Vascular extracellular matrix and arterial mechanics

Abstract

An important factor in the transition from an open to a closed circulatory system was a change in vessel wall structure and composition that enabled the large arteries to store and release energy during the cardiac cycle. The component of the arterial wall in vertebrates that accounts for these properties is the elastic fiber network organized by medial smooth muscle. Beginning with the onset of pulsatile blood flow in the developing aorta, smooth muscle cells in the vessel wall produce a complex extracellular matrix (ECM) that will ultimately define the mechanical properties that are critical for proper function of the adult vascular system. This review discusses the structural ECM proteins in the vertebrate aortic wall and will explore how the choice of ECM components has changed through evolution as the cardiovascular system became more advanced and pulse pressure increased. By correlating vessel mechanics with physiological blood pressure across animal species and in mice with altered vessel compliance, we show that cardiac and vascular development are physiologically coupled, and we provide evidence for a universal elastic modulus that controls the parameters of ECM deposition in vessel wall development. We also discuss mechanical models that can be used to design better tissue-engineered vessels and to test the efficacy of clinical treatments.

FIG. 1

Nonlinear mechanical behavior of the adult mouse aorta. A: average circumferential stress versus stretch ratio. B: circumferential incremental elastic modulus (E_inc) versus stretch ratio. E_inc was calculated by determining the local slope of the stress-stretch ratio relationship in A. The physiological region is highlighted in gray for each graph. Note the decreased incremental elastic modulus at low stretch ratios where elastin dominates the vessel mechanical behavior and the increased modulus at high stretch ratios where collagen dominates. The physiological range is at the intersection of these two regions. The sharp increase in modulus just beyond the physiological range prevents distension of, and damage to, the vessel with increased pressure. [Data replotted from Wagenseil et al. (290).]

FIG. 2

Immunofluorescence micrograph of E17 mouse aorta. Sections through the aorta of an E17 mouse were stained with an antibody for elastin (green) and for flk, a marker for endothelial cells (red). On the left is the lumen (L) of the artery. The intima (I) is evident as a single layer of red-staining endothelial cells. The media (M) contains dense layers of elastin, whereas the elastin in the adventitia (Ad) consists of fine fibers. The vein (V) on the top right shows the presence of endothelial cells but no elastin, whereas the small artery (Ar) directly below shows both. Scale bar = 100 μm. (Micrograph provided by Dr. Sean McLean.)

FIG. 3

Elastic lamellae in human aorta. Top: electron micrograph of human aorta in cross-section showing the arrangement of smooth muscle cell layers separated by the darkly stained elastic lamellae. The lumen of the vessel is at the top. The image on the bottom shows the network of elastin after all cells and other extracellular matrix (ECM) proteins are removed by autoclaving. The circumferential sheets of elastin are joined across the wall by numerous interlamellar elastin connections, which are important for transferring stress across the wall and throughout the elastic fiber network. Scale bar = 20 μm.

FIG. 4

Hemodynamic parameters and ECM expression increase sharply in late embryonic and early postnatal development in mice. A: systolic pressure and cardiac output versus age are replotted from published studies (131, 132, 147, 297). Dotted lines were interpolated between different studies, as data on the hemodynamic parameters for the last third of embryonic development in mice has not been published. Age was calculated assuming that embryonic development lasts 21 days and the mice are born on day 0. B: median normalized values for elastin and collagen type I expression are replotted from Kelleher et al. (146). Expression of both proteins steadily increases from E14 through P14-21, then rapidly decreases to low levels by ~P30. C: sum of the median normalized elastin and collagen expression versus age. The sum was calculated by totaling all median normalized gene expression from the start of development to the current age. This graph also illustrates that little new elastin or collagen protein accumulates after expression of the two genes are downregulated. Note the logarithmic scale on the vertical axes of graphs in A and C. The similar developmental timeline of hemodynamic parameters and total ECM expression suggests correlations between these events.

FIG. 5

Electron micrographs of developing elastic fibers. A: electron micrograph showing a developing elastic fiber adjacent to an elastin-producing cell. Bar = 1.0 μm. B: at higher magnification, the elastic fibers are seen to consist of black amorphous elastin deposited within a bundle of microfibrils. Bar = 0.25 μm. C: in cross section, the microfibrils have a tubular appearance. Bar = 0.25 μm. D: an elastic fiber visualized using quick-freeze, deep etch microscopy (104). Unlike standard transmission microscopy, quick-freeze, deep-etch images provide insight into organization of elastin (E) within the fiber (184, 187). The major feature is a densely packed matrix of 5-7 nm tropoelastin molecules that are associated so tightly that little or no etching occurs during sample preparation. Microfibrils (MF) are seen along the periphery of the fiber and at the end. [A-C from Mecham and Davis (184), copyright Elsevier 1994.]

FIG. 6

Elastin domain structure and cross-link formation. Top: schematic diagram of exon and domain structure of human tropoelastin. Shaded squares represent lysine cross-linking domains that contain prolines (KP) or are enriched in alanines (KA). White squares are hydrophobic sequences. Bottom: cross-linking of elastin monomers is initiated by the oxidative deamination of lysine side chains by the enzyme lysyl oxidase in a reaction that consumes molecular oxygen and releases ammonia. The aldehyde (allysine) that is formed can condense with another modified side chain aldehyde (1) to form the bivalent aldol condensation product (ACP) cross-link. Reaction with the amine of an unmodified side chain through a Schiff base reaction (2) produces dehydrolysinonorleucine (dLNL). ACP and dLNL can then condense to form the tetrafunctional cross-link desmosine or its isomer isodesmosine.

