A Novel Murine Model of Early Abdominal Aortic Aneurysm Reflects Immunopathology in Human Abdominal Aortic Aneurysms

Abstract

The pathogenesis of abdominal aortic aneurysm (AAA) remains poorly understood but there is mounting evidence that AAA may be an immune mediated condition. Current mouse models of AAA were developed primarily to reproduce mature aneurysms and may poorly reflect the pathology found in early AAA. We propose a novel murine model of AAA incorporating sensitization to aortic antigens, which Results in more robust AAA and duplicates the proposed autoimmune mechanisms in humans. C57B/L6 mice were randomized to sensitization to fragmented elastin (fELA) or control. One week later AAA was induced in all mice with the topical elastase method. Prior to sacrifice, development of a delayed-type hypersensitivity (DTH) reaction was assessed by cutaneous challenge in the ear pinna with fELA. Blood and periaortic lymph nodes (LN) were collected for quantification of type 1 regulatory T-cells (Tr1) and T-helper 17 (Th17) cells by flow cytometry. The mice were perfused with a silicone injection compound to fill and opacify their vasculature and microcomputed tomography scans were obtained for three-dimensional reconstruction of the infrarenal aorta (Fig 1) to evaluate volume and surface area of the aneurysms. Mice sensitized to fELA had larger aneurysms: volume 5.31 mm3 versus 4.16 mm3 (P = .034) and surface area 34.55 mm2 versus 30.87 mm2 (P = .038). They had positive DTH reactions to fELA represented by a greater weight difference between challenged and unchallenged ears: 4.13 mg vs 0.21 mg (P = .0004). The degree of DTH response was found to positively correlate with both volume (Fig 2) (R = 0.79; P = .00025) and surface area of aneurysm (R = 0.71; P = .0019). Flow cytometry Results were noTable for a significantly decreased Tr1/Th17 ratio in the sensitized group both in the blood (P = .0005) and LNs (P = .031) and in periaortic LNs this Tr1/Th17 ratio negatively correlated with AAA volume (R = –0.52; P = .021) with a trend toward a similar correlation in the blood (R = –0.49; P = .056). The addition of sensitization to fELA to a surgical topical elastase model of AAA Results in the development of larger aneurysms whose size correlates both with the DTH response to the antigen and with a decreasing Tr1/Th17 ratio in blood and periaortic LNs. The decreased Tr1/Th17 ratio is of particular interest as this was noted to be highly predictive of the presence of AAA in a study of Veterans Affairs patients presenting for screening ultrasound examination (0.5287 in AAA vs 2.6570 in non-AAA; P < .00001; receiver operating characteristic curve with an area under the curve of 0.925; 58 patients).Fig 2The strength of the delayed type-hypersensitivity (DTH) reaction to cutaneous challenge with elastin fragments correlates with aneurysm volume as calculated from three-dimensional reconstructions of the infrarenal aorta on microcomputed tomography scans (MicroCT). Cutaneous challenge was performed on the pinna of the ear and reaction quantified by weight difference compared to contralateral ear which was injected with vehicle.View Large Image Figure ViewerDownload Hi-res image Download (PPT)