191 Interleukin-1 Alpha is Directly Activated During Haemostasis by Thrombin Cleavage

Abstract

<h3></h3> The clinical consequences of atherosclerosis are closely associated with thrombus formation following plaque rupture. Thrombin primarily cleaves fibrinogen into fibrin which forms a thrombus at sites of injury, but can also induce inflammation indirectly via PAR receptors. Chronic inflammation is a key feature of atherosclerosis that promotes both plaque growth and instability, and links between inflammation and haemostasis are widely proposed. We now present data showing that the powerful proatherogenic cytokine IL-1α is directly activated by thrombin. We find that thrombin treatment of pro-IL-1α produces a truncated form, indicating cleavage of the protein in a similar manner to its canonical calpain cleavage. Mutation of the thrombin consensus in IL-1α prevented cleavage, confirming the site. Thrombin cleavage of pro-IL-1α induced similar cytokine activity as calpain, while recombinant proteins equivalent to the thrombin- and calpain-cleaved forms had identical activity. Importantly, cleavage and activation of pro-IL-1α also occurred during coagulation of platelet-rich plasma and whole blood, suggesting physiological relevance. Again, thrombin-site mutant pro-IL-1α was uncleaved and lacked activity, proving the specific action of thrombin. Murine pro-IL-1α also requires processing for cytokine activity and again thrombin cleavage induced full activity, supporting an evolutionary conserved mechanism. Importantly, specific mutation of the site in mouse pro-IL-1α prevents thrombin cleavage, which has enabled us to generate a novel knock-in mouse to study the <i>in vivo</i> relevance of these findings. We show that thrombin cleaves and activates pro-IL-1α at a novel site that is conserved across species. This suggests that thrombin-mediated activation of IL-1α during thrombosis may directly promote inflammation. We propose that this represents an unappreciated fundamental link between the key enzyme of thrombosis and the principal cytokine within immunity. These findings have broad implications for normal physiology and the pathogenesis of diseases such as atherothrombosis.