Melanocortin receptors are novel effectors of macrophage efferocytosis and promote resolution of inflammation

Trinidad Montero-Melendez1, Thomas EN Jonassen3, Michael P Seed2, Mauro Perretti1. 1Biochemical Pharmacology, William Harvey Research Institute, School of Medicine and Dentistry, Queen Mary University of London, London, UK, 2Experimental Medicine and Rheumatology, William Harvey Research Institute, School of Medicine and Dentistry, Queen Mary University of London, London, UK, 3Biochemical Sciences, University of Copenhagen, Copenhagen, Denmark/

Inflammation is a defensive response of the body against pathogens and injury in which immune cells and soluble factors take part to neutralize the injurious agent and initiate tissue repair. However, a loss of regulation of these mechanisms can prevent the final resolution of the inflammatory process leading to a chronic status. The macrophage plays an orchestrating role in these settings, and its dynamic activation can be modulated by several mediators and receptors. Our group has focused on Melanocortin (MC) receptors (adenylate-cyclase associated GPCR) which conveys the anti-inflammatory actions of melanocortin peptides. We report here that MC receptor agonists αMSH (natural agonist), AP214 (synthetic peptide derivative) and AP1189 (small molecule) evoke pro-resolving responses.

Using biogel-elicited peritoneal macrophages we have observed increased phagocytosis of zymosan particles (given at ratio 1:10, macrophage to zymosan, 0.5x106 macrophages per well) when cells were treated with MC agonists (36% phagocytic cells for vehicle and 69% for 1nM AP1189 treated cells). Furthermore, they also increased phagocytosis of apoptotic neutrophils (process of efferocytosis) in vitro (16%phagocytic cells for vehicle and 22% for 1nM AP1189 treated cells) and in vivo (44% phagocytic cells for vehicle and 54% for AP1189 -800μg/kg- treated mice). This pro-efferocytic effect was mainly driven by MC3, and to a less extent by MC1, as demonstrated using macrophages from MC1-/- and MC3-/- mice. In addition, cells from these mice presented a mild defect in phagocytosis and efferocytosis, with basal responses being reduced vs. wild-type cells. To assess the pro-resolving nature of MC receptor activation, we tested AP1189 in vivo, administering it (1 mg i.p.) at the peak of an acute peritonitis (12h post-zymosan) observing marked improvement of resolution indices, for AP1189-treated and control mice (PBS treated), respectively:

T50 (time when max neutrophil numbers are reduced by 50%) was 21h and 38h;

Ri (time from maximal response to T50) was 9h and 26h.

These effects of AP1189 were accompanied by significant reduction in inflammatory monocytes (-42% at 22h; 2.4x106 and 1.4x106 cells in vehicle and AP1189 treated mice respectively) and PGE2 levels (-80% at 44h; 413 and 88 pg/ml in vehicle and AP1189 treated mice respectively), together with enhanced extent of macrophages engulfed with apoptotic neutrophils. We conclude MC receptor can be successfully harnessed – using diverse pharmacological approaches - to induce pro-resolving effects with a potential beneficial application in chronic inflammatory diseases.

This project was funded by an Action Pharma/WHR Foundation collaboration.