Cell elasticity with altered cytoskeletal architectures across multiple cell types

Abstract

The cytoskeleton is primarily responsible for providing structural support, localization and transport of organelles, and intracellular trafficking. The structural support is supplied by actin filaments, microtubules, and intermediate filaments, which contribute to overall cell elasticity to varying degrees. We evaluate cell elasticity in five different cell types with drug-induced cytoskeletal derangements to probe how actin filaments and microtubules contribute to cell elasticity and whether it is conserved across cell type. Specifically, we measure elastic stiffness in primary chondrocytes, fibroblasts, endothelial cells (HUVEC), hepatocellular carcinoma cells (HUH-7), and fibrosarcoma cells (HT 1080) subjected to two cytoskeletal destabilizers: cytochalasin D and nocodazole, which disrupt actin and microtubule polymerization, respectively. Elastic stiffness is measured by atomic force microscopy (AFM) and the disruption of the cytoskeleton is confirmed using fluorescence microscopy. The two cancer cell lines showed significantly reduced elastic moduli values (~0.5kPa) when compared to the three healthy cell lines (~2kPa). Non-cancer cells whose actin filaments were disrupted using cytochalasin D showed a decrease of 60-80% in moduli values compared to untreated cells of the same origin, whereas the nocodazole-treated cells showed no change in elasticity. Overall, we demonstrate actin filaments contribute more to elastic stiffness than microtubules but this result is cell type dependent. Cancer cells behaved differently, exhibiting increased stiffness as well as stiffness variability when subjected to nocodazole. We show that disruption of microtubule dynamics affects cancer cell elasticity, suggesting therapeutic drugs targeting microtubules be monitored for significant elastic changes.

Mapping intracellular mechanics on micropatterned substrates

“…Similarly, the local contribution of actin in intracellular rheology can be investigated in detail using our method, as best illustrated by the differences we observed between the effects of two actin-destabilizing drugs, LatA and CytoD. Although actin destabilization is mostly thought to soften the cell interior, its precise effect on intracellular mechanics is still not clear (44) and probably depends on the cell type (61). Surprisingly, although in LatA-treated cells the shear modulus dropped and bead diffusion was facilitated, we did not detect any effect upon treatment with CytoD (Fig.…”

Intracellular nanoparticle dynamics affected by cytoskeletal integrity

“…We found that the mobility of QDs within fibroblasts was less affected by nocodazole treatment than cytochalasin D treatement, which is consistent with prior studies showing that the mechanical properties of fibroblasts do not change significantly when exposed to nocodazole. 2 From the particle tracks, MSD was determined using MATLAB 43 . MSDs for QDs within fibroblasts, fibrosarcoma cells, and cytochalasin D treated fibroblasts are shown in Figure 1d–f respectively.…”

“…A contributor to this confinement is the cytoskeleton itself, which features a mesh-like network with open spaces that direct the diffusion of macromolecules. The structural role of the actin and microtubule network within cell mechanics has been previously investigated 2–4 and together with the whole cytoskeleton network, cytoplasmic organelles, and molecular crowding, serves as obstacles to non-directed intracellular motion. Intracellular particle diffusion has become increasingly important within nanomedicine, where nano-scale photodynamic therapies 5–7 and gene or drug delivery systems 8–11 have made significant advances in disease treatment options.…”

“…Here we discuss the influence of cytoskeletal integrity, namely filamentous actin and microtubules, on intracellular nanoparticle diffusion manifested in different cell conditions. Motivated by prior elasticity measurements, 2 we identify differences between two cell types, one cancer (fibrosarcoma cells HT 1080) and one non-cancer (human dermal fibroblasts), as well as drug-altered cytoskeletons within those two lines by cytochalasin D treatment (disrupts filamentous actin) and nocodazole treatment (disrupts microtubules). Within the cytoplasm of these cells, the nanoparticles act as internal probes that reflect the influence of the cytoskeleton and its caged network on transport.…”

Characterization of the mechanical properties of resected porcine organ tissue using optical fiber photoelastic polarimetry

“…For example, the large sensor area in rheology systems results in a poor spatial resolution, on the order of centimeters. To address this, researchers turned to Atomic Force Microscopy (AFM) [3][4][5]. However, destructive sample preparation is required, preventing further imaging and molecular analysis.…”