Abstract
Epithelial–mesenchymal transition (EMT) is a complex series of cellular reprogramming events culminating in striking alterations in morphology towards an invasive mesenchymal phenotype. Increasingly, evidence suggests that EMT exerts a pivotal role in pathophysiological situations including fibrosis and cancer. Core to these dynamical changes in cellular polarity and plasticity is discrete modifications in cytoskeletal structure. In particular, newly established actin-stress fibres supplant a preceding system of highly organised cortical actin. Although cumulative studies have contributed to elucidation of the detailed signalling pathways that underpin this elaborate molecular process, there remains a deficiency regarding its precise contribution to cellular biomechanics. The advent of atomic force microscopy (AFM) and high-content analysis (HCA) provides two innovative technologies for dissecting the relationship between EMT-related morphological and structural alterations and cell mechanical properties. AFM permits acquisition of high resolution topographical images and detailed analysis of cellular viscoelasticity while HCA facilitates a comprehensive and perspicacious assessment of morphological changes. In combination, they offer the possibility of novel insights into the dynamic traits of transitioning cells. Herein, a detailed protocol describing AFM and HCA techniques for evaluation of transforming growth factor-β1-induced EMT of alveolar epithelial cells is provided.
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Acknowledgements
STB is funded by an IRCSET Government of Ireland Postgraduate Scholarship in Science, Engineering and Technology. This work has been funded in part by a Strategic Research Cluster grant (07/SRC/B1154) under the National Development Plan co-funded by EU Structural Funds and Science Foundation Ireland.
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Buckley, S.T., Davies, A.M., Ehrhardt, C. (2011). Atomic Force Microscopy and High-Content Analysis: Two Innovative Technologies for Dissecting the Relationship Between Epithelial–Mesenchymal Transition-Related Morphological and Structural Alterations and Cell Mechanical Properties. In: O'Driscoll, L. (eds) Gene Expression Profiling. Methods in Molecular Biology, vol 784. Humana Press. https://doi.org/10.1007/978-1-61779-289-2_14
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DOI: https://doi.org/10.1007/978-1-61779-289-2_14
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