Characterizing Cell-Scale Architecture-Dependent Mechano-Adaptation In Vitro

Abstract

Mechano-adaptation is the process whereby cells or tissues change their structure or function following mechanical perturbation. Necessarily, changes in structure and function impact mechanical state, and these changes influence subsequent changes in structure and function, establishing a feedback loop between physical state and physiologic response. It has been posited that mechano-adaptation is driven by mechanical stress such that changes in structure and function happen when cell or tissue stress deviates from a homeostatic setpoint. While this notion of “tensional homeostasis” has been demonstrated across length scales and tissue types and contexts, little work has explored the role of the cytoskeleton – the primary mechanical constituent of biological tissue – in this process. Here, I use Cellular Microbiaxial Stretching (CµBS) in tandem with traction force microscopy and immunocytochemistry to explore mechano-adaptation in single cells and multicellular tissues micropatterned into various shapes to determine the role of cytoskeletal architecture in stress response. First, I demonstrate that, following a step change in strain held for 24 h, single vascular smooth muscle cell stresses increase and over time, stresses equilibrate at their pre-stretch values in all strain magnitude and cell shape conditions tested. I formulated a continuum-based model that accounted for experimentally measured orientations of the actomyosin cytoskeleton, and attributed stress evolution to actomyosin targeted growth, and found this model to strongly capture experimental behavior. Next, I sought to determine if this same approach could be used to describe mechano-adaptive behavior in multicellular tissues. I micropatterned Madin-Darby canine kidney epithelial cells into microtissues of various shape, subjected these tissues to a step change in strain held for 24 h, and tracked tissue architecture, proliferation, and stresses over the duration of the experiment. Although the cytoskeleton underwent little realignment over time, stresses decreased over time in tissues with or without stretch, which led me to suspect that proliferation resulted in a decrease in tissue stress. I found that tissues proliferated at a rate that did not depend on tissue shape or external stretch, but that this proliferation resulted in a decrease in tissue stress that resulted from increased cell packing and change in tissue geometry. I then modified the model used to capture single cell mechano-adaptation to include proliferation and found that this model recapitulated changes in tissue geometry and stress when accounting for experimentally measured cytoskeletal alignment and isotropic volumetric growth. Lastly, I offer insight into preliminary work that applied these methods towards investigating mechano-adaptation of the embryonic heart in vitro. This work demonstrates that accounting for the organization of cytoskeletal architecture in a continuum-based model of mechano-adaptation is sufficient to capture stress response across length scales.