BS20 Macrophage-specific deletion of neuropilin-2 inhibits inflammatory signalling and attenuates atherosclerotic plaque development in apoe-deficient mice

• Published in: Heart (British Cardiac Society) Vol. 109; no. Suppl 3; p. A260
• Main Authors: Pellet-Many, Caroline, Fernando-Sayers, Jacob, Zachary, Ian
• Format: Journal Article
• Language: English
• Published: London BMJ Publishing Group Ltd and British Cardiovascular Society 02.06.2023; BMJ Publishing Group LTD
• Subjects: Atherosclerosis; Basic Science; Bone marrow; Kinases; Macrophage; Neuropilin
• ISSN: 1355-6037; 1468-201X
• DOI: 10.1136/heartjnl-2023-BCS.234

IntroductionAtherosclerosis is a multifaceted vascular disease involving several cell types and characterised by arterial wall inflammation resulting from the subendothelial retention, and modification, of lipoproteins by macrophages. Macrophages are highly plastic cells that can alter their phenotype and undergo polarisation in response to cytokines or other environmental stimuli. They become pro-inflammatory in several disease settings, making them an attractive therapeutic target. Neuropilins (NRPs) are widely expressed multifunctional single-pass transmembrane proteins which lack their own kinase activity. Instead, NRPs form co-receptors and have been shown to enhance cell responses to several growth factors. Both Neuropilin-1 (NRP1) and NRP2 play essential roles in angiogenesis and axonal guidance, and both are expressed by macrophages. However, little is known about the role of NRP2 in macrophage function or atherosclerosis.PurposeTo investigate whether NRP2 plays a role in macrophage signalling and recruitment in atherosclerosis, we characterised the consequences of conditional NRP2 deletion in macrophage function and atherosclerotic plaque development.MethodsWe generated mice with a macrophage-specific (LysM-Cre) deletion of Nrp2, and lineage tracing, on the pro-atherogenic Apolipoprotein E-deficient background (Nrp2-KOMac, Apoe-/-, EYFP). These mice were fed a high-fat diet (HFD) for 16 weeks, after which, plaque coverage was assessed by staining their aortas with Oil-red-O. Plaque content was further characterised by sectioning and staining the aortic roots. The role of NRP2 in modulating pro-inflammatory macrophage polarisation and signalling, was investigated using functional assays and transcriptome analysis of Nrp2-KO bone marrow-derived macrophages (M0 BMDMs).ResultsHFD-induced atherosclerotic plaque coverage was significantly reduced in Nrp2-KOMac, Apoe-/-, EYFP mice, with 17.4% total aortic plaque coverage in Nrp2-KOMac, Apoe-/-, EYFP mice, compared to 24.9% in WTApoe-/-, EYFP mice (p=0.0021, n=10). Plaques from Nrp2-KOMac, Apoe-/-, EYFP mice displayed features consistent with increased plaque stability, including reduced necrotic core area, plaque lipid content, and increased cap thickness. NRP2 was significantly upregulated upon M-CSF-mediated differentiation of bone marrow progenitors into BMDMs, and further upregulated by pro-inflammatory polarisation using LPS and IFNg. Furthermore, upregulation of Il1b, Tnfa and Il6, induced by LPS and IFNg, was significantly reduced in Nrp2-KO BMDMs, and these cells displayed a decreased migration towards MCP-1. Transcriptome analysis revealed that NFkB signalling pathway genes, and genes regulating monocyte chemotaxis, were downregulated in Nrp2-KO BMDMs.ConclusionThis investigation leads us to conclude that macrophage-derived NRP2 is pro-atherogenic, likely resulting from its role in positively regulating pro-inflammatory signalling and macrophage migration. Targeting NRP2 expressed on the surface of macrophages could therefore offer a novel therapeutic approach for reducing the disease burden associated with atherosclerosis.Conflict of InterestN/A