TMEM16F enhances procoagulant activity and thrombosis through platelet microparticle generation
Date
2016
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
University of Delaware
Abstract
Increased procoagulant plasma microparticles are detected in prothrombotic
and hypercoagulable states and may contribute significantly to thrombosis, the single
greatest cause of death worldwide. Platelet microparticles are strongly procoagulant
because of the exposed membrane phosphatidylserine (PS) that interacts with the
coagulation cascade enabling rapid, localized and efficient thrombin generation
subsequently resulting in fibrin formation and further platelet activation. Recently,
TMEM16F, a novel putative 8-transmembrane spanning calcium-activated calciumpermeable
channel, emerged as an essential component required for calciumdependent
PS exposure on platelet membranes, procoagulant activity and thrombosis,
yet the role of TMEM16F in microparticle formation and subsequent TMEM16Fdependent
microparticle generation on thrombosis has been unclear. ☐ To determine whether TMEM16F is required for optimal microparticle
formation and PS exposure, platelets from mice lacking (TMEM16F KO) or
expressing TMEM16F (WT) were treated with physiological agonists or calcium
ionophore and microparticle numbers and PS exposure on platelets were assessed by
flow cytometry. TMEM16F KO platelets displayed an impaired ability to generate
optimal PS+ and total microparticles and also PS exposure in response to either
thrombin and collagen or A23187. Moreover, both CD41+ (platelet- and
megakaryocyte- derived) and total microparticles in healthy circulation were lower in
TMEM16F KO mice compared to WT mice, underscoring the physiological
requirement for TMEM16F in microparticle formation in vivo. ☐ It was demonstrated that TMEM16F-dependent platelet microparticles are
important for procoagulant activity and thrombosis, since both platelet-rich plasma
(PRP) and platelet-poor plasma (PPP) from TMEM16F KO mice had diminished
thrombin generation compared to those isolated from WT mice. Subsequent addition
of platelet-derived microparticles to either TMEM16F-deficient PRP or PPP
completely restored thrombin generation to the level of that observed in WT PRP.
Furthermore, TMEM16F KO mice displayed defective thrombosis in vivo using a
FeCl3-injury to the carotid artery model and the thrombotic defect was overcome with
injection of isolated platelet microparticles, underscoring the requirement of
TMEM16F-dependent platelet microparticles for thrombosis. Notably, number of
platelets, leukocytes and erythrocytes in addition to platelet aggregation responses
remained intact in TMEM16F KO mice, though mean platelet volume was slightly
higher compared to WT mice, suggesting that loss of TMEM16F does not interrupt the
steady-state levels of platelets and other circulating cells or platelet aggregation
pathways, but platelets may be larger on average because of reduced microparticle
formation. ☐ Taken together, these data indicate that TMEM16F is required for platelet
microparticle formation and that TMEM16F-dependent platelet microparticles
subsequently play a role in procoagulant activity that can contribute to thrombosis.