Buka, Richard John 
ORCID: 0000-0002-2738-6236
(2025).
Mechanisms of platelet activation in vaccine-induced thrombotic thrombocytopenia and heparin-induced thrombocytopenia.
University of Birmingham.
Ph.D.
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Buka2025PhD.pdf
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Abstract
Vaccine-induced immune thrombocytopenia with thrombosis (VITT) and heparin-induced thrombocytopenia (HIT) are disorders mediated by antibodies against platelet factor 4 (PF4), an abundant, positively charged, platelet-derived chemokine. In VITT, immune complexes consisting of PF4 and anti-PF4 antibodies form and activate platelets through the low-affinity immune receptor FcγRIIA whereas in HIT, the immune complexes also contain the anticoagulant heparin. Both heparin and PF4 are known to activate platelets independently. Heparin acts through platelet endothelial aggregation receptor 1 (PEAR1) but the mechanism of activation by PF4 is unknown.
The aims of this thesis were therefore to firstly ascertain the mechanism of platelet activation by PF4 and then to investigate whether PF4 and heparin play a role in directly activating platelets in VITT and HIT. A third, separate aim was to investigate endothelial activation in VITT and HIT to elucidate any differences that may contribute to the common occurrence of cerebral venous sinus thrombosis in VITT but not in HIT.
Herein, PF4 is shown to lead to platelet aggregation, α-granule release, and spreading in a dose-dependent manner starting at 10 μg/mL (1.28 μM). This occurs through binding to the thrombopoietin receptor c-Mpl and signalling through Janus kinase 2 (JAK2), a process that can be blocked by the JAK2 inhibitor, ruxolitinib, and a c-Mpl blocking antibody. Next, it was shown that blockade of the PF4-c-Mpl interaction and downstream signalling could reduce platelet aggregation to sera and isolated immunoglobulin G from patients with VITT. There was no such clear effect with HIT sera and antibodies. However, blockade of the heparin-PEAR1 interaction with an inhibitory anti-PEAR1 nanobody did lead to reduction in platelet aggregation and α-granule release to HIT sera and antibodies. These findings may have therapeutic relevance when developing targeted therapies for use in these disorders. Endothelial cell work showed no direct induction of endothelial activation by VITT or HIT sera or antibodies, nor did they lead to platelet recruitment.
| Type of Work: | Thesis (Doctorates > Ph.D.) | |||||||||||||||
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| Award Type: | Doctorates > Ph.D. | |||||||||||||||
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| Licence: | All rights reserved | |||||||||||||||
| College/Faculty: | Colleges > College of Medicine and Health | |||||||||||||||
| School or Department: | Department of Cardiovascular Sciences | |||||||||||||||
| Funders: | British Heart Foundation, National Institute for Health Research | |||||||||||||||
| Subjects: | Q Science > QP Physiology R Medicine > RB Pathology R Medicine > RC Internal medicine R Medicine > RM Therapeutics. Pharmacology |
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| URI: | http://etheses.bham.ac.uk/id/eprint/16248 |
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