FIG. 7

Mechanical properties and wall structure of invertebrate aortae. A: pressure-volume inflation-deflation curves for aortas from lamprey (top) and lobster (bottom). Relative volume is given as V/V₀, where V is the instantaneous volume of the vessel and V₀ is the volume at the pressure at which the inflation-deflation cycle was started. B: light micrograph showing a transverse section of the abdominal aorta of the lobster, stained with modified Weigert’s technique, showing the positively stained fibrous material forming the internal lamina closest to the lumen (top) dense fibrous matrix within the middle lamina (ML), which contains striated muscle cells (STM) and fibroblasts. No positively staining fibers are observed in the connective tissue of the external layer (EL). Scale bar = 20 μm. C: electron micrograph showing the fibrils within a dense fiber in the middle lamina. Arrowheads indicate the periodicity of the beaded fibrils. Scale bar = 190 nm. [From Davison et al. (50), with permission from the Company of Biologists Ltd.]

FIG. 8

Changes in lamellar ECM composition during transition from an open to a closed circulatory system. The vessel wall in invertebrates with an open circulatory system is exposed to low intraluminal pressure and little or no pulsatile flow. Under these circumstances, the vessel wall does not require elastic recoil for proper function. Ultrastructural analysis of the aorta from the marine whelk, an invertebrate with an open circulatory system, found abundant fibers with the typical appearance of collagen bundles (50). As organisms became more complex and the circulatory system transitions from an open to a closed circuit with increasing pulsatile pressure, the vessel wall developed an extracellular matrix that provides elastic recoil. In invertebrates with a highly developed open circulatory system and in lower vertebrates that have not completed the transition to a fully closed circulatory system (e.g., lamprey and hagfish), the vessel wall contains dense layers of fibrillin-containing microfibrils, which provide elastic recoil not found in the collagen-only vessel of lower invertebrates (50). With the emergence of a fully closed circulation, the major vessels experienced much higher pulse pressure as the entire cardiac output is ejected from the heart during systole. It is at this point in evolution that elastin appears in the wall where it associates with microfibrils to form large elastic structures that provide the elastic recoil required for normal vessel function in vertebrates. [Electron micrographs of whelk and lobster aorta from Davison et al. (50), with permission from The Company of Biologists Ltd., and of bovine aorta from Mecham and Davis (184), copyright Elsevier 1994.]

FIG. 9

Incremental elastic modulus of the aorta in a variety of animal species. Top: plots of incremental elastic modulus (note logarithmic scale) as a function of inflation pressure. Although substantial differences exist among the various animals, in each case the aorta exhibits nonlinear elasticity and the modulus increases dramatically with pressure. Bottom: the same data as above but plotted with pressure normalized to the mean blood pressure for each species. The physiological elastic modulus at P/P_mean = 1 ranged from 0.3 to 1 MPa, with most clustering around 0.4 MPa. The incremental elastic modulus for human (young and old) aorta and wild-type and Eln+/- mouse aorta was determined from data in Learoyd and Taylor (157) and Wagenseil et al. (290), respectively. The modulus for all other animals is from Shadwick (253), with permission from The Company of Biologists Ltd.

FIG. 10

Increased residual shear strain causes the abdominal aorta and left carotid artery of Eln+/- mice to change shape upon excision. Small carbon particles were placed vertically along the length of each vessel in vivo to measure the difference between in vivo and ex vivo length and to document residual shear. Ex vivo, the particles shift to the right along the vessel length, indicating residual shear. The rightward shift is small in wild-type (WT) vessels and becomes more pronounced in Eln+/- vessels. The small residual shear in WT vessels causes only slight curvature in the ex vivo vessel. The increased residual shear in Eln+/- vessels causes either sharp changes in curvature (abdominal aorta) or complete loops (carotid artery) in the ex vivo vessel. Scale bar = 1 mm. [Modified from Wagenseil et al. (290).]

FIG. 11

Vessel inner diameters are similar at physiological blood pressures of WT and Eln+/- genotypes. Comparison of ascending aorta inner diameters of adult WT and Eln+/- mice at their respective physiological pressures shows that the vessel in Eln+/- animals is smaller at every intravascular pressure. Because the basal blood pressure in the Eln+/- genotype is higher, however, its effective working diameter (open arrows) is comparable to that of the WT animal (solid arrows). Green lines show the working diameter of the WT artery at its physiological systolic (S) and diastolic (D) pressures. Red lines show that the systolic (s) and diastolic (d) dimensions of the Eln+/- artery at its higher pressures are similar to the WT values.

FIG. 12

Relationships between the vessel wall dimensions, stresses, and physiological parameters. The dimensions include inner radius (r_i), length (l), and thickness (t). The stresses include wall shear stress (τ_w), longitudinal stress (σ_z), and average circumferential stress (σ_θ). The physiological parameters include volumetric blood flow (Q), longitudinal tethering forces [F(l)], and blood pressure (P). The viscosity of blood (μ) is also included in the wall shear stress equation. The equations show how changes in the vessel wall dimensions and/or physiological parameters can alter the wall stresses.