Epithelial-Mesenchymal Transition (EMT) is a critical process in embryonic development in which epithelial cells undergo a transdifferentiation into mesenchymal cells. This process is essential for tissue patterning and organization, and it has also been implicated in a wide array of pathologies. While the intracellular signaling pathways that regulate EMT are well-understood, there is increasing evidence that the mechanical properties and composition of the extracellular matrix (ECM) also play a key role in regulating EMT. In turn, EMT drives changes in the mechanics and composition of the ECM, creating a feedback loop that is tightly regulated in healthy tissues, but is often dysregulated in disease. Here we present a review that summarizes our understanding of how ECM mechanics and composition regulate EMT, and how in turn EMT alters ECM mechanics and composition.

Cells respond to mechanical cues from the substrate to which they are attached. These mechanical cues drive cell migration, proliferation, differentiation, and survival. Previous studies have highlighted three specific mechanisms through which substrate stiffness directly alters cell function: increasing stiffness drives 1) larger contractile forces; 2) increased cell spreading and size; and 3) altered nuclear deformation. While studies have shown that substrate mechanics are an important cue, the role of the extracellular matrix (ECM) has largely been ignored. The ECM is a crucial component of the mechanosensing system for two reasons: 1) many ECM fibrils are assembled by application of cell-generated forces, and 2) ECM proteins have unique mechanical properties that will undoubtedly alter the local stiffness sensed by a cell. We specifically focused on the role of the ECM protein fibronectin (FN), which plays a critical role in de novo tissue production. In this study, we first measured the effects of substrate stiffness on human embryonic fibroblasts by plating cells onto microfabricated pillar arrays (MPAs) of varying stiffness. Cells responded to increasing substrate stiffness by generating larger forces, spreading to larger sizes, and altering nuclear geometry. These cells also assembled FN fibrils across all stiffnesses, with optimal assembly occurring at approximately 6 kPa. We then inhibited FN assembly, which resulted in dramatic reductions in contractile force generation, cell spreading, and nuclear geometry across all stiffnesses. These findings suggest that FN fibrils play a critical role in facilitating cellular responses to substrate stiffness.

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Epithelial cells form continuous sheets of cells that exist in tensional homeostasis.Homeostasis is maintained through cell-to-cell adhesions that distribute tension and balance forces between cells and their underlying matrix. Disruption of tensional homeostasis can lead to Epithelial-Mesenchymal Transition (EMT), which is a transdifferentiation process in which epithelial cells adopt a mesenchymal phenotype, where cell-cell adhesion is lost and individual cell migration is acquired. This process is critical during embryogenesis and wound healing, but is also dysregulated in many disease states. To further understand the role of intercellular tension in spatial patterning of epithelial cell monolayers, we developed a multicellular computational model of cell-cell and cell-substrate forces. This work builds on a hybrid Cellular Potts-finite element model to evaluate cell-matrix mechanical feedback of an adherent multicellular cluster. Thermodynamically-constrained cells migrate by generating traction forces on a finite element substrate to minimize the total energy of the system. Junctional forces at cell-cell contacts balance these traction forces, thereby producing a mechanically stable epithelial monolayer. Simulations were compared to in vitro experiments using fluorescence-based junction force sensors in clusters of cells undergoing EMT. Results indicate that the multicellular CPM model can reproduce many aspects of EMT, including epithelial monolayer formation dynamics, changes in cell geometry, and spatial patterning of cell geometry and cell-cell forces in an epithelial colony. Author summaryEpithelial cells line all organs of the human body and act as a protective barrier by forming a continuous sheet. These cells exert force on both their neighboring cells as well as the underlying extracellular matrix, which is a network of proteins that creates the structure of tissues. Here we develop a model that encompasses both cell-cell forces and cell-matrix forces in an epithelial cell sheet. The model accounts for cell migration and proliferation, and regulates how cell-cell adhesions are formed. We demonstrate how the interplay between cell-cell forces and cell-matrix forces can regulate the formation of July 9, 2019 1/27 the epithelial cell sheet, the organization of cells within the sheet, and the pattern of cell geometries and cell forces within the sheet. We compare computational results with experiments in which epithelial cell sheets are disrupted and cell-cell junction forces are measured, and demonstrate that the model captures many aspects of epithelial cell dynamics observed experimentally. 42 cellular geometry. This results in a pattern of traction forces directed towards the cell 43 centroid and proportional to their distance from the cell centroid. These traction forces 44 generate substrate strains which, in addition to cell-cell and cell-matrix interactions, 45 July 9, 2019 2/27 impose a thermodynamic constraint and govern the dynamics of individual cells in the 46 CPM. In the current work, we ...

Currently, there are no high-throughput experiments to quantify all biochemical interactions of small molecules in an organism (human) that is essential to ascertain both toxicity and efficacy of a drug. Furthermore, the synthesis of compound/drug libraries explores diverse chemical space that is unrelated to the disease with no guidance from disease-specific biological pathways (targets and antitargets). Recognizing this unexplored area, we have developed proteome-scale target/antitarget guided lead optimization methods (CANDESIGN) for discovery and de novo design of safe chemical libraries. Selection of target/anti-target networks are identified iteratively using machine learning on experimental data for desired cellular phenotypes in vitro and in vivo. We model interactions with proteomes from different organisms (currently 48,278) to infer homology of compound/drug behavior. Our approach has been verified for drug repurposing, design then synthesis of potent non-toxic anticancer pathway-specific libraries, and compounds that alter specific cell functions in vivo. Repurposed leads are identified from 1 billion predicted compound-proteome interactions with 35% accuracy across nine indications (immunological, metabolic, infectious). Compound libraries synthesized for specific hormone signaling pathways result in picoMolar anticancer leads to combat resistant cancers. Our leads are nontoxic in vitro and in vivo and more efficacious than current treatments on patient-derived tumor xenograft mouse models. Similarly, we designed compounds to specifically change function of Myeloid Derived Suppressor Cells (MDSC) from upregulated gene networks of monocytic MDSCs in the cancer microenvironment. Cancer cell lines (human, mouse, dog) are insensitive to our MDSC-specific library in vitro but exerts antitumor impact in vivo via immunomodulation. We observed decreased frequencies of MDSC expressing suppressive functional markers (iNOS, PD-L1) and increased frequency of IFN-g þ CD8 þ T-cells for antitumor activity. We conclude that proteomescale networks result in safer ''chemical probes'' to effect similar pathways across different diseases therapeutically.