Moreover, microtubule and vimentin density decreased from the center to the periphery of the cell. methods, lower table) for radial orientation functions (Fig 3) of actin, microtubules and vimentin TPT-260 (Dihydrochloride) at an angle of 90 towards stretch. Sample sizes are given in Fig 3 caption.(XLS) pone.0210570.s003.xls (47K) GUID:?D8E875C4-E9DF-4B10-9DD6-0090AEF6B356 S2 Dataset: Statistical significances (KS test, upper table) and effect sizes (see Materials and methods, lower table) for intrinsic DPP4 radial orientation functions (Fig 5) of actin, microtubules and vimentin at an angle of 90 towards stretch. Sample sizes are given in Fig 3 caption.(XLS) pone.0210570.s004.xls (36K) GUID:?28FC4CBF-4974-410A-A7CB-D7D374106179 S3 Dataset: Statistical significances (KS test) and effect sizes (see Materials and methods) for any comparison of radial orientation functions of the TPT-260 (Dihydrochloride) actin cytoskeleton (values at 90, see Fig 12) of cells treated with nocodazole and control cells treated with DMSO alone. Moreover, same analysis for intrinsic radial orientation of actin, i.e., alignment of correlograms before averaging.(XLSX) pone.0210570.s005.xlsx (10K) GUID:?B465E5A4-F7A7-4989-9E0D-84474B357F06 Data Availability StatementThe data underlying this study have been uploaded to the Image Data Resource repository and are accessible using the following URL: https://doi.org/10.17867/10000119. Abstract In mammalian cells, actin, TPT-260 (Dihydrochloride) microtubules, and various types of cytoplasmic intermediate TPT-260 (Dihydrochloride) filaments respond to external stretching. Here, we investigated the underlying processes in endothelial cells plated on soft substrates from silicone elastomer. After cyclic stretch (0.13 Hz, 14% strain amplitude) for periods ranging from 5 min to 8 h, cells were fixed and double-stained for microtubules and either actin or vimentin. Cell images were analyzed by a two-step routine. In the first step, micrographs were segmented for potential fibrous structures. In the second step, the producing binary masks were auto- or cross-correlated. Autocorrelation of segmented images provided a sensitive and objective measure of orientational and translational order of the different cytoskeletal systems. Aligning of correlograms from individual cells removed the influence of only partial alignment between cells and enabled determination of intrinsic cytoskeletal order. We found that cyclic stretching affected the actin cytoskeleton most, microtubules less, and vimentin mostly only via reorientation of the whole cell. Pharmacological disruption of microtubules experienced TPT-260 (Dihydrochloride) barely any influence on actin ordering. The similarity, i.e., cross-correlation, between vimentin and microtubules was much higher than the one between actin and microtubules. Moreover, prolonged cyclic stretching slightly decoupled the cytoskeletal systems as it reduced the cross-correlations in both cases. Finally, actin and microtubules were more correlated at peripheral regions of cells whereas vimentin and microtubules correlated more in central regions. Introduction Within the organism most tissue cells are permanently exposed to mechanical deformation. For example, cells of the myocard experience strains of up to 30% with each heart beat [1] and cells lining the alveoli of the lung experience comparable strains during breathing [2]. Even larger strains, of up to 80%, have been inferred for soft tissues of the shoulder as a result of transporting a backpack [3]. Consequently, most tissues exhibit structures that are clearly adapted to these intense mechanical deformations. Obviously, cells embedded in these tissues must sense the mechanical signal and adapt to it. In cases where these cellular adaptations to mechanical strain are compromised or maladapted, severe pathological disorders like enlargement of cerebral aneurysms [4] and right heart failure in response to pulmonary arterial hypertension [5] occur. Thus, the interplay of tissue cells and mechanical signals is usually of high interest. Unraveling the processes underlying cellular reactions to deformation is usually a challenging task, as it is very difficult to apply well-defined mechanical signals and to quantify the ensuing responses. This challenge can be met in experiments on cells cultivated on elastomeric substrates undergoing uniaxial or biaxial strain [6C10] because here substrate strain can be cautiously controlled and cellular reactions can be well analyzed by most techniques of molecular cell