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Abstract IntroductionTraction force microscopy (TFM) is a widely used technique to measure cell contractility on compliant substrates that mimic the stiffness of human tissues. For every step in a TFM workflow, users make choices which impact the quantitative results, yet many times the rationales and consequences for making these decisions are unclear. We have found few papers which show the complete experimental and mathematical steps of TFM, thus obfuscating the full effects of these decisions on the final output. MethodsTherefore, we present this “Field Guide” with the goal to explain the mathematical basis of common TFM methods to practitioners in an accessible way. We specifically focus on how errors propagate in TFM workflows given specific experimental design and analytical choices. ResultsWe cover important assumptions and considerations in TFM substrate manufacturing, substrate mechanical properties, imaging techniques, image processing methods, approaches and parameters used in calculating traction stress, and data-reporting strategies. ConclusionsBy presenting a conceptual review and analysis of TFM-focused research articles published over the last two decades, we provide researchers in the field with a better understanding of their options to make more informed choices when creating TFM workflows depending on the type of cell being studied. With this review, we aim to empower experimentalists to quantify cell contractility with confidence. 

Abstract The epithelial microenvironment is incredibly dynamic, subjected to mechanical cues including cyclic stretch. While cyclic cell stretching platforms have revealed epithelial cell reorientation and gap formation, few studies have investigated the long-term effects of cyclic stretch on cell migration. We measured the migratory response of the epithelium to a range of physiologically relevant frequencies and stretch. Our results indicate that lower stretch frequencies (i.e., 0.1 Hz) suppress epithelial migration, accompanied by cell reorientation and high cell shape solidity. We found that this response is also accompanied by increased recruitment of vinculin to cell-cell contacts, and this recruitment is necessary to suppress cell movements. These results confirm the mechanosensitive nature of vinculin within the adherens junction, but independently reveal a novel mechanism of low frequency stress response in supporting epithelial integrity by suppressing cell migration. 

Abstract The heart is a dynamic pump whose function is influenced by its mechanical properties. The viscoelastic properties of the heart, i.e., its ability to exhibit both elastic and viscous characteristics upon deformation, influence cardiac function. Viscoelastic properties change during heart failure (HF), but direct measurements of failing and non-failing myocardial tissue stress relaxation under constant displacement are lacking. Further, how consequences of tissue remodeling, such as fibrosis and fat accumulation, alter the stress relaxation remains unknown. To address this gap, we conducted stress relaxation tests on porcine myocardial tissue to establish baseline properties of cardiac tissue. We found porcine myocardial tissue to be fast relaxing, characterized by stress relaxation tests on both a rheometer and microindenter. We then measured human left ventricle (LV) epicardium and endocardium tissue from non-failing, ischemic HF and non-ischemic HF patients by microindentation. Analyzing by patient groups, we found that ischemic HF samples had slower stress relaxation than non-failing endocardium. Categorizing the data by stress relaxation times, we found that slower stress relaxing tissues were correlated with increased collagen deposition and increased α-smooth muscle actin (α-SMA) stress fibers, a marker of fibrosis and cardiac fibroblast activation, respectively. In the epicardium, analyzing by patient groups, we found that ischemic HF had faster stress relaxation than non-ischemic HF and non-failing. When categorizing by stress relaxation times, we found that faster stress relaxation correlated with Oil Red O staining, a marker for adipose tissue. These data show that changes in stress relaxation vary across the different layers of the heart during ischemic versus non-ischemic HF. These findings reveal how the viscoelasticity of the heart changes, which will lead to better modeling of cardiac mechanics for in vitro and in silico HF models. 

The behavior of cells is governed by signals originating from their local environment, including mechanical forces exerted on the cells. Forces are transduced by mechanosensitive proteins, which can impinge on signaling cascades that are also activated by growth factors. We investigated the cross-talk between mechanical and biochemical signals in the regulation of intracellular signaling networks in epithelial monolayers. Phosphoproteomic and transcriptomic analyses on epithelial monolayers subjected to mechanical strain revealed the activation of extracellular signal–regulated kinase (ERK) downstream of the epidermal growth factor receptor (EGFR) as a predominant strain-induced signaling event. Strain-induced EGFR-ERK signaling depended on mechanosensitive E-cadherin adhesions. Proximity labeling showed that the metalloproteinase ADAM17, an enzyme that mediates shedding of soluble EGFR ligands, was closely associated with E-cadherin. A probe that we developed to monitor ADAM-mediated shedding demonstrated that mechanical strain induced ADAM activation. Mechanically induced ADAM activation was essential for mechanosensitive, E-cadherin–dependent EGFR-ERK signaling. Together, our data demonstrate that mechanical strain transduced by E-cadherin adhesion triggers the shedding of EGFR ligands that stimulate downstream ERK activity. Our findings illustrate how mechanical signals and biochemical ligands can operate within a linear signaling cascade. 

The feedback between mechanical and chemical signals plays a key role in controlling many biological processes and collective cell behavior. Here we focus on the emergence of spatiotemporal density waves in a one-dimensional “cell train.” Combining a minimal theoretical model with experiments on MDCK epithelial cells confined to a linear pattern, we examine the spontaneous oscillations driven by feedback between myosin activation and mechanical deformations, as well as their effect on the response of the tissue to externally applied deformations. We show that the nature and frequency of spontaneous oscillations is controlled by the size of the cell train, with a transition from size-dependent standing waves to intrinsic spontaneous waves at the natural frequency of the tissue. The response to external boundary perturbations exhibits a resonance at this natural frequency, providing a possible venue for inferring the mechanochemical couplings that control the tissue behavior from rheological experiments.