The interplay of extracellular fluid viscosity, stiffness and confinement in regulating cell responses

Summary- Cell migration is a fundamental biological process underlying diverse (patho)physiological events, such as embryonic development, immune response, and disease progression. Cells in vivo travel through different tissues of varying extracellular fluid viscosities. Viscosity is thus a physiologically relevant physical cue, which has been largely overlooked. We were the first to establish that elevated, albeit physiological, levels of viscosity regulate tumor epithelial cell migration and invasion in vitro and in vivo. Cells not only sense and respond to fluid viscosity but also develop memory. Yet, our understanding of how memory is imprinted on epithelial cells and what changes viscosity imparts to the cell’s molecular machinery and chromatin architecture represent major unanswered questions of fundamental and potential translational value. Along these lines, how fluid viscosity affects the function of other cell types, such as immune cells, as well as their crosstalk with tumor epithelial cells, is currently unknown. In addition to fluid viscosity, cells in vivo are also exposed to other important physical cues, such as microenvironmental stiffness and confinement, which all trigger intracellular responses that allow cells to adapt to the diverse landscapes they encounter. We were the first to demonstrate that confinement alone regulates epithelial cell migration mechanisms. Intriguing preliminary data reveal that microchannel stiffness alters cell migration mechanisms in confinement. Yet, the interplay among fluid viscosity, confinement and stiffness on cell function represents an open and unaddressed question. To develop a comprehensive understanding of how physical cues modulate cell responses, we will examine the cell as whole using a holistic and integrated approach. Although prior work has established that the cell nucleus and plasma membrane molecules, such as mechanosensitive ion channels (MICs), sense and respond to mechanical cues, our understanding of the crosstalk between the nucleus and ion channel-mediated mechanosensation is at best limited. We are uniquely positioned to address all these key gaps in the literature by employing a holistic approach integrating cutting-edge in vitro models (hydrogel-based microfluidic channels of precisely tunable stiffness and degree of confinement) and sophisticated techniques (3-dimensional (3D) traction force microscopy, optogenetics, high-throughput sequencing etc) with the chick embryo model. Taken together, our proposed work over the next five years will generate new conceptual information on how key mechanical cues are sensed, interpreted and imprinted on cells via the crosstalk among the cell cytoskeleton, the nucleus and chromatin architecture, and MIC mechanosensing. Our work will unravel novel signaling mechanisms that may lead to the discovery of new therapeutic targets potentially applicable to various pathologies, like cancer, where the elevated viscosity of the local microenvironment has emerged as a key, but understudied, modulator.