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Ferrata Storti Foundation
Haematologica 2019 Volume 104(9):1866-1878
Platelet Biology & its Disorders
Fatal dysfunction and disintegration of thrombin-stimulated platelets
Oleg V. Kim,1,2 Tatiana A. Nevzorova,3 Elmira R. Mordakhanova,3 Anastasia A. Ponomareva,3,4 Izabella A. Andrianova,3 Giang Le Minh,3 Amina G. Daminova,3,4 Alina D. Peshkova,3 Mark S. Alber,2 Olga Vagin,5,6 Rustem I. Litvinov1,3 and John W. Weisel1
1University of Pennsylvania Perelman School of Medicine, Department of Cell and Developmental Biology, Philadelphia, PA, USA; 2University of California Riverside, Department of Mathematics, Riverside, CA, USA; 3Kazan Federal University, Institute of Fundamental Medicine and Biology, Kazan, Russian Federation; 4Kazan Institute of Biochemistry and Biophysics, FRC Kazan Scientific Center of RAS, Kazan, Russian Federation; 5Geffen School of Medicine at UCLA, Department of Physiology, Los Angeles, CA, USA and 6VA Greater Los Angeles Healthcare System, Los Angeles, CA, USA
ABSTRACT
Platelets play a key role in the formation of hemostatic clots and obstructive thrombi as well as in other biological processes. In response to physiological stimulants, including thrombin, platelets change shape, express adhesive molecules, aggregate, and secrete bioactive substances, but their subsequent fate is largely unknown. Here we exam- ined late-stage structural, metabolic, and functional consequences of throm- bin-induced platelet activation. Using a combination of confocal microscopy, scanning and transmission electron microscopy, flow cytome- try, biochemical and biomechanical measurements, we showed that throm- bin-induced activation is followed by time-dependent platelet dysfunction and disintegration. After ~30 minutes of incubation with thrombin, unlike with collagen or ADP, human platelets disintegrated into cellular fragments containing organelles, such as mitochondria, glycogen granules, and vac- uoles. This platelet fragmentation was preceded by Ca2+ influx, integrin aIIbβ3 activation and phosphatidylserine exposure (activation phase), fol- lowed by mitochondrial depolarization, generation of reactive oxygen species, metabolic ATP depletion and impairment of platelet contractility along with dramatic cytoskeletal rearrangements, concomitant with platelet disintegration (death phase). Coincidentally with the platelet frag- mentation, thrombin caused calpain activation but not activation of caspas- es 3 and 7. Our findings indicate that the late functional and structural dam- age of thrombin-activated platelets comprise a calpain-dependent platelet death pathway that shares some similarities with the programmed death of nucleated cells, but is unique to platelets, therefore representing a special form of cellular destruction. Fragmentation of activated platelets suggests that there is an underappreciated pathway of enhanced elimination of platelets from the circulation in (pro)thrombotic conditions once these cells have performed their functions.
Introduction
Platelets are blood cells that play a pivotal role in preventing bleeding (hemostasis) and obstructing blood vessels (thrombosis), in addition to other biological functions.1,2 Activated platelets mechanically contract blood clots and thrombi,3 which is an important pathogenic mechanism in thrombosis.4–6 Under (patho)physiological con- ditions, platelets activated by stimulants change their morphology, express adhesive molecules, undergo aggregation, and secrete bioactive substances. Disruption of these functions can have pathological consequences, including heart attack and stroke. However, the mechanisms underlying the subsequent fate of activated platelets, including platelet clearance, remain poorly defined. The structural and func-
Correspondence:
JOHN WEISEL
weisel@pennmedicine.upenn.edu
Received: July 18, 2018. Accepted: February 14, 2019. Pre-published: February 21, 2019.
doi:10.3324/haematol.2018.202309
Check the online version for the most updated information on this article, online supplements, and information on authorship & disclosures: www.haematologica.org/content/104/9/1866
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