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Published on: August 13, 2015
Mice Lacking PECAM-1 and Ceacam1 Have Enhanced Platelet Secretion and Thrombus Growth: Novel Link with PAR4
Fahd A Kuriri1,2, Genia Burchall1, Fehaid Alanazi1,3
1Thrombosis and Vascular Diseases Laboratory, School of Health and Biomedical Sciences, STEM College, RMIT University, Bundoora, Victoria, Australia.
Abstract:
The Ig-ITIM bearing receptors, PECAM-1 and CEACAM1, have been shown net negative regulators of platelet-collagen interactions and hemiITAM signaling pathways. In this study, a double knockout (DKO) mouse was developed with deleted PECAM-1 and CEACAM1 to study their combined contribution in platelet activation by glycoprotein VI, C-type lectin-like receptor 2, protease activated receptor (PAR4), ADP purinergic receptors, and thromboxane receptor (TP) A2 pathways. In addition, their collective contribution was examined in thrombus formation under high shear and microvascular thrombosis using in vivo models. DKO platelets responded normally to ADP purinergic receptors and the TP A2 pathway. However, DKO platelets released significantly higher amounts of P-selectin compared with hyper-responsive Pecam-1-/- or Ceacam1-/- versus wild-type (WT) upon stimulation with collagen-related peptide or rhodocytin. In contrast, DKO platelets showed increased amounts of P-selectin exposure upon stimulation with PAR4 agonist peptide or thrombin but not Pecam-1-/- , Ceacam1-/- , or WT platelets. Blockade of phospholipase C (PLC) or Rho A kinase revealed that DKO platelets enhanced α-granule release via PAR4/Gαq/PLC signaling without crosstalk with Src/Syk or G12/13 signaling pathways. Severely delayed clot retraction in vitro was observed in DKO phenotype. The DKO model revealed a significant increase in thrombus formation compared with the hyper-responsive Ceacam1-/- or Pecam-1-/- versus WT phenotype. DKO platelets have similar glycoprotein surface expression compared with Pecam-1-/- , Ceacam1-/- , and WT platelets. This study demonstrates that PECAM-1 and CEACAM1 work in concert to negatively regulate hemiITAM signaling, platelet-collagen interactions, and PAR4 Gαq protein- coupled signaling pathways. Both PECAM-1 and CEACAM1 are required for negative regulation of platelet activation and microvascular thrombosis in vivo.
Insights
Platelet receptors PECAM-1 and CEACAM1 negatively regulate collagen and PAR4 signaling. Deleting both enhances platelet activation and thrombus formation, revealing their combined role in thrombosis.
Area of Science:
- Immunology
- Hematology
- Cellular Signaling
Background:
- Platelet activation is crucial for hemostasis but also implicated in thrombosis.
- PECAM-1 and CEACAM1 are inhibitory receptors regulating platelet function.
- Their combined role in platelet activation pathways and thrombosis is not fully understood.
Purpose of the Study:
- To investigate the combined role of PECAM-1 and CEACAM1 in regulating platelet activation.
- To examine their collective contribution to thrombus formation in vivo.
Main Methods:
- Generated a double knockout (DKO) mouse lacking PECAM-1 and CEACAM1.
- Assessed DKO platelet responses to various agonists (collagen-related peptide, PAR4, ADP, TP A2).
- Evaluated thrombus formation under high shear and in microvascular thrombosis models.
Main Results:
- DKO platelets showed hyper-responsiveness to collagen-related peptide and rhodocytin, releasing more P-selectin.
- DKO platelets exhibited enhanced P-selectin exposure upon PAR4 stimulation, mediated by Gαq/PLC signaling.
- DKO mice displayed significantly increased thrombus formation in vivo compared to single knockouts or WT.
Conclusions:
- PECAM-1 and CEACAM1 cooperate to negatively regulate platelet activation via hemiITAM and PAR4 signaling.
- Both receptors are essential for controlling platelet responses to collagen and thrombin.
- Combined deficiency of PECAM-1 and CEACAM1 leads to heightened platelet activation and increased microvascular thrombosis.
