Frequency-modulated atomic force microscopy localises viscoelastic remodelling in the ageing sheep aorta

Abstract

Age-related aortic stiffening is associated with cardiovascular diseases such as heart failure. The mechanical functions of the main structural components of the aorta, such as collagen and elastin, are determined in part by their organisation at the micrometer length scale. With age and disease both components undergo aberrant remodelling, hence, there is a need for accurate characterisation of the biomechanical properties at this length scale. In this study we used a frequency-modulated atomic force microscopy (FM-AFM) technique on a model of ageing in female sheep aorta (young: ~18 months, old: >8 years) to measure the micromechanical properties of the medial layer of the ascending aorta. The novelty of our FM-AFM method, operated at 30 kHz, is that it is non-contact and can be performed on a conventional AFM using the ׳cantilever tune’ mode, with a spatial (areal) resolution of around 1.6 μm2. We found significant changes in the elastic and viscoelastic properties within the medial lamellar unit (elastic lamellae and adjacent inter-lamellar space) with age. In particular, there was an increase in elastic modulus (Young; geometric mean (geometric SD)=42.9 (2.26) kPa, Old=113.9 (2.57) kPa, P<0.0001), G′ and G″ (storage and loss modulus respectively) (Young; G′=14.3 (2.26) kPa, Old G′=38.0 (2.57) kPa, P<0.0001; Young; G″=14.5 (2.56) kPa, Old G″=32.8 (2.52) kPa, P<0.0001). The trends observed in the elastic properties with FM-AFM matched those we have previously found using scanning acoustic microscopy (SAM). The utility of the FM-AFM method is that it does not require custom AFM hardware and can be used to simultaneously determine the elastic and viscoelastic behaviour of a biological sample.

Macro- and Micro-mechanical Properties of the Ovine Aorta: Correlation with Regional Variations in Collagen, Elastin and Glycosaminoglycan Levels

“…By presenting both uniaxial and nanoindentation data we are able to compare our data with other studies and also correlate our novel nanoindentation method with more conventional, established mechanical testing methods. We believe that these data will serve as a useful reference given the increase in use of indentation-based techniques for aortic biomechanical analysis [29][30][31][32].…”

Nano-scientific Application of Atomic Force Microscopy in Pathology: from Molecules to Tissues

“…A simple indentation experiment was carried out to determine the elastic moduli of samples. Some attempts were made to obtain both the storage and loss moduli of biological samples by superimposing the oscillating motion on the probe while indenting the samples [113,115,119]. For the clinical applications of AFM techniques, there have been an increasing number of studies investigating tissues obtained from a minimally invasive surgical intervention such as biopsy [47,50,120].…”

“…Mechanical stiffening has also been noted as a hallmark of cardiovascular complications [119,121]. A recent AFM study directly observed a significant increase in the elastic moduli of ventricular tissues freshly harvested from mice with pressure overloadinduced cardiac hypertrophy [121].…”

“…The findings of the study provided information not only on the direct quantification of mechanical changes in the myocardium but also on novel pharmacological interventions to slow down the detrimental progress of cardiac hypertrophy. Moreover, age-related aortic stiffening, another cause of heart failure, was quantitatively evaluated by an AFM-based biomechanical assay [119]. The study was unique because frequency modulated atomic force microscopy (FM-AFM) was used to determine both the storage and loss moduli in localized regions.…”

Nanomechanics and ultrastructure of the internal mammary artery adventitia in patients with low and high pulse wave velocity

“…For example, age-related arterial stiffening is largely attributed to changes in the intima due to atherosclerosis, and degeneration of the media [4] . Furthermore, within these layers, alterations can be localized to individual components at the nano- and micro- scale [5] , [6] . Hence, to understand the mechanisms driving arterial stiffening, the nano-structure and mechanical properties of individual layers within the artery need to be considered.…”