AA9. Macrophage-Derived Netrin 1 Promotes Abdominal Aortic Aneurysms by Activating Matrix Metalloproteinase 3 in Vascular Smooth Muscle Cells

Abstract

Aberrant extracellular matrix (ECM) fragmentation and sustained inflammation are hallmarks of abdominal aortic aneurysms (AAAs). However, the mechanisms by which these events are coupled, thereby fueling focal vascular damage, are undefined. We have previously demonstrated critical immunoregulatory functions of the neuronal guidance cue netrin 1 in regulating macrophage accumulation in the vasculature. Here, we investigate the novel role of netrin 1 in the pathogenesis of AAA and test the hypothesis that netrin 1 mediates crosstalk between infiltrating macrophages and vascular smooth muscle cells (VSMCs), thereby promoting ECM degradation in AAA. AAA was experimentally induced in apolipoprotein E-deficient (ApoE−/−) C57Bl/6 mice infused with angiotensin II for 28 days. Aneurysm progression and severity were monitored by ultrasound Doppler imaging and elastin staining. A t-distributed stochastic neighbor embedding analysis of data sets from single-cell RNA sequencing of diseased aortas profiled the distribution of netrin 1 among the aortic cellular subpopulations. Conditional deletion of netrin 1 in macrophages was obtained with Ntn1flox/floxLysMCre+/− transgenic mouse models, and AAA was induced. RNA sequencing identified the specific regulation of matrix metalloprotease 3 (MMP3) by netrin 1 in VSMCs. MMP3-deficient mice were exposed to angiotensin II, and the contribution to ECM degradation in response to netrin 1 signaling was analyzed. Netrin 1-induced calcium influx and nuclear translocation of nuclear factor of activated T cells, cytoplasmic 3 (NFATc3) were measured in VSMCs. Netrin 1 expression peaked in human and murine aneurysmal macrophages. Targeted deletion of netrin 1 macrophage protected mice from disease occurrence and complications compared with control mice. Macrophage-derived netrin 1 induced robust intracellular calcium flux necessary for the nuclear localization of the transcription factor NFATc3, leading to transcriptional regulation and sustained catalytic activity of MMP3 by adjacent VSMCs. The deficiency of MMP3 reduced ECM damage and the susceptibility of mice to develop AAA. Here, we established netrin 1 as a major signal that mediates the crosstalk between inflammation and chronic erosion of the ECM in AAA. Strategies to target netrin 1 could be of potential therapeutic interest to reduce the progression and morbidity of AAA.