Understanding Viscoelasticity Changes in Single Cells using Variable Indentation-Rate Viscoelastic Analysis

Abstract

Increased myocardial stiffness correlates with reduced cardiac performance, arrhythmias and increased mortality, yet the contribution of the cytoskeleton to these mechanical alterations is poorly understood. Our group has recently identified that post-translational modifications of microtubules, particularly detyrosination, correlates with declining cardiac function in hypertrophic cardiomyopathy. To determine if changes in microtubule detyrosination induce changes in cardiomyocyte mechanical properties, we employed a new AFM method, variable indentation-rate viscoelastic analysis, VIVA, to compare the viscoelasticity of isolated rat cardiomyoctes expressing different levels of microtubule detyrosination. Individual cardiomyocytes were either treated with parthenolide or transduced with adenovirus encoding tubulin tyrosine ligase (TTL) to reduce the amount of detyrosination. Alternatively, detyrosination was increased by genetically silencing TTL with shRNA. Using VIVA to determine viscoelasticity, we found a substantial correlation between the detyrosination level and cardiomyocyte viscoelasticity. In particular, increases in detyrosination increased both the stiffness and viscosity of cardiomyocytes, while reduction of detyrosination decreased the stiffness and viscosity roughly 2 fold. These AFM results agree with contractile measurements that show TTL overexpression leads to a higher contractile velocity and faster relaxation time. This can be explained by a lower viscosity of myocytes expressing lower levels of microtubule detyrosination. Our findings identify microtubule detyrosination as a regulator of myocyte stiffness, and suggest that targeting this cytoskeletal modification could be beneficial in certain cardiomyopathies.