The endocannabinoid anandamide activates pro‐resolving pathways in human primary macrophages by engaging both CB2 and GPR18 receptors

Resolution of inflammation is the cellular and molecular process that protects from widespread and uncontrolled inflammation and restores tissue function in the aftermath of acute immune events. This process is orchestrated by specialized pro‐resolving mediators (SPM), a class of bioactive lipids able to reduce immune activation and promote removal of tissue debris and apoptotic cells by macrophages. Although SPMs are the lipid class that has been best studied for its role in facilitating the resolution of self‐limited inflammation, a number of other lipid signals, including endocannabinoids, also exert protective immunomodulatory effects on immune cells, including macrophages. These observations suggest that endocannabinoids may also display pro‐resolving actions. Interestingly, the endocannabinoid anandamide (AEA) is not only known to bind canonical type 1 and type 2 cannabinoid receptors (CB1 and CB2) but also to engage SPM‐binding receptors such as GPR18. This suggests that AEA may also contribute to the governing of resolution processes. In order to interrogate this hypothesis, we investigated the ability of AEA to induce pro‐resolving responses by classically‐activated primary human monocyte‐derived macrophages (MoDM). We found that AEA, at nanomolar concentration, enhances efferocytosis in MoDMs in a CB2‐ and GPR18‐dependent manner. Using lipid mediator profiling, we also observed that AEA modulates SPM profiles in these cells, including levels of resolvin (Rv)D1, RvD6, maresin (MaR)2, and RvE1 in a CB2‐dependent manner. AEA treatment also modulated the gene expression of SPM enzymes involved in both the formation and further metabolism of SPM such as 5‐lipoxygenase and 15‐Prostaglandin dehydrogenase. Our findings show, for the first time, a direct effect of AEA on the regulation of pro‐resolving pathways in human macrophages. They also provide new insights into the complex interactions between different lipid pathways in activation of pro‐resolving responses contributing to the reestablishment of homeostasis in the aftermath of acute inflammation. N‐arachidonoylethanolamine (AEA or anandamide) is one of the two main endocannabinoid (eCB) lipids and possesses well‐described immunomodulatory properties. This lipid is able to bind both type‐2 endocannabinoid receptor (CB2) and GPR18 – i.e., the receptor of resolving D2 (RvD2), a specialized pro‐resolving mediator (SPM). This suggests an interaction between these two lipid systems in modulating inflamm