Abstract Number: OC 56.2
Meeting: ISTH 2022 Congress
Background: RalA and RalB are small GTPases which have important functions in cell proliferation and tumour metastasis but their exact roles in thrombosis are less clear. We reported previously that platelet restricted deletion of RalA and RalB (RalAB-DKO) in mice resulted in near complete defect in P-selectin externalisation upon activation but no other significant secretion defects were detected. We also showed that RalAB-DKO resulted in near abolishment of thrombus formation.
Aims: We caught to investigate the underlying mechanism how deletion of RalAB resulted in near complete inhibition of venous thrombosis in mice.
Methods: In vitro NETosis was initiated in cultured mouse neutrophils by adding platelets and their releasates. NET+ cells were detected with confocal microscopy using anti-CitH3. Thrombus sections were immunohistochemically stained and imaged as detailed in the figure legends.
Results: Confocal microscopy showed that in the WT mice P-selectin colocalised with HMGB1 in the thrombi to form a multi-layer scaffolding structure to stabilise the thrombi while P-selectin and HMGB1 were completely separated in RalAB-DKO (Fig 1), indicating that platelet RalAB are required for both NETosis and interaction of P-selectin-HMGB1 during venous thrombosis. In an in vitro NETosis assay WT platelet releasates initiated NETosis in 27.83% neutrophils compared to only 1.99% neutrophils in RalAB-DKO platelets. But 17.38% neutrophils were NET positive when RalAB-DKO platelet releasates were substituted with WT platelet releasates, partially restoring the NETosis of RalAB-DKO platelets (Fig2).
Conclusion(s): Our data suggest that activated platelets secrete signalling factors to initiate NETosis. It has been reported that deletion of platelet HMGB1 resulted in decreased venous thrombosis and NETosis in mice. We hypothesize that HMGB1 and P-selectin could be the signalling factors platelets secrete during flow restriction in venous thrombosis.
This work was funded by The British Heart Foundation.
To cite this abstract in AMA style:Li Y, Williams C, Amulic B, Poole A. Platelet RalA/B play pivotal roles in regulation of venous thrombosis by co-ordinating P-selectin-HMGB1 interaction [abstract]. https://abstracts.isth.org/abstract/platelet-rala-b-play-pivotal-roles-in-regulation-of-venous-thrombosis-by-co-ordinating-p-selectin-hmgb1-interaction/. Accessed February 28, 2024.
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