Contractility defects hinder glycoprotein VI-mediated platelet activation and affect platelet functions beyond clot contraction

Martin Kenny; Alice Y Pollitt; Smita Patil; Dishon W Hiebner; Albert Smolenski; Natalija Lakic; Robert Fisher; Reema Alsufyani; Sebastian Lickert; Viola Vogel; Ingmar Schoen

doi:10.1016/j.rpth.2024.102322

Contractility defects hinder glycoprotein VI-mediated platelet activation and affect platelet functions beyond clot contraction

Res Pract Thromb Haemost. 2024 Jan 22;8(1):102322. doi: 10.1016/j.rpth.2024.102322. eCollection 2024 Jan.

Authors

Martin Kenny^{1

2}, Alice Y Pollitt³, Smita Patil^{1

2}, Dishon W Hiebner^{1

2}, Albert Smolenski⁴, Natalija Lakic⁵, Robert Fisher⁵, Reema Alsufyani⁵, Sebastian Lickert⁶, Viola Vogel⁶, Ingmar Schoen^{1

2}

Affiliations

¹ School of Pharmacy and Biomolecular Sciences, Royal College of Surgeons in Ireland, Dublin, Ireland.
² Irish Centre for Vascular Biology, Royal College of Surgeons in Ireland, Dublin, Ireland.
³ School of Biological Sciences, University of Reading, Reading, United Kingdom.
⁴ School of Medicine, Conway Institute, University College Dublin, Belfield, Dublin, Ireland.
⁵ School of Medicine, Royal College of Surgeons in Ireland, Dublin, Ireland.
⁶ Department of Health Sciences and Technologies, ETH Zurich, Zurich, Switzerland.

Abstract

Background: Active and passive biomechanical properties of platelets contribute substantially to thrombus formation. Actomyosin contractility drives clot contraction required for stabilizing the hemostatic plug. Impaired contractility results in bleeding but is difficult to detect using platelet function tests.

Objectives: To determine how diminished myosin activity affects platelet functions, including and beyond clot contraction.

Methods: Using the myosin IIA-specific pharmacologic inhibitor blebbistatin, we modulated myosin activity in platelets from healthy donors and systematically characterized platelet responses at various levels of inhibition by interrogating distinct platelet functions at each stage of thrombus formation using a range of complementary assays.

Results: Partial myosin IIA inhibition neither affected platelet von Willebrand factor interactions under arterial shear nor platelet spreading and cytoskeletal rearrangements on fibrinogen. However, it impacted stress fiber formation and the nanoarchitecture of cell-matrix adhesions, drastically reducing and limiting traction forces. Higher blebbistatin concentrations impaired platelet adhesion under flow, altered mechanosensing at lamellipodia edges, and eliminated traction forces without affecting platelet spreading, α-granule secretion, or procoagulant platelet formation. Unexpectedly, myosin IIA inhibition reduced calcium influx, dense granule secretion, and platelet aggregation downstream of glycoprotein (GP)VI and limited the redistribution of GPVI on the cell membrane, whereas aggregation induced by adenosine diphosphate or arachidonic acid was unaffected.

Conclusion: Our findings highlight the importance of both active contractile and passive crosslinking roles of myosin IIA in the platelet cytoskeleton. They support the hypothesis that highly contractile platelets are needed for hemostasis and further suggest a supportive role for myosin IIA in GPVI signaling.

Keywords: blebbistatin; hemostasis; nonmuscle myosin type IIA; thrombosis; traction.