Ultrasound Biomicroscopic Imaging for Interleukin-1 Receptor Antagonist-Inhibiting Atherosclerosis and Markers of Inflammation in Atherosclerotic Development in Apolipoprotein-E Knockout Mice

Abstract

We sought to validate the hypothesis that the development of atherosclerosis can be suppressed by the interleukin-1 receptor antagonist (IL-1Ra) in murine models of atherosclerosis in vivo, noninvasively seen by means of high-resolution ultrasound biomicroscopy, and we studied changes in inflammatory markers such as IL-1 and C-reactive protein (CRP) plasma levels in these models of atherosclerosis. We divided IL-1Ra(+/-)/apolipoprotein-E (apoE)(-/-) and IL-1Ra(+/+)/apoE(-/-) mice into 2 age groups, used as atherosclerotic models. The control groups were age-matched IL-1Ra(+/+)/apoE(+/+) mice. Plaque thickness was measured in the ascending aorta in short-axis images by means of ultrasound and histology. Plasma levels of IL-1 and CRP were quantified in the 3 murine groups. At 16 weeks, plaque thickness in the ascending aortas of the IL-1Ra(+/-)/apoE(-/-) mice was significantly greater than that in the IL-1Ra(+/+)/apoE(-/-) mice, on ultrasound and histology (P <0.01). In contrast, at 32 weeks, the differences between these 2 genotypes were not statistically significant. Serum IL-1 levels were lower in the IL-1Ra(+/-)/apoE(-/-) mice than in the IL-1Ra(+/+)/apoE(-/-) mice at 16 and 32 weeks (P <0.05). At 16 weeks, serum CRP levels in the IL-1Ra(+/-)/apoE(-/-) mice were higher than in the IL-1Ra(+/+)/apoE(-/-) mice (P <0.01). Our results suggest that ultrasound biomicroscopy enables evaluation of atherosclerotic lesions in vivo, noninvasively and in real-time, in apoE(-/-) mice. Partial IL-1Ra deficiencies might promote early plaque development in 16-week-old apoE(-/-) mice. The balance of IL-1 and IL-1Ra might influence atherosclerotic development. Finally, CRP might affect the initiation of atherosclerosis, rather than its progression.

Keywords: Apolipoproteins E/deficiency/genetics; atherosclerosis/diagnosis/genetics; disease models/animal; inflammation/physiopathology; interleukin-1 receptor antagonist protein; interleukin-1/genetics/physiology; mice, knockout; microscopy/methods; time factors.

Fig. 1.

Images at 16 weeks show plaque development in the vascular lumen. In the IL-1Ra^+/−/apoE^−/− mice, the ascending aorta is shown on A) ultrasound biomicroscopy (UBM) (short-axis view) with B) the corresponding histologic image; C) histologic image of the aortic sinus (short-axis view). In the IL-1Ra^+/+/apoE^−/− mice, the ascending aorta is shown in D) UBM short-axis view with E) the corresponding histologic image; F) histologic image of the aortic sinus. The UBM images show the border of the plaque (circled) within the curvature of the ascending aorta. Histologic images are H & E stains, orig. ×4. apoE = apolipoprotein E; IL-1Ra = interleukin-1 receptor antagonist

Fig. 2.

Images at 32 weeks show plaque in the vascular lumen. In the IL-1Ra^+/−/apoE^−/− mice, the ascending aorta is shown on A) ultrasound biomicroscopy (UBM) (short-axis view) with B) the corresponding histologic image; C) histologic image of the aortic sinus (short-axis view). In the IL-1Ra^+/+/apoE^−/− mice, the ascending aorta is shown in D) UBM short-axis view with E) the corresponding histologic image; F) histologic image of the aortic sinus. The UBM images show the border of the plaque (circled). Histologic images are H & E stains, orig. ×4. apoE = apolipoprotein E; IL-1Ra = interleukin-1 receptor antagonist

Fig. 3.

A) At 16 weeks, the aortic sinus plaque area in the IL-1Ra^+/−/apoE^−/− mice was significantly larger than in the IL-1Ra^+/+/apoE^−/− mice (P <0.01). B) There was no significant difference in plaque area between the groups at 32 weeks. apoE = apolipoprotein E; IL-1Ra = interleukin-1 receptor antagonist P <0.05 was considered statistically significant.

Fig. 4.

Plaque thickness measurements on ultrasound biomicroscopy (UBM) significantly correlated to those on histopathology in both A) the IL-1Ra^+/+/apoE^−/− mice and B) the IL-1Ra^+/−/apoE^−/− mice. apoE = apolipoprotein E; IL-1Ra = interleukin-1 receptor antagonist P <0.05 was considered statistically significant.