Elastic properties of hydrogels and decellularized tissue sections used in mechanobiology studies probed by atomic force microscopy

Giménez, Adrià; Uriarte, Juan J.; Vieyra, Joan; Navajas, Daniel; Alcaraz, Jordi

doi:10.1002/jemt.22740

Cited by 29 publications

(23 citation statements)

References 72 publications

(183 reference statements)

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“…It is worth noting that the stiffness measured in the reseeded decellularized matrices (mean 40-60 kPa) was higher than the values measured in native human lung parenchymal tissue (1.34±0.36 kPa; Liu et al, 2010 ) and IPF tissue (16.52±2.25 kPa; Booth et al, 2012 ). It has been reported in the literature that the decellularization method affects the stiffness of acellular tissue ( Melo et al, 2014 ) and, more generally, that decellularized tissue is stiffer than normal lung tissue ( Giménez et al, 2017 ), although the origins of these differences in decellularized versus normal tissue stiffness remain poorly understood. The reseeded decellularized matrices did not form alveolar structures in the time span of this study and were densely packed ( Fig.…”

Section: Discussionmentioning

confidence: 99%

Lysyl oxidases regulate fibrillar collagen remodelling in idiopathic pulmonary fibrosis

Tjin

White

Faiz

et al. 2017

Disease Models &Amp; Mechanisms

123

110

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Idiopathic pulmonary fibrosis (IPF) is a progressive scarring disease of the lung with few effective therapeutic options. Structural remodelling of the extracellular matrix [i.e. collagen cross-linking mediated by the lysyl oxidase (LO) family of enzymes (LOX, LOXL1-4)] might contribute to disease pathogenesis and represent a therapeutic target. This study aimed to further our understanding of the mechanisms by which LO inhibitors might improve lung fibrosis. Lung tissues from IPF and non-IPF subjects were examined for collagen structure (second harmonic generation imaging) and LO gene (microarray analysis) and protein (immunohistochemistry and western blotting) levels. Functional effects (collagen structure and tissue stiffness using atomic force microscopy) of LO inhibitors on collagen remodelling were examined in two models, collagen hydrogels and decellularized human lung matrices. LOXL1/LOXL2 gene expression and protein levels were increased in IPF versus non-IPF. Increased collagen fibril thickness in IPF versus non-IPF lung tissues correlated with increased LOXL1/LOXL2, and decreased LOX, protein expression. β-Aminoproprionitrile (β-APN; pan-LO inhibitor) but not Compound A (LOXL2-specific inhibitor) interfered with transforming growth factor-β-induced collagen remodelling in both models. The β-APN treatment group was tested further, and β-APN was found to interfere with stiffening in the decellularized matrix model. LOXL1 activity might drive collagen remodelling in IPF lungs. The interrelationship between collagen structural remodelling and LOs is disrupted in IPF lungs. Inhibition of LO activity alleviates fibrosis by limiting fibrillar collagen cross-linking, thereby potentially impeding the formation of a pathological microenvironment in IPF.

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Section: Discussionmentioning

confidence: 99%

Lysyl oxidases regulate fibrillar collagen remodelling in idiopathic pulmonary fibrosis

Tjin

White

Faiz

et al. 2017

Disease Models &Amp; Mechanisms

123

110

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“…To culture fibroblasts in conditions that mimic the fibrotic mechanical microenvironment, we adapted two independent and complementary assays based on collagen-rich hydrogels with tunable elasticity as outlined in Figure 1 . Both assays have been widely used in previous mechanobiology studies [ 27 , 28 ]. The floating gel assay is based on culturing fibroblasts embedded in three-dimensional (3D) collagen-I gels that remain either attached to their container or floating, thereby providing high or low mechanical resistance, respectively.…”

Section: Resultsmentioning

confidence: 99%

Dysregulated Collagen Homeostasis by Matrix Stiffening and TGF-β1 in Fibroblasts from Idiopathic Pulmonary Fibrosis Patients: Role of FAK/Akt

