Abstract 114: Macrophages Mediate the Ability of the Receptor for Advanced Glycation End Products to Prevent Formation of Abdominal Aortic Aneurysm in a Murine Model

Abstract

INTRODUCTION: The pathophysiology of AAAs is multifactorial, involving chronic inflammation, increased MMP expression/activity, and medial elastic fiber destruction. RAGE transduces the biological impact of ligand families including AGEs, S100, and HMGB1 to upregulate inflammatory and tissue injury-provoking genes including MMPs. We previously identified increased RAGE expression in human AAAs and have shown that the absence of RAGE, whether by genetic deletion or pharmacologic blockade with sRAGE, prevents AAA formation in a mouse model. We sought to characterize the cellular mediators of this effect. METHODS: C57BL/6 wild-type (WT) and RAGE null (KO) mice underwent transient elastase perfusion to induce AAAs. Mice were sacrificed at day 14 and flow cytometry performed to characterize differences in activation (CD69+) of lymphocytes (CD4/8), neutrophils (GR1+/CD11b+), macrophages (F480), NK cells (NK1.1) (n=24/group). Immunohistochemistry was performed for Mac-3 and Arg 1 (M2 marker) or Arg 2 (M1 marker). Primary BMDMs were isolated and skewed towards M1 (LPS/IFN ¡) or M2 (LPS, IL-4, IL-13) with the addition of S100b or HMGB1 to determine the ligands' effect upon macrophage skewing by ELISA. RESULTS: Flow cytometry revealed increased aortic expression of activated CD8+ cells (p=0.002), macrophages (p-0.002), and NK cells (p=0.03). No differences were noted in cellular activation from spleens from the same animals. Immunohistochemistry revealed increased macrophage expression in WT vs KO aortae, highest at day 5 (p=0.04). Arg 1 and 2 expression on macrophages was elevated in WT vs KO animals (p=0.03). Skewing BMDMs from WT and KO towards M1 led to elevated TNFα with addition of either S100b (p=0.02) or HMGB1 (p=0.03). Skewing BMDMs towards M2 led to elevated IL-10 in KO vs WT cells with the addition of S100b (p=0.01). HMGB1 has no effect upon skewing towards M2. CONCLUSIONS: The effect of RAGE in the development of murine AAA appears to be mediated by macrophages. Differential macrophage skewing towards an M1 by both S100b and HMGB1 likely plays a role in this effect. Skewing towards M2 is mediated by S100b and not HMGB1. Further studies will seek to identify at which stage in AAA formation activation of the RAGE-ligand axis becomes functionally important.