Background: The ubiquitously expressed small GTPases RalA and RalB are involved in cell proliferation and tumour metastasis. We reported that RalA/B conditional double knockout (RalAB DKO) in mouse platelets resulted in near complete defect in P-selectin externalisation on platelets upon activation, with no other significant secretion defect.

Aims: To address whether platelet-expressed Ral GTPases, through regulation of platelet surface expression of P selectin, control venous thrombosis in vivo in mice.

Methods: We used an in vivo inferior vena cava ligation model of venous thrombosis in mice, and analyses of neutrophil NET formation.

Results: RalAB DKO led to ablation of venous thrombosis in mice (thrombus weight in DKO: 0.4±0.2 mg vs 15.35±3.20 mg in Wt control, n=8 in DKO and n=10 in WT, p< 0.001). Immunohistochemical staining of thrombus sections demonstrated that in DKO mice there was no NET formation, and less neutrophil infiltration than in WT mice, where strong NETosis was detected. Platelet-specific conditional knockout of SNAP23 however did form (diminished) venous thrombi, even though absence of SNAP23 resulted in complete ablation of secretion. Histological examination of SNAP23^-/- thrombi showed similar structures to WT, although diminished in size, and NETosis formation was normal, suggesting that secretion defects, particularly P-selectin externalisation, are only partially responsible for the ablated venous thrombosis in RalAB DKO mice. In addition to the defect in NETosis, we also found that P-selectin and HMGB1 did not colocalize in thrombus sections of RalAB DKO as they did in WT control mice.

Conclusions: Platelets play a critical role in venous thrombosis in a ligation model in mouse. Platelet-derived P-selectin and HMGB1 recruit and activate neutrophils and NET formation, coordinated by platelet-expressed RalA and RalB. As Rals are targets for drug development, future work is required to determine whether platelet Rals represent good novel targets in venous thrombosis management.

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ISTH Congress Abstracts - https://abstracts.isth.org/abstract/critical-role-for-platelet-ral-gtpases-in-the-regulation-of-venous-thrombosis/