Giménez

Duch

Puig

et al. 2017

IJMS

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Idiopathic pulmonary fibrosis (IPF) is an aggressive disease in which normal lung parenchyma is replaced by a stiff dysfunctional scar rich in activated fibroblasts and collagen-I. We examined how the mechanochemical pro-fibrotic microenvironment provided by matrix stiffening and TGF-β1 cooperates in the transcriptional control of collagen homeostasis in normal and fibrotic conditions. For this purpose we cultured fibroblasts from IPF patients or control donors on hydrogels with tunable elasticity, including 3D collagen-I gels and 2D polyacrylamide (PAA) gels. We found that TGF-β1 consistently increased COL1A1 while decreasing MMP1 mRNA levels in hydrogels exhibiting pre-fibrotic or fibrotic-like rigidities concomitantly with an enhanced activation of the FAK/Akt pathway, whereas FAK depletion was sufficient to abrogate these effects. We also demonstrate a synergy between matrix stiffening and TGF-β1 that was positive for COL1A1 and negative for MMP1. Remarkably, the COL1A1 expression upregulation elicited by TGF-β1 alone or synergistically with matrix stiffening were higher in IPF-fibroblasts compared to control fibroblasts in association with larger FAK and Akt activities in the former cells. These findings provide new insights on how matrix stiffening and TGF-β1 cooperate to elicit excessive collagen-I deposition in IPF, and support a major role of the FAK/Akt pathway in this cooperation.

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“…Prior work by this group Melo et al (2014b) demonstrated that the specific decellularization process affects the elastic modulus tissues, thus it is important to recognize that the additional variable of tissue preparation (frozen intact tissue versus decellularized) is not identical between the two studies and may also account for some of the observed differences. In support of this concept, it is notable that the mechanical properties of acellular ECM tissues are stiffer than normal lung tissue (Gimenez et al, 2017; Ott et al, 2010), though the explanation for this effect remains to be determined. In contrast to these comparable results with sharp tips, Meng et al (2015) reported similar elastic modulus values to those of Liu et al (2016) and our current results using larger spherical tips, but did so with the use of a sharp pyramidal tip (40 nm radius) and relatively thick tissue section.…”

Section: Discussionmentioning

confidence: 98%

Measured pulmonary arterial tissue stiffness is highly sensitive to AFM indenter dimensions

Sicard

Fredenburgh

Tschumperlin

2017

Journal of the Mechanical Behavior of Biomedical Materials

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The mechanical properties of pulmonary tissues are important in normal function and the development of diseases such as pulmonary arterial hypertension. Hence it is critical to measure lung tissue micromechanical properties as accurately as possible in order to gain insight into the normal and pathological range of tissue stiffness associated with development, aging and disease processes. In this study, we used atomic force microscopy (AFM) micro-indentation to characterize the Young’s modulus of small human pulmonary arteries (vessel diameter less than 100 μm), and examined the influence of AFM tip geometry and diameter, lung tissue section thickness and the range of working force applied to the sample on the measured modulus. We observed a significant increase of the measured Young’s modulus of pulmonary vessels (one order of magnitude) associated with the use of a pyramidal sharp AFM tips (20 nm radius), compared to two larger spherical tips (1 and 2.5 μm radius) which generated statistically indistinguishable results. The effect of tissue section thickness (ranging from 10 to 50 microns) on the measured elastic modulus was relatively smaller (<1-fold), but resulted in a significant increase in measured elastic modulus for the thinnest sections (10 micron) relative to the thicker (20 and 50 micron) sections. We also found that the measured elastic modulus depends modestly (again <1-fold), but significantly, on the magnitude of force applied, but only on thick (50 micron) and not thin (10 micron) tissue sections. Taken together these results demonstrate a dominant effect of indenter shape/radius on the measured elastic modulus of pulmonary arterial tissues, with lesser effects of tissue thickness and applied force. The results of this study highlight the importance of AFM parameter selection for accurate characterization of pulmonary arterial tissue mechanical properties, and allow for comparison of literature values for lung vessel tissue mechanical properties measured by AFM across a range of indenter and indentation parameters.

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Elastic properties of hydrogels and decellularized tissue sections used in mechanobiology studies probed by atomic force microscopy

Cited by 29 publications

References 72 publications

Lysyl oxidases regulate fibrillar collagen remodelling in idiopathic pulmonary fibrosis

Lysyl oxidases regulate fibrillar collagen remodelling in idiopathic pulmonary fibrosis

Dysregulated Collagen Homeostasis by Matrix Stiffening and TGF-β1 in Fibroblasts from Idiopathic Pulmonary Fibrosis Patients: Role of FAK/Akt

Measured pulmonary arterial tissue stiffness is highly sensitive to AFM indenter dimensions

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