Force-driven evolution of mesoscale structure in engineered 3D microtissues and the modulation of tissue stiffening

Abstract

The complex structures of tissues determine their mechanical strength. In engineered tissues formed through self-assembly in a mold, artificially imposed boundary constraints have been found to induce anisotropic clustering of the cells and the extracellular matrix in local regions. To understand how such tissue remodeling at the intermediate length-scale (mesoscale) affects tissue stiffening, we used a novel microtissue mechanical testing system to manipulate the remodeling of the tissue structures and to measure the subsequent changes in tissue stiffness. Microtissues were formed through cell driven self-assembly of collagen matrix in arrays of micro-patterned wells, each containing two flexible micropillars that measured the microtissues’ contractile forces and also their elastic moduli via magnetic actuation. We manipulated tissue remodeling by inducing myofibroblast differentiation with TGF-β1, by varying the micropillar spring constants or by blocking cell contractility with blebbistatin and collagen cross-linking with BAPN. We showed that increased anisotropic compaction of the collagen matrix, caused by increased micropillar spring constant or elevated cell contraction force, contributed to tissue stiffening. Conversely, collagen matrix and tissue stiffness were not affected by inhibition of cell-generated contraction forces.. Together, these measurements showed that mesoscale tissue remodeling is an important middle step linking tissue compaction forces and tissue stiffening.

Molecular Mechanism Responsible for Fibronectin-controlled Alterations in Matrix Stiffness in Advanced Chronic Liver Fibrogenesis

“…Consistent with previous in vitro and in vivo observations (11,56,57), our findings indicate that inhibition of collagen crosslinking by BAPN affects TGF-␤-induced structural and mechanical integrity of collagenous ECM networks in Fn-null hepatic stellate cells. We have demonstrated functional links between tissue/organ functions and regulatory mechanisms of LOX underlying chronic fibrotic disease such as liver fibrosis/ cirrhosis.…”

Supersoft lithography: candy-based fabrication of soft silicone microstructures

“…The effective stiffness of the cantilever is reduced by decreasing the geometric cross-section of the cantilever, enabling the measurement of small contractile forces. Although extremely useful 35,36 , the dog-bone shaped tissues may exhibit strong necking profiles 37 leading to varied cross-sectional areas, and thus, significant variations in internal tissue stresses.…”

Magnetically actuated cell-laden microscale hydrogels for probing strain-induced cell responses in three dimensions

“…[18][19][20] In these systems, 3D microtissues were created and constrained between patterned poly(dimethylsiloxane) cantilevers attached to a magnetically responsive microsphere. Such microtissue force gauges have allowed fast diffusion of soluble factors to multiple samples simultaneously, offered the capability to apply external mechanical loading to the specimens and enabled measurement of the contractility of multicellular tissues under applied force.…”