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

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.