Aorta Atherosclerosis Lesion Analysis in Hyperlipidemic Mice

Abstract

Atherosclerosis is a chronic inflammatory disease of large and medium-sized arteries. Apolipoprotein E-deficient (ApoE^-/-) mice are used as experimental models to study human atherosclerosis. ApoE^-/- mice are constitutively hyperlipidemic and develop intima plaques that resemble human plaques. Various issues including experimental design for lesion analysis, dietary conditions, isolation of the aorta, staining methods, morphometry, group size, age, the location within the arterial tree, and statistical analyses are important parameters that need to be addressed to obtain robust data. Here, we provide detailed methods to quantify aorta atherosclerosis.

Figure 1. Preparation of aorta and schematic view of aorta with its major branches.

Image of in-situ aorta preparation before isolation showing preparation of aorta segments corresponding to the right schematic view, perivascular adipose tissue in thorax and abdomen, and paraaortic lymph nodes (left). Schematic view of freshly isolated aorta depicting the upper thoracic part with its branches (cyan) and lower abdominal part with its branches (yellow). Diaphragm indicates border between thorax and abdomen (right).

Figure 2. Atherosclerosis lesion analysis of whole aorta.

Removal of perivascular adipose tissue from freshly isolated aorta and trimming of branching arteries under the dissection microscope (left, middle left). Longitudinally split and pinned whole aorta before en-face staining (middle right). Enumeration of total aorta surface area and plaque surface area for morphometric analysis of Sudan-IV-stained whole aorta (right).

Figure 3. Atherosclerosis lesion analysis of aorta cross sections.

Freshly isolated aorta with thoracic and abdominal perivascular adipose tissue (left). Note that perivascular adipose tissue in aortic arch was removed for better visibility of the branching arteries. Schematic presentation of isolation and embedding of different aorta segments including thorax-I with its branches (cyan) and thorax-II (green) above diaphragm, abdomen-I with its branches (yellow) and abdomen-II with its branches (red) below diaphragm (middle). Rotate thorax-I at 180° angle before embedding. Arrow indicates direction of cutting. Enumeration of lumen area, internal elastic lamina (IEL) and external elastic lamina (EEL) area for morphometric analysis of Oil Red O and hematoxylin-stained innominate artery (right).