Mechanical heterogeneities in the subendothelial matrix develop with age and decrease with exercise

Abstract

Arterial stiffening occurs with age and is associated with lack of exercise. Notably both age and lack of exercise are major cardiovascular risk factors. While it is well established that bulk arterial stiffness increases with age, more recent data suggest that the intima, the innermost arterial layer, also stiffens during aging. Micro-scale mechanical characterization of individual layers is important because cells primarily sense the matrix that they are in contact with and not necessarily the bulk stiffness of the vessel wall. To investigate the relationship between age, exercise, and subendothelial matrix stiffening, atomic force microscopy was utilized here to indent the subendothelial matrix of the thoracic aorta from young, aged-sedentary, and aged-exercised mice, and elastic modulus values were compared to conventional pulse wave velocity measurements. The subendothelial matrix elastic modulus was elevated in aged-sedentary mice compared to young or aged-exercised mice, and the macro-scale stiffness of the artery was found to linearly correlate with the subendothelial matrix elastic modulus. Notably, we also found that with age, there exists an increase in the point-to-point variations in modulus across the subendothelial matrix, indicating non-uniform stiffening. Importantly, this heterogeneity is reversible with exercise. Given that vessel stiffening is known to cause aberrant endothelial cell behavior, and the spatial heterogeneities we find exist on a length scale much smaller than the size of a cell, these data suggest that further investigation in the heterogeneity of the subendothelial matrix elastic modulus is necessary to fully understand the effects of physiological matrix stiffening on cell function.

Beneficial Effects of Exercise on Subendothelial Matrix Stiffness Are Short-Lived

“…Swimming Protocol. Mice were swum in groups of 2-5 mice per tank as previously described [5]. Mice swam five days a week for 10 min per day in week 1, 30 min per day in weeks 2-3, and 45 min per day for weeks 4-8.…”

“…Increased stiffness within the vasculature is independently associated with cardiovascular disease risk, and is often measured using pulse wave velocity (PWV), a standard clinical measurement of bulk arterial stiffness [1]. Well-known risk factors for cardiovascular disease include increased age and lack of exercise [2,3], which are also associated with increased artery stiffness [1,4,5]. Changes in subendothelial matrix stiffness are associated with endothelial cell contraction, endothelial cell-cell junction disruption, and increased vessel permeability [6,7].…”

“…Changes in subendothelial matrix stiffness are associated with endothelial cell contraction, endothelial cell-cell junction disruption, and increased vessel permeability [6,7]. Previously, we showed that increased vascular stiffness in aged mice can be decreased through exercise on both the macro and microscales [5]. However, the lasting effects of exercise on microscale intimal stiffness after exercise cessation are unknown.…”

Microscale Interrogation of 3D Tissue Mechanics

“…Viscoelasticity can be measured when the cantilever is operated in an oscillating mode (Rother et al, 2014;Connizzo and Grodzinsky, 2017). Surface mapping of tissue mechanics using AFM can be used to identify mechanical heterogeneities in tissues, such as age-related heterogeneous vessel stiffening (Kohn et al, 2016) and malignancy-related heterogeneous tumor cell softening (Plodinec et al, 2012). However, the surface probing nature of AFM makes it difficult to use for 3D mapping of tissue mechanics.…”

Mapping cellular-scale internal stiffness in 3D tissues with smart material hydrogel probes

“…Exquisitely structured tissues and organs arise from a homogenous blastomere through spatial patterns of cell proliferation, migration, and differentiation, in concert with matrix secretion and remodelling [1][2][3][4] . Mechanical features of the local microenvironment are critical regulators of these cellular processes [5][6][7][8][9][10][11][12] , and tissue stiffness is now well-established to drive fate-function relationships during development 13,14 ; disease progression [15][16][17][18] and tissue homeostasis [19][20][21] . However, our technical ability to monitor stiffness at the cellular length scale during tissue development remains severely limited, and could be critically important in elucidating biophysical mechanisms of tissue morphogenesis and disease.…”