Hydroxychloroquine inhibits hemolysis-induced arterial thrombosis ex vivo and improves lung perfusion in hemin-treated mice

Joshua H Bourne; Gina Perrella; Juma El-Awaisi; Lauren V Terry; Veronika Tinkova; Rebecca L Hogg; Poppy Gant; Beata Grygielska; Neena Kalia; Dean Kavanagh; Alexander Brill; Jordan D Dimitrov; Steve P Watson; Julie Rayes

doi:10.1016/j.jtha.2024.04.008

Hydroxychloroquine inhibits hemolysis-induced arterial thrombosis ex vivo and improves lung perfusion in hemin-treated mice

J Thromb Haemost. 2024 Apr 24:S1538-7836(24)00226-5. doi: 10.1016/j.jtha.2024.04.008. Online ahead of print.

Affiliations

¹ Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, United Kingdom.
² Institute of Immunology and Immunotherapy, University of Birmingham, Birmingham, United Kingdom.
³ Centre de Recherche des Cordeliers, Institut national de la santé et de la recherche médicale, Sorbonne Université, Université Sorbonne Paris-Cité, Université Paris Descartes, Université Paris Diderot, Paris, France.
⁴ Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, United Kingdom; Centre of Membrane Proteins and Receptors (COMPARE), Universities of Birmingham and Nottingham, the Midlands, United Kingdom.
⁵ Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, United Kingdom. Electronic address: j.rayes@bham.ac.uk.

PMID: 38670315
DOI: 10.1016/j.jtha.2024.04.008

Abstract

Background: Free labile hemin acts as a damage-associated molecular pattern during acute and chronic hemolysis and muscle injury, supporting platelet activation and thrombosis.

Objectives: To investigate the anti-thrombotic potential of hydroxychloroquine on hemolysis-induced platelet activation and arterial thrombosis.

Methods: The effect of hydroxychloroquine on hemin-induced platelet activation and hemolysis-induced platelet recruitment and aggregation was measured in washed platelets and hemolyzed blood, respectively. Its effect on ferric-chloride (FeCl₃)-induced arterial thrombosis and lung perfusion following hemin injection was assessed in wild-type mice.

Results: Erythrocyte lysis and endothelial cell activation cooperatively supported platelet aggregation and thrombosis at arterial shear stress. This thrombotic effect was reversed by hydroxychloroquine. In a purified system, hydroxychloroquine inhibited platelet build-up on immobilized von Willebrand factor in hemolyzed blood without altering initial platelet recruitment. Hydroxychloroquine inhibited hemin-induced platelet activation and phosphatidylserine exposure independently of reactive oxygen species generation. In the presence of hemin, hydroxychloroquine did not alter glycoprotein VI shedding but reduced C-type-lectin-like-2 expression on platelets. In vivo, hydroxychloroquine reversed pulmonary perfusion decline induced by exogenous administration of hemin. In arterial thrombosis models, hydroxychloroquine inhibited ferric-chloride-induced thrombosis in the carotid artery and reduced von Willebrand factor accumulation in the thrombi.

Conclusion: Hydroxychloroquine inhibited hemolysis-induced arterial thrombosis ex vivo and improved pulmonary perfusion in hemin-treated mice, supporting a potential benefit of its use as an adjuvant therapy in hemolytic diseases to limit arterial thrombosis and to improve organ perfusion.

Keywords: hemin; hemolysis; hydroxychloroquine; platelets; thrombosis.