangement, although the glycoprotein attachment status in these two subpopulations is not clear. We developed a new, flow cytometry-based, single-cell approach to investigate attachment of membrane glycoproteins to the cytoskeleton in cell subpopulations. In PS-negative platelets, adhesive glycoproteins integrin αIIbβ3, glycoprotein Ib and, as shown for the first time, P-selectin were associated with the cytoskeleton. In contrast, this attachment was disrupted in PS-positive platelets; it was retained to some extent only in the small convex regions or 'caps'. It correlated with the degradation of talin and filamin observed only in PS-positive platelets. Calpain inhibitors essentially prevented the disruption of membrane glycoprotein attachment in PS-positive platelets, as well as talin and filamin degradation. With the suggestion that detachment of glycoproteins from the cytoskeleton may affect platelet adhesive properties, we investigated the ability of PS-positive platelets to resist shear-induced breakaway from the immobilized fibrinogen. Shear rates of 500/s caused PS-positive platelet breakaway, but their adhesion stability increased more than 10-fold after pretreatment of the platelets with calpain inhibitor. In contrast, the ability of PS-positive platelets to adhere to immobilized von Willebrand's factor at 100/s was low, but this was not affected by the preincubation of platelets with a calpain inhibitor. Our data suggest that calpain-controlled detachment of membrane glycoproteins is a new mechanism that is responsible for the loss of ability of the procoagulant platelets to resist detachment from thrombi by high shear stress.

KEYWORDS:

adhesion; adhesive glycoproteins; calpain; cytoskeleton; platelet; shear

PMID:
26607836
DOI:
10.1042/BJ20150779
[Indexed for MEDLINE]

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