PB0831 - Cyclophilin A in platelet-mediated thrombo-inflammation

Background: Upon platelet activation, crucial for hemostasis and thrombosis, Cyclophilin A (CypA), a damage-associated molecular pattern (DAMP) stored in platelet α-granules, is released and critically influences cardiovascular diseases. CypA contributes to the bidirectional signaling of integrin αIIbβ3, thereby influencing platelet function. However, mechanisms governing CypA recruitment, integrin binding, and release remain unexplored. Despite evidence that extracellular CypA contributes to cardiovascular events, the exact origin of extracellular CypA, especially the role of platelet-derived CypA and its interaction with immune cells, has not yet been investigated.

Aims: Our Ppiafl/flPF4Cre mouse model will address this gap, exploring platelet-derived CypA functions and the impact on immune response after tissue injury.

Methods: Methods applied include Western blotting, qPCR and standard platelet phenotyping to characterize the Ppiafl/flPF4Cre mouse model. Super-resolution microscopy and platelet-macrophage co-cultures were used to determine the influence of CypA in the context of thrombo-inflammation.

Results: Initial experiments with Ppiafl/flPF4Cre mice confirmed CypA deletion specifically in platelets at the transcriptomic and protein level, but not in vascular smooth muscle cells. Importantly, platelet count remained unaffected. Platelet activation was significantly impaired in Ppiafl/fl PF4Cre mice following thrombin, ADP and CRP-XL stimulation, and a trend towards reduced thrombus volume under shear stress, without changes in thrombus surface area, highlighted the crucial role of intracellular CypA in platelet function. Using super-resolution technology, we furthermore visualized CypA not only intracellularly, but also extracellularly on the surface of resting murine platelets. Preliminary experiments indicated an increased expression of inflammatory genes in macrophages after stimulation with extracellular CypA. Similar effects were observed in co-cultures with resting and thrombin-activated platelets, suggesting a contribution of platelet-surface CypA in thrombo-inflammation and emphasizing its potential significance in tissue injury.

Conclusion(s): In summary, our Ppiafl/fl PF4Cre mouse model confirms the crucial role of platelet-derived CypA in platelet function and thrombus formation, highlighting its potential contribution to thrombo-inflammation and tissue injury.