Abstract-Preis der Segnitz-Ackermann-Stiftung:Olfactory receptor 2 drives abdominal aortic aneurysm by promoting CX3CR1-mediated monocyte recruitment

Clin Res Cardiol (2024). DOI 10.1007/s00392-024-02526-y
P. Schelemei (Köln)¹, F. S. R. Picard (Köln)¹, H. Nemade (Köln)¹, D. Mehrkens (Köln)², Y. Park (Köln)¹, S. Grimm (Köln)², K. Tinaz (Köln)², M. Orecchioni (Augusta)³, J. Roy (Solna)⁴, M. L. Liljeqvist (Solna)⁴, M. Wagenhäuser (Düsseldorf)⁵, H. Schelzig (Düsseldorf)⁵, S. Baldus (Köln)⁶, M. Mollenhauer (Köln)², H. Winkels (Köln)^2
1Universitätsklinikum Köln Klinik III für Innere Medizin - Experimentelle Kardiologie Köln, Deutschland; ²Herzzentrum der Universität zu Köln Klinik III für Innere Medizin Köln, Deutschland; ³Immunology Center of Georgia Augusta, USA; ⁴Karolinska Institutet Solna, Schweden; ⁵Universitätsklinikum Düsseldorf Klinik für Gefäß- und Endovaskularchirurgie Düsseldorf, Deutschland; ⁶Herzzentrum der Universität zu Köln Klinik für Kardiologie, Angiologie, Pneumologie und Internistische Intensivmedizin Köln, Deutschland

Background:
Abdominal aortic aneurysm (AAA) is a chronic, age-associated disease defined by localized enlargement of the aorta above 3.5cm or 50% of its original diameter. Pathomechanistically, smooth muscle cell death, fibrosis, and inflammation contribute to maladaptive vascular remodelling. Macrophage responses are important contributors to the progression of AAA, yet the underlying mechanisms and mediators are poorly understood. The olfactory receptor 2 (Olfr2) is a G-protein coupled receptor involved in mediating the sense of smelling. Extra-nasal expression of Olfr2 has been identified on vascular macrophages, but its role in AAA-associated macrophages responses is unknown.

Methods & Results:
We performed in-silico micro array analysis of 246 abdominal aorta samples with and without thrombus formation of 76 patients as well as 13 controls and detected significantly increased expression of OR6A2 (the human ortholog of Olfr2) in AAA with thrombus formation compared to control tissue. OR6A2 expression was confirmed by immunofluorescence exclusively on 30% of macrophages residing in human AAA tissue (n=3).
We next aimed to investigate the dynamics of Olfr2 expression on aortic mouse leukocytes in the porcine pancreatic elastase (PPE) infusion model. Applying a 27 marker panel and spectral flow cytometry enabled us to detect 20 unique immune cell cluster. Olfr2 was specifically expressed by aortic MHCII+ CCR2low monocytes and macrophages at day 7 post AAA induction, while Olfr2 expression levels were reduced to baseline levels at day 28.
To study the functional role of Olfr2 in AAA, we induced AAA formation in Olfr2-deficient (KO) and WT control mice and observed a marked protection from aneurysm formation by ultrasound analysis in KO mice. Histological and immunofluorescence analysis further revealed reduced elastin degradation and preservation of smooth muscle cells in KO mice. Moreover, spectral flow cytometry demonstrated a marked reduction of aortic leukocytes, particularly of monocytes and macrophages, at days 7 and 28 post AAA induction in KO mice. This coincided with reduced expression of the chemokine receptor CX3CR1 on Ly6Chigh monocytes in the blood of KO mice already at baseline which was aggravated at day 7.
To test whether reduced CX3CR1 expression on monocytes is of functional relevance, we isolated monocytes from the bone marrow of WT and KO mice for a chemotaxis assay. Olfr2-deficient monocytes showed impaired migration towards the CX3CR1 ligand CX3CL1. We confirmed these findings in-vivo in WT recipient mice, which have been adoptively transferred with WT and KO monocytes labeled with different cell tracker dyes at day 3 post PPE. FACS analysis of aortic leukocytes 24h post injection revealed reduced infiltration of injected KO monocytes compared to WT. 
Finally, pharmacological modulation of Olfr2 in AAA formation was assessed. Injection with the Olfr2 antagonist citral reduced aortic dilation and macrophage content in WT mice, whereas treatment with the Olfr2 agonist octanal aggravated AAA formation and aortic inflammation.

Conclusion:
We highlight a novel mechanism by which Olfr2 drives the development of AAA by promoting CX3CR1 mediated monocyte recruitment.