Multiphoton microscopy observations of 3D elastin and collagen fiber microstructure changes during pressurization in aortic media

Abstract

Elastin and collagen fibers play important roles in the mechanical properties of aortic media. Because knowledge of local fiber structures is required for detailed analysis of blood vessel wall mechanics, we investigated 3D microstructures of elastin and collagen fibers in thoracic aortas and monitored changes during pressurization. Using multiphoton microscopy, autofluorescence images from elastin and second harmonic generation signals from collagen were acquired in media from rabbit thoracic aortas that were stretched biaxially to restore physiological dimensions. Both elastin and collagen fibers were observed in all longitudinal-circumferential plane images, whereas alternate bright and dark layers were observed along the radial direction and were recognized as elastic laminas (ELs) and smooth muscle-rich layers (SMLs), respectively. Elastin and collagen fibers are mainly oriented in the circumferential direction, and waviness of collagen fibers was significantly higher than that of elastin fibers. Collagen fibers were more undulated in longitudinal than in radial direction, whereas undulation of elastin fibers was equibiaxial. Changes in waviness of collagen fibers during pressurization were then evaluated using 2-dimensional fast Fourier transform in mouse aortas, and indices of waviness of collagen fibers decreased with increases in intraluminal pressure. These indices also showed that collagen fibers in SMLs became straight at lower intraluminal pressures than those in EL, indicating that SMLs stretched more than ELs. These results indicate that deformation of the aorta due to pressurization is complicated because of the heterogeneity of tissue layers and differences in elastic properties of ELs, SMLs, and surrounding collagen and elastin.

Kinematics of collagen fibers in carotid arteries under tension-inflation loading

“…Using a 870 nm excitation wavelength [23], the collagen second harmonic generation (SHG) signal was acquired through 400-492 nm band-pass filters. The multiphoton imaging modality was well documented in previous studies [51,39,49], should additional information be needed. The imaging resolution of the microscope was set to 0.5 µm in all directions [23,44] Fig.…”

“…Upon mechanical loading, the collagen bundles progressively unfold [8,21]. The measurement of fiber waviness [11,44,53,49] revealed in particular that recruitment, or engagement of collagen fibers, is gradual (unsynchronized among fibers) and starts at a finite strain [23,13]. The process can differ whether the observed vascular region is close or remote from the heart (distal regions vs. proximal regions) [55].…”

“…In particular, the non-linear mechanical behavior of the arterial tissue is often associated with the progressive rearrangements of these different fiber networks, at the 10 to 100 µm scale. In the present contribution, we chose to focus on the adventitial collagen, since it was recently shown to be the fiber network the most prone to mechanically-induced rearrangements [44,12,49,31]. In particular, this network, composed of thick bundles of type I collagen fibers [15], is the major structural component of the adventitia, due to its important load-bearing function preventing over-distensions [26].…”

Micromechanics of elastic lamellae: unravelling the role of structural inhomogeneity in multi-scale arterial mechanics

“…Multiphoton microscopy [27,[35][36][37] has been employed to study the ECM organization in blood vessels. Multiphoton microscopy has the ability to image the ECM architecture of biological tissues with minimal sample preparation [26], and has been employed to examine the structure of collagen and elastic fibres in various types of biological tissues such as skin, tendon, ligament and blood vessels [38 -41].…”

Multiscale Mechanical Behavior of Large Arteries

“…At a larger scale, the elastic lamellae progressively unfold with the load application and then stretch, as observed by means of polarized light microscopy by (Sokolis et al 2006). The cohesive pericellular interlaced bundles also straighten and reorient by application of a load (Sokolis et al 2006), and this recruitment process was shown to be faster than the recruitment of the circumferentially oriented parallel bundles covering the elastic lamellae (Sugita & Matsumoto 2016); the latter authors propose the stiffness difference between elastin and smooth muscle cells as the possible explanation, the more compliant surrounding medium allowing faster recruitment of the initially crimped fibers.…”