Exploring cellular mechanics in ARSACS : an AFM-based analysis of elasticity and viscoelasticity

Belo, João Augusto Antunes

http://hdl.handle.net/10400.5/97427

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Autores

Belo, João Augusto Antunes

Orientador(es)

Rodrigues, Mário

Herrera García, Federico

Resumo(s)

Studying cell mechanical properties is crucial for understanding various biological processes and disease mechanisms. Incorporating both elastic and viscoelastic properties into the analysis allows for a more complete understanding of cellular mechanics and supports biomedical applications. While measuring viscoelasticity in cells is challenging due to their complexity and current technological limitations, advances in measurement techniques are enhancing our ability to study these properties. This knowledge is essential for accurately modeling biological systems and has significant implications for understanding health and disease. This study explores the impact of cytoskeletal alterations on single-cell mechanical properties, including stiffness, elasticity, and viscoelasticity. These properties were studied by disrupting the cytoskeleton, particularly the intermediate filaments (IF). This disruption was achieved in two ways: first, using C6Sacs−/− cells, a cellular model for ARSACS (a disease known to impair intermediate filaments); and second, pharmacologically by incubating healthy C6 cells with Withaferin A, a naturally occurring steroidal lactone, that has also shown to cause intermediate filament disruption. To obtain the mechanical properties of the cells, cells were studied using Atomic Force Microscopy (AFM). The data was analysed using Hertz model to obtain the Young’s modulus for approach and retract curves. The Hertz model chosen was the parabolic approximation for a spherical indenter, which was compared to a more robust approximation of a spherical indenter obtained from the Sneddon solution to the contact problem. To obtain the viscoelastic properties, E1, E2, τ and η, the data was analysed using a modified SLS model. Average values for topography, Young’s modulus, and viscoelastic parameters were calculated for wild-type (WT), mutant (MT), WT treated with Withaferin A (WFA), and WT treated with Dimethyl sulfoxide (DMSO). Key differences in cellular mechanics were identified between these groups, providing insights into how cytoskeletal disruptions affect cellular behaviour, particularly in diseases like ARSACS.