Integrating structural heterogeneity, fiber orientation, and recruitment in multiscale ECM mechanics
Introduction
Extracellular matrix (ECM) provides the principal avenue for mechanical and biochemical communications between tissue and cells. Cells actively sense the external mechanical forces applied through the ECM and also their surrounding local stiffness (Throm Quinlan et al., 2011). External mechanical forces and ECM stiffness have been shown to play important roles in regulating fundamental cellular morphology (Wells, 2008) and functions, such as proliferation (Wells, 2008; Schrader et al., 2011), differentiation (Engler et al., 2006; Even-Ram et al., 2006) and mobility (Pelham and Wang, 1997; Zaman et al., 2006). Many pathological conditions involve significant alterations of ECM mechanical properties (Baker et al., 2009; Janmey and Miller, 2011; Lu et al., 2012; Engler et al., 2008; Chaturvedi et al., 2010; Georges et al., 2007; Wells, 2008; Shkumatov et al., 2015; White, 2015). Understanding the translation of mechanical forces from the tissue to cellular level is crucial and a requisite bridge between research in the fields of cell and tissue mechanics. However, the force translation in ECM is inherently a multiscale process ranging from the macroscopic to microscopic level. The mechanical properties of ECM, known to have a 3D hierarchical structure, have been shown to be highly dependent on the scale of measurement (Aifantis et al., 2011; Li et al., 2017). As a result, the distribution of mechanical force in the ECM is highly dependent on the hierarchical structure and the local mechanical properties of the ECM (Pizzo et al., 2005; Mow et al., 1994).
The macro- and microscopic mechanical properties of ECM have been broadly studied. To investigate the macroscopic mechanical properties of collagen matrix, various experimental methods, such as uniaxial tension (Roeder et al., 2002; Feng et al., 2003; Duan and Sheardown, 2006; Van Oosten et al., 2016), biaxial tension (Jhun et al., 2009; Sander et al., 2009; Xu et al., 2011), and rheological tests (Vader et al., 2009; Xu et al., 2011; Kurniawan et al., 2012; Li and Zhang, 2014; Motte and Kaufman, 2012) have been performed. In microscopic mechanical measurements, controlled mechanical forces were applied using a variety of techniques including indentation (Wenger et al., 2007; Aifantis et al., 2011; Mckee et al., 2011), laser tracking microrheology (Velegol and Lanni, 2001; Parekh and Velegol, 2007; Sun et al., 2004; Shayegan and Forde, 2013), magnetic twisting cytometry (Leung, et al., 2007), and optical magnetic twisting cytometry (OMTC) (Li et al., 2017). These studies on the mechanical properties of ECM are focused on a single measurement scale. Advances in imaging techniques promote the integration of multiscale mechanical loading and imaging modalities to provide perspectives on load induced changes in ECM microstructure, cellular morphology and alignment (Bell et al., 2012; Throm Quinlan et al., 2011). An integrated mechanical loading-confocal microscopy system was developed and used to study three-dimensional (3D) mechanical strains in collagen matrix at multiple length scales in response to uniaxial (Roeder et al., 2004, Roeder et al., 2009) and biaxial stretching (Bell et al., 2012). To obtain local mechanical properties, local strains were measured, and the macro-scale mechanical properties of a substrate are usually used in the calculation of local traction forces (Franck et al., 2011; Toyjanova et al., 2014).
To date, there has been limited understanding of multiscale ECM mechanical properties. Several previous studies have attempted to measure the local mechanical properties of ECM and showed that the local mechanical properties of an interconnected network do not always match its global properties. Microscopic mechanical properties of ECM measured by laser trap microscopy (Velegol and Lanni, 2001) and AFM (Gautreau et al., 2006; Throm Quinlan and Billar, 2012)yielded consistently greater stiffness measurements than those obtained from macroscopic rheometry and uniaxial tensile tests. Significant differences in the mechanical properties of ECM were shown ranging from the macro to nano scales as Etendon < Efiber < Efibri (Aifantis et al., 2011). The microscopic mechanical properties were found to be less sensitive to changes in collagen concentration, as compared to the macroscopic mechanical properties (Li et al., 2017; Baniasadi and Minary-Jolandan, 2015). The scale of measurements was found to play an important role in multiscale ECM mechanical properties. The differences in multiple ECM mechanical properties emerge when the probe size is comparable to or smaller than the average pore size of the sample (Costa et al., 2003).
