HAuCl4, Putative General Aquaporins Blocker, Reduces Platelet Spreading, Filopodia Formation, Procoagulant Response, and Thrombus Formation Under Flow

Tomasz Misztal; Agata Golaszewska; Justyna Branska-Januszewska; Natalia Marcinczyk; Ewa Chabielska; Marian Tomasiak; Tomasz Rusak

doi:10.3389/fphys.2020.01025

HAuCl₄, Putative General Aquaporins Blocker, Reduces Platelet Spreading, Filopodia Formation, Procoagulant Response, and Thrombus Formation Under Flow

Front Physiol. 2020 Aug 21:11:1025. doi: 10.3389/fphys.2020.01025. eCollection 2020.

Authors

Tomasz Misztal¹, Agata Golaszewska¹, Justyna Branska-Januszewska², Natalia Marcinczyk³, Ewa Chabielska³, Marian Tomasiak¹, Tomasz Rusak¹

Affiliations

¹ Department of Physical Chemistry, Medical University of Bialystok, Bialystok, Poland.
² Department of Biology, Medical University of Bialystok, Bialystok, Poland.
³ Department of Biopharmacy, Medical University of Bialystok, Bialystok, Poland.

Abstract

Background: Recent studies indicate that aquaporin (AQP) water channels have a regulatory function in human platelet secretion and in procoagulant response of murine platelets. However, the engagement of AQPs in morphological changes, procoagulant response, and thrombus formation in human blood has never been investigated. Methods: Confocal microscopy was used to study platelet spreading, filopodia formation, ballooning, and thrombus formation under flow. Flow cytometry was utilized to assess platelet phosphatidylserine (PS) exposure and microparticles shedding. Kinetics of clot formation in vitro was evaluated by thromboelastometry. Mouse model of ferric chloride (III) (FeCl₃)-induced thrombosis was used to investigate thrombus formation in vivo. Results: We found that chloroauric(III) acid (HAuCl₄), a classical AQP inhibitor (10-100 μM), reduced spreading of human platelets on collagen-coated surfaces and inhibited filopodia formation in a fluid phase. Under flow conditions, HAuCl₄ (100 μM) attenuated thrombi growth on collagen, platelet secretion, and PS exposure. Thrombus formation was restored by the addition of exogenous adenosine diphosphate (ADP). Collagen-evoked platelet procoagulant response (evaluated as PS exposure, shedding of microparticles, platelet-dependent thrombin generation, and membrane ballooning) was distinctly reduced by HAuCl₄ (25-200 μM), as well as the dynamics of clot formation. In mouse model of thrombosis, reduction of surface of PS-positive cells within thrombus was observed in the presence of HAuCl₄ (1-10 mg/kg). Conclusion: These results suggest that in human platelets AQPs are crucial for agonist-evoked morphological changes, thrombus formation under flow, and in development of procoagulant response. Antithrombotic effect in vivo suggests that nontoxic inhibitors of AQPs may be considered as potential candidates for a novel class of antiplatelet drugs.

Keywords: aquaporins; mouse model of thrombosis; phosphatidylserine exposure; platelets; procoagulant activities; thrombus formation.