A vacuum-actuated microtissue stretcher for long-term exposure to oscillatory strain within a 3D matrix

Abstract

Although our understanding of cellular behavior in response to extracellular biological and mechanical stimuli has greatly advanced using conventional 2D cell culture methods, these techniques lack physiological relevance. We developed the microtissue vacuum-actuated stretcher (MVAS) to probe cellular behavior within a 3D multicellular environment composed of innate matrix protein, and in response to continuous uniaxial stretch. The MVAS consists of an array of fifty self-assembled microtissues bordered by vacuum chambers. When a vacuum is applied, the microtissues stretch in plane allowing live imaging. The MVAS is highly suitable for biomedical research and pharmaceutical discovery due to a high-throughput array format and scalable fabrication steps outlined in this paper. We validated our approach by characterizing the bulk microtissue strain, the microtissue strain field and single cell strain, and by assessing F-actin expression in response to chronic cyclic strain of 10%. The MVAS was shown to be capable of delivering reproducible dynamic bulk strain amplitudes up to 13% and the strain field had local maxima around each of the cantilevers. The strain at the single cell level was found to be 10.4% less than the microtissue axial strain due to cellular rotation. Chronic cyclic strain produced a 35% increase in F-actin expression consistent with previously observed cytoskeletal reinforcement in 2D cell culture. The MVAS may further our understanding of the reciprocity shared between cells and their environment, which is critical to meaningful biomedical research and successful therapeutic approaches.

Bioengineering a Miniaturized In Vitro 3D Myotube Contraction Monitoring Chip For Modelization of Muscular Dystrophies

“…The classical experimental tools to quantify forces generated by single cells include traction force microscopy (TFM) (Plotnikov et al, 2014)(Lo et al, 2000) (Balaban et al, 2001) or micropillar substrates (Tan et al, 2017) (Ghibaudo et al, 2011) (Trichet et al, 2012). The latter has been adapted to measure forces generated within 3D microtissues (Kural and Billiar 2013) (Boudou et al, 2012) (Kalman et al, 2016) (Walker et al, 2018) (Legant et al, 2009). In the context of myotube formation, it appears as a well-adapted system.…”

Light-driven biological actuators to probe the rheology of 3D microtissues

“…7i-k) after immuno-staining. Altogether, these results suggest that, similarly to long term, external mechanical stimulations, a long term optogenetic conditioning during tissue formation impacts the expression and/or maturation of the actomyosin machinery 11,34 , with the additional ability to do it locally.…”

Light-driven biological actuators to probe the rheology of 3D microtissues

“…7.I-K) after immuno-staining. Altogether, these results suggest that, similarly to long term, external mechanical stimulations, a long term optogenetic conditioning during tissue formation impacts the expression and/or maturation of the actomyosin machinery 11,33 , with the additional ability to do it locally.…”