The role of the G protein-coupled receptor ChemR23 in cardiovascular inflammation

Cardiovascular inflammation is a biological process characterized by the immune cell response to harmful stimuli within the cardiovascular system, wherein homeostasis is reestablished by neutralizing the insult and promoting the repair of the damaged tissue. However, a failure in the resolution of inflammation can lead to a maladaptive inflammatory response, contributing to the development of cardiovascular diseases such as aortic valve stenosis (AVS) and atherosclerosis, as well as vascular complications such as intimal hyperplasia. Omega-3 polyunsaturated fatty acids (n-3 PUFA) serve as a substrate for the generation of a group of bioactive specialized pro-resolving lipid mediators (SPMs) that mediate the resolution of inflammation. The SPM resolvinE1 (RvE1) derived from eicosapentaenoic acid reduces cardiovascular inflammation by signaling through the G protein-coupled receptor ChemR23. The aims of the current thesis were to establish the role of ChemR23 and its activation via RvE1 in the context of AVS, intimal hyperplasia, and atherosclerotic plaque calcification.

In Article I, we reported for the first time that n-3 PUFA as well as RvE1 were decreased in calcified regions compared with non-calcified parts of human aortic valves from AVS patients. Stimulation of valve interstitial cells with RvE1 reduced cell calcification after phosphate supplementation. Genetic deletion of ChemR23 in apolipoprotein E-deficient (Apoe-/-) mice exacerbated hemodynamic and histological signs of AVS. Moreover, presence of the Fat-1 transgene (Fat-1tg), which enables the endogenous synthesis of n-3 PUFA, halted AVS progression by reducing hemodynamic and histological signs of AVS, and promoting M2 macrophage polarization in the mouse aortic valves. Importantly, Fat-1tg effects were only detected in the presence of ChemR23, suggesting that the beneficial actions were mediated via RvE1 signaling through ChemR23.

In Article II, we showed that ChemR23 genetic deletion in mice increased the development of intimal hyperplasia after a carotid ligation. In vitro, ChemR23-deficient vascular smooth muscle cells (VSMCs) exhibited lower proliferation compared with ChemR23 wild-type VSMC. Moreover, ChemR23-deficient macrophages presented a more pro-inflammatory phenotype. Finally, conditioned media transfer from ChemR23-deficient macrophages to VSMCs increased VSMC proliferation compared with conditioned media from ChemR23 wild-type macrophages. This points to a dual effect of ChemR23 in the vasculature depending on the degree of inflammation, with ChemR23 directly stimulating VSMC proliferation and at the same time suppressing macrophage-induced VSMC proliferation.

In Article III, we demonstrated that the presence of Fat-1tg increased n-3 PUFA incorporation in atherosclerotic plaques of Apoe-/- mice, and decreased atherosclerotic plaque calcification. As also seen in aortic valves, ChemR23 deletion increased muri ne atherosclerotic plaque calcification independently of the local levels of n-3 PUFA, demonstrating the importance of a correct signaling through ChemR23. Moreover, in atherosclerotic plaques of Fat-1tgxApoe-/- mice, the protein expression of the M2-macrophage marker arginase-1 was increased and ChemR23 deletion enhanced the levels of tissue non-specific alkaline phosphatase (TNAP).

In conclusion, the results included in this thesis demonstrate the importance of ChemR23 signaling in cardiovascular inflammation and provide novel mechanistic insights into the beneficial actions of n-3 PUFA and its downstream SPM RvE1 in cardiovascular diseases.

List of scientific papers

I. Artiach G, Carracedo M, Plunde O, Wheelock CE, Thul S, Sjövall P, Franco- Cereceda A, Laguna-Fernandez A, Arnardottir H, Bäck M. Omega-3 Polyunsaturated Fatty Acids Decrease Aortic Valve Disease through the Resolvin E1 and ChemR23 Axis. Circulation. 2020;142:776-789.
https://doi.org/10.1161/CIRCULATIONAHA.119.041868

II. Artiach G*, Carracedo M*, Clària J, Laguna-Fernandez A, Bäck M. Opposing Effects on Vascular Smooth Muscle Cell Proliferation and Macrophage-induced Inflammation Reveal a Protective Role for the Proresolving Lipid Mediator Receptor ChemR23 in Intimal Hyperplasia. Front Pharmacol. 2018;9:1327. *Both authors contributed equally.
https://doi.org/10.3389/fphar.2018.01327

III. Artiach G, Carracedo M, Sjövall P, Arnadottir H, Laguna-Fernandez A, Bäck M. Inhibition of atherosclerotic plaque calcification by Omega-3 polyunsaturated fatty acids involves the resolvin E1 receptor ChemR23, macrophage polarization and alkaline phosphatase alteration. [Manuscript]