The C-Type Lectin Receptor CD93 Regulates Platelet Activation and Surface Expression of the Protease Activated Receptor 4

Abstract

Background The C-type lectin receptor CD93 is a single pass type I transmembrane glycoprotein involved in inflammation, immunity, and angiogenesis. This study investigates the role of CD93 in platelet function. CD93 knockout (KO) mice and wild-type (WT) controls were compared in this study.

Methods Platelet activation and aggregation were investigated by flow cytometry and light transmission aggregometry, respectively. Protein expression and phosphorylation were analyzed by immunoblotting. Subcellular localization of membrane receptors was investigated by wide-field and confocal microscopy.

Results The lack of CD93 in mice was not associated to any evident bleeding defect and no alterations of platelet activation were observed upon stimulation with thromboxane A2 analogue and convulxin. Conversely, platelet aggregation induced by stimulation of the thrombin receptor PAR4 was significantly reduced in the absence of CD93. This defect was associated with a significant reduction of α-granule secretion, integrin αIIbβ3 activation, and protein kinase C (PKC) stimulation. Resting WT and CD93-deficient platelets expressed comparable amounts of PAR4. However, upon stimulation with a PAR4 activating peptide, a more pronounced clearance of PAR4 from the platelet surface was observed in CD93-deficient platelets compared with WT controls. Confocal microscopy analysis revealed a massive movement of PAR4 in cytosolic compartments of activated platelets lacking CD93. Accordingly, platelet desensitization following PAR4 stimulation was more pronounced in CD93 KO platelets compared with WT controls.

Conclusion These results demonstrate that CD93 supports platelet activation triggered by PAR4 stimulation and is required to stabilize the expression of the thrombin receptor on the cell surface.

Keywords

blood platelets - C-type lectin receptors - G protein-coupled receptor - platelet activation - thrombin

Authors' Contribution

S.M.G.T., M.V., S.R., I.C.: performed the experiments, analyzed the data, and edited the manuscript; F.G., M.O.: provided vital reagents, discussed the results, and edited the manuscript; M.T., G.F.G.: designed the study, analyzed and interpreted the data, and drafted the manuscript.

Publikationsverlauf

Eingereicht: 18. April 2023

Angenommen: 04. September 2023

Accepted Manuscript online:
05. September 2023

Artikel online veröffentlicht:
30. September 2023

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