The endocannabinoid anandamide mediates anti‐inflammatory effects through activation of NR4A nuclear receptors

Abstract

Endocannabinoids are lipid mediators, which elicit complex biological effects that extend beyond the central nervous system. The tissue concentrations of endocannabinoids increase in atherosclerosis, and for the endocannabinoid N-arachidonoyl-ethanolamine (anandamide; AEA), this has been linked to a potential anti-inflammatory function. Focusing on vascular smooth muscle cells, we set out to determine the underlying mechanism. Confirming previous results, AEA pretreatment attenuated the cytokine-mediated induction of inflammatory gene expression like CCL2. Unexpectedly, this effect was also observed in preparations of cannabinoid receptor (CB) 1 and 2 knockout mice and after pertussis toxin treatment but was restricted to fairly high concentrations of the lipid. The anti-inflammatory effect of AEA required preincubation, suggesting an effect through gene induction. RNA-seq revealed that AEA caused an increased expression of the nuclear receptors NR4A1 and NR4A2. Knockdown of these receptors blocked the AEA-mediated anti-inflammatory effect. Conversely, NR4A agonists (CsnB, C-DIM12) also attenuated inflammatory gene expression. Microscale thermophoresis, NMR-spectroscopy, and Gal4 reporter gene assays confirmed the binding of AEA to NR4A. Moreover, AEA caused an NR4A-dependent recruitment of the nuclear corepressor NCoR1 to the CCL2 promotor. Finally, AEA also elicited anti-inflammatory effects in the vascular organ culture of mice, which was not observed after the genetic deletion of NR4A1 or NR4A2.AEA elicits a specific anti-inflammatory response in vascular smooth muscle cells through activation of NR4A. AEA analogous with high potential to bind NR4A receptors might act as novel anti-inflammatory drugs.