Cyclophilin A is a ligand for RAGE in thrombo-inflammation.

AIMS: Cyclophilin A (CyPA) induces leucocyte recruitment and platelet activation upon release into the extracellular space. Extracellular CyPA therefore plays a critical role in immuno-inflammatory responses in tissue injury and thrombosis upon platelet activation. To date, CD147 (EMMPRIN) has been described as the primary receptor mediating extracellular effects of CyPA in platelets and leucocytes. The receptor for advanced glycation end products (RAGE) shares inflammatory and prothrombotic properties and has also been found to have similar ligands as CD147. In this study, we investigated the role of RAGE as a previously unknown interaction partner for CyPA. METHODS AND RESULTS: Confocal imaging, proximity ligation, co-immunoprecipitation, and atomic force microscopy were performed and demonstrated an interaction of CyPA with RAGE on the cell surface. Static and dynamic cell adhesion and chemotaxis assays towards extracellular CyPA using human leucocytes and leucocytes from RAGE-deficient Ager-/- mice were conducted. Inhibition of RAGE abrogated CyPA-induced effects on leucocyte adhesion and chemotaxis in vitro. Accordingly, Ager-/- mice showed reduced leucocyte recruitment and endothelial adhesion towards CyPA in vivo. In wild-type mice, we observed a downregulation of RAGE on leucocytes when endogenous extracellular CyPA was reduced. We furthermore evaluated the role of RAGE for platelet activation and thrombus formation upon CyPA stimulation. CyPA-induced activation of platelets was found to be dependent on RAGE, as inhibition of RAGE, as well as platelets from Ager-/- mice showed a diminished activation and thrombus formation upon CyPA stimulation. CyPA-induced signalling through RAGE was found to involve central signalling pathways including the adaptor protein MyD88, intracellular Ca2+ signalling, and NF-κB activation. CONCLUSION: We propose RAGE as a hitherto unknown receptor for CyPA mediating leucocyte as well as platelet activation. The CyPA-RAGE interaction thus represents a novel mechanism in thrombo-inflammation.

Myeloid But Not Endothelial Expression of the CB2 Receptor Promotes Atherogenesis in the Context of Elevated Levels of the Endocannabinoid 2-Arachidonoylglycerol.

The endocannabinoid 2-arachidonoylglycerol (2-AG) is an inflammatory mediator and ligand for the cannabinoid receptors CB1 and CB2. We investigated the atherogenic mechanisms set in motion by 2-AG. Therefore, we created two atherosclerotic mouse models with distinct cell-specific knockouts of the CB2 receptor on either myeloid or endothelial cells. These mice were treated with JZL184, resulting in elevated plasma levels of 2-AG. After a high-fat high-cholesterol diet, atherosclerotic plaques were analyzed. The atherogenic effect of 2-AG was abrogated in mice lacking myeloid expression of the CB2 receptor but not in mice lacking endothelial expression of the CB2 receptor. In vitro, treatment of human monocytes with 2-AG led to the increased production of reactive oxygen species (ROS) and IL-1ß. In conclusion, 2-AG shows an atherogenic effect in vivo, dependent on the presence of the CB2 receptor on myeloid cells. In addition, our in vitro data revealed 2-AG to promote inflammatory signalling in monocytes. 2-Arachidonoylglycerol shows an atherogenic effect that is abrogated in mice lacking myeloid expression of the CB2 receptor.