Disparate Changes in the Mechanical Properties of Murine Carotid Arteries and Aorta in Response to Chronic Infusion of Angiotensin-II

Abstract

Chronic infusion of angiotensin-II has proved useful for generating dissecting aortic aneurysms in atheroprone mice. These lesions preferentially form in the suprarenal abdominal aorta and sometimes in the ascending aorta, but reasons for such localization remain unknown. This study focused on why these lesions do not form in other large (central) arteries. Toward this end, we quantified and compared the geometry, composition, and biaxial material behavior (using a nonlinear constitutive relation) of common carotid arteries from three groups of mice: non-treated controls as well as mice receiving a subcutaneous infusion of angiotensin-II for 28 days that either did or did not lead to the development of a dissecting aortic aneurysm. Consistent with the mild hypertension induced by the angiotensin-II, the carotid wall thickened as expected and remodeled modestly. There was no evidence, however, of a marked loss of elastic fibers or smooth muscle cells, each of which appear to be initiating events for the development of aneurysms, and there was no evidence of intramural discontinuities that might give rise to dissections.

Keywords: constitutive properties; mechanics; stiffness; stress-strain.

Figure 1

Method for quantifying constituent area fractions illustrated for a representative untreated control vessel. (a) VVG images were used to identify elastin (H = 0° −360°, S = 0.0 -1.0, L = 0.0 - 0.22). (b) MTC images were used to quantify both smooth muscle cell (SMC) cytoplasm (H = 275° −10°, S = 0.13 - 1.0, L = 0.14 − 1.0) and collagen (H = 150° −260°, S = 0.12 - 1.0, L = 0.14 − 1.0). In each case, we show both the original image and the extracted constituent area fractions. All images are at the same scale and can be overlaid to verify the results of the pixel isolation and subsequent quantification.

Figure 2

Passive axial force-stretch behavior during cyclic extension tests at three fixed pressures. The vertical line represents the cross-over point, which indicates the in-vivo value of axial stretch λ_z^iv (Humphrey et al., 2009). (a) Representative data from the untreated control group, vessel #4. (b) Representative data from the Ang-II infusion without AAA group, vessel #1. (c) Representative data from the Ang-II infusion with AAA group, vessel #1. Despite the trend toward axial stiffening and decrease in the in-vivo axial stretch, there was no significant difference between the two Ang-II groups and the untreated control group (cf. Table 1).

Figure 3

Representative plots of the theoretical fits (solid lines) to experimental data (○) from passive pressure-diameter and pressure-force responses during cyclic pressurization tests at three fixed axial stretches (i.e., at λ_z^iv and ± 5% this value). (a),(d) Representative results from the untreated control group, vessel #4. (b),(e) Representative results from the Ang-II infusion without AAA group, vessel #1. (c),(f) Representative results from the Ang-II infusion with AAA group, vessel #1. The associated best-fit parameters are in Table 2.

Figure 4

Plots of the theoretically predicted (solid lines) and experimentally inferred (○) passive circumferential stress-stretch behavior during cyclic pressurization tests at three fixed axial stretches (at λ_z^iv and ± 5% this value) for all three groups. (a) Representative data from the untreated control group, vessel #4. (b) Representative data from the Ang-II infusion without AAA group, vessel #1. (c) Representative data from the Ang-II infusion with AAA group, vessel #1.

Figure 5

Mean values of the strain energy function W (equation 7) at the respective in vivo axial stretch and over a physiological pressure range (60 − 120 mmHg), shown here in kPa. Associated specimen-specific circumferential and axial stretches were used at each pressure in conjunction with the best-fit parameters for each vessel to compute the strain energy. Error bars representing the standard deviation at each pressure value were omitted for clarity. All comparisons across groups and pressures were not significantly different at a confidence level of p < 0.05.

Figure 6

Histological images of representative vessels from the untreated control group (row 1), the Ang-II infusion without AAA group (row 2), and the Ang-II infusion with AAA group (row 3). The leftmost images show a Verhoeff van Gieson (VVG) stain with elastic fibers in black and collagen in red-pink; the rightmost images show a Masson's Trichrome (MTC) stain with collagen in blue and smooth muscle cells (SMC) in red. Note the increase in medial thickness in response to Ang-II infusion (rows 2 and 3), as indicated both by the increased distance between elastic lamellae in the VVG stain and the SMC hypertrophy/hyperplasia in the MTC stain. Note, too, the increased collagen deposition in the adventitia as shown by the darker blue staining in the Ang-II infused mice as compared to the control group. Thus, Ang-II infusion appeared to cause a modest adaptive growth and remodeling consistent with mild hypertension. Finally, note that the outermost images were acquired using a 20X objective and the innermost images using a 60X objective. The black scale bar represents 100 μm in each image.

Figure 7

Histological images of a section of (a) an untreated control vessel and (b) an Ang-II treated vessel with AAA, both imaged using a 60X objective and zoomed-in further for observation. Visual comparisons suggested that the lamellae in the Ang-II treated vessels were thicker than those in the control vessels, with increased inter-lamellar distance and overall greater medial thickness. The yellow arrows in (b) indicate locations of discontinuity in staining for elastin, potentially revealing some initial fraying or less dense packing of elastin in the lamellae. The black scale bar represents 100 μm.