In the present study, we created a multiscale experimental approach that combines optical magnetic twisting cytometry (OMTC) and biaxial tensile testing techniques to study the changes in local ECM mechanical properties with controlled global biaxial mechanical loading. The biaxial tensile test considers the multi-axial loading state under physiological conditions and has shown promises in fully characterizing the anisotropic nature of soft biological tissues (Sacks and Sun, 2003; Nerurkar et al., 2010; Bell et al., 2012). By oscillating microscale ferromagnetic beads bound to collagen fibers, OMTC was used to measure the local mechanical properties of collagen matrix. OMTC allows the measurements of a population of beads and can effectively eliminate unbound or loosely bound beads (Fabry et al., 2001). Porcine adventitia layer was used as an ECM equivalent due to several reasons: 1) Porcine adventitia is the outmost layer of the artery and consists primarily of type I collagen fibers. 2) It is semi-transparent so it is possible to make optical measurements using OMTC. 3) Compared to collagen gel, it is more robust to work with when biaxial loading is applied. Multiphoton microscopy was also used to capture the structural changes of collagen fibers under tissue-level biaxial loading.
Section snippets
Sample preparation
Descending thoracic aortas of porcine (12 – 24 month old) were harvested at a local abattoir and transported to the laboratory on ice. Before experiments, the aortas were cleaned of adherent tissues and fat and rinsed in deionized water. All samples were taken from approximately the same longitudinal region of the aorta to avoid changes in mechanical properties along the longitudinal anatomic position (Zeinali-Davarani et al., 2015). The adventitial layer was carefully peeled from the porcine
Tissue-level mechanical properties and ECM structure
Fig. 2a shows the average Cauchy stress vs. stretch curves from biaxial tensile testing of the adventitia tissue sample, which is highly nonlinear with the longitudinal direction stiffening slightly earlier than the circumferential direction. To better compare the mechanical properties in the longitudinal and circumferential directions, tangent modulus was obtained by differentiating the Cauchy stress-stretch curves in Fig. 2a as dσ/dλ. As shown in Fig. 2b, a prominent increase in tangent
Discussion
In this study, a new experimental methodology was developed to study the changes of microscopic ECM mechanics under macroscopic loading. Specifically, OMTC was used to measure the local mechanical properties of the collagen network when subjected to controlled global biaxial mechanical loading. Our study suggests that it is important to consider the complex interplay among ECM fiber orientation, fiber engagement, and the tissue-level mechanical loading when understanding the scale dependency of
Conclusions
A novel experimental approach that allows the measurements of local ECM mechanical properties with controlled tissue-level biaxial mechanical loading was established and used to study the multiscale ECM mechanics of the collagen network in the adventitial layer of the arterial wall. This study reported several interesting findings of the role of ECM fiber network structure in determining the local ECM mechanical properties upon tissue-level mechanical loading. Our study shows that the ECM fiber
Acknowledgements
The authors would like to acknowledge the funding support from the National Science Foundation (CAREER 0954825) and National Institutes of Health (2R01HL098028).
References (72)
- et al.
Extracellular matrix stiffness and architecture govern intracellular rheology in cancer
Biophys. J.
(2009) - et al.
Multiscale strain analysis of tissue equivalents using a custom-designed biaxial testing device
Biophys. J.
(2012) - et al.
Arterial extracellular matrix: a mechanobiological study of the contributions and interactions of elastin and collagen
Biophys. J.
(2014) From single fiber to macro-level mechanics: a structural finite element model for elastomeric fibrous biomaterials
J. Mech. Behav. Biomed. Mater.
(2014)- et al.
Dendrimer crosslinked collagen as a corneal tissue engineering scaffold: mechanical properties and corneal epithelial cell interactions
Biomaterials
(2006) - et al.
Matrix elasticity directs stem cell lineage specification
Cell
(2006) - et al.
Matrix control of stem cell fate
Cell
(2006) - et al.
Flow and magnetic field induced collagen alignment
Biomaterials
(2007) - et al.
Mechanical design criteria for intervertebral disc tissue engineering
J. Biomech.
(2010) - et al.
Stretching type II collagen with optical tweezers
J. Biomech.
(2004)
Cell traction forces on soft biomaterials. I. Microrheology of type I collagen gels
Biophys. J.
Mechanical properties of collagen fibrils
Biophys. J.
Transverse mechanical properties of collagen fibers from nanoidentation
J. Mater. Sci.: Mater. Med.
Alginate-collagen fibril composite hydrogel
Materials
Affine versus non-affine fibril kinematics in collagen networks: theoretical studies of network behavior
J. Biomech. Eng.
Passive stiffness of myocardium from congenital heart disease and implications for diastole
Circulation
Embryonic cardiomyocytes beat best on a matrix with heart-like elasticity: scar-like rigidity inhibits beating
J. Cell Sci.
Time scale and other invariants of integrative mechanical behavior in living cells
Phys. Rev. E
Signal Transduction in Smooth Muscle Selected Contribution: time course and heterogeneity of contractile responses in cultured human airway smooth muscle cells
J. Appl. Physiol.
Investigation on the mechanical properties of contracted collagen gels as a scaffold for tissue engineering
Artif. Organs
Three-dimensional traction force microscopy: a new tool for quantifying cell-matrix interactions
PLoS One
Fibrillar structure and mechanical properties of collagen
J. Struct. Biol.
Hyperelastic modelling of arterial layers with distributed collagen fibre orientations
J. R. Soc. Interface
Characterizing Viscoelastic Properties of Polyacrylamide Gels
Increased stiffness of the rat liver precedes matrix deposition: implications for fibrosis
Am. J. Physiol. Gastrointest. Liver Physiol.
Pseudostatic and dynamic nanomechanics of the tunica adventitia in elastic arteries using atomic force microscopy
ACS Nano
Mechanisms of mechanical signaling in development and disease
J. Cell Sci.
Planar biaxial mechanical behavior of bioartificial tissues possessing prescribed fiber alignment
J. Biomech. Eng.
Early stiffening and softening of collagen: interplay of deformation mechanisms in biopolymer networks
Biomacromolecules
A new microrheometric approach reveals individual and cooperative roles for TGF-β1 and IL-1β in fibroblast-mediated stiffening of collagen gels
FASEB J.
Multiscale measurements of the mechanical properties of collagen matrix
ACS Biomater. Sci. Eng.
Modeling of the viscoelastic behavior of collagen gel from dynamic oscillatory shear measurements
Biorheology
The extracellular matrix: a dynamic niche in cancer progression
J. Cell Biol.
Glycosaminoglycans contribute to extracellular matrix fiber recruitment and arterial wall mechanics
Biomech. Model Mechanobiol.
Cited by (13)
The effects of axial twisting and material non-symmetry on arterial bent buckling
2023, Journal of BiomechanicsReconstructing auto tissue engineering lamellar cornea with aspartic acid modified acellular porcine corneal stroma and preconditioned limbal stem cell for corneal regeneration
2022, BiomaterialsCitation Excerpt :With the purpose of reconstructing the ATELC, favorite surface properties of the scaffold are also required. The scaffold surface is expected to possess favorable hydrophilic and adhesive, which could allow seeded cells to grow and proliferate [60,61]. Owing to the destruction of delicate collagen fibers ultrastructure following decellularization, the hydrophilic property of APCS scaffold surface was decreased compared to the NPC.
Effects of material non-symmetry on the mechanical behavior of arterial wall
2022, Journal of the Mechanical Behavior of Biomedical MaterialsCitation Excerpt :There has been strong evidence of nonsymmetric two fiber families in arteries. First, several studies reported nonsymmetric fiber dispersion in porcine and human arteries (Schriefl et al. 2012a, 2012b; Mottahedi and Han, 2016; Li et al., 2019) although the differences have been ignored in modeling. A 19% difference in dispersion parameters between the two fiber families (k1 = 0.1173, k2 = 0.1396) was reported in human aorta (Schriefl et al., 2012a) though its effect was not examined.
An ultrastructural 3D reconstruction method for observing the arrangement of collagen fibrils and proteoglycans in the human aortic wall under mechanical load
2022, Acta BiomaterialiaCitation Excerpt :Some studies focused on the mechanical characterization of an isolated collagen monomer [18] or fibril [19], but it should be taken into account that collagen fibrils do not act independently in the aortic wall [20]. Recent studies have shown that bundles of collagen fibrils, i.e. collagen fibers, change their microstructural organization in loaded arterial tissues [21–25]. Nevertheless, the mechanical properties of the tissue may not only be limited to the micro-architecture, but also be influenced by the quality [26] and integrity [27] of the collagen fibers.