Mechanical, structural, and dynamical modifications of cholesterol exposed porcine aortic elastin

Abstract

Elastin is a protein of the extracellular matrix that contributes significantly to the elasticity of connective tissues. In this study, we examine dynamical and structural modifications of aortic elastin exposed to cholesterol by NMR spectroscopic and relaxation methodologies. Macroscopic measurements are also presented and reveal that cholesterol treatment may cause a decrease in the stiffness of tissue. ²H NMR relaxation techniques revealed differences between the relative populations of water that correlate with the swelling of the tissue following cholesterol exposure. ¹³C magic-angle-spinning NMR spectroscopy and relaxation methods indicate that cholesterol treated aortic elastin is more mobile than control samples. Molecular dynamics simulations on a short elastin repeat VPGVG in the presence of cholesterol are used to investigate the energetic and entropic contributions to the retractive force, in comparison to the same peptide in water. Peptide stiffness is observed to reduce following cholesterol exposure due to a decrease in the entropic force.

Keywords: (13)C MAS NMR; (2)H NMR; Cholesterol; Elastin.

Figure 1

Longitude, circumference, and thickness of elastin treated with cholesterol for 7 days (n=6). The sizes of elastin samples after treatment were normalized to the sizes before treatment, (p = 0.7228, 0.3060, and 0.1092 for the longitudinal, circumferential, and thickness directions, respectively).

Figure 2

Average Cauchy stress versus stretch curves of untreated (UT) and cholesterol treated (CT) elastin along the A) longitudinal (L, p=0.7159) and B) circumferential (C, p=0.4735) (n=6).

Figure 3

Normalized tangent modulus of cholesterol treated (CT) elastin in the longitudinal (L, p=0.3038) and circumferential(C, p=0.1404) directions (n=6).

Figure 4

Direct polarization (DP) ¹³C NMR spectra of untreated (A1-B1) and cholesterol treated (A2-B2) porcine aortic elastin samples acquired with decoupling at 10 kHz magic angle spinning. Spectra labeled (B) correspond to the aliphatic region and spectra labeled (A) the carbonyl region. Differences are observed in the resolution of the various peaks between the samples, as discussed in the text.

Figure 5

¹H - ¹³C cross polarization (CP) ¹³C NMR spectra of untreated (A1-B1) and cholesterol treated (A2-B2) porcine aortic elastin samples acquired with decoupling at 10 kHz magic angle spinning.

Figure 6

²H T₁-T₂ results from untreated (A) and cholesterol-treated (B) porcine aortic elastin at 37°C. Four distinguishable peaks are observed in each sample, denoted by α₁, α₂, β, and γ. Differences in the relative populations of water in the assigned γ and β regions are observed between the samples, as discussed in the text. The numerical values of the T₁ and T₂ times and the relative intensities are provided in Table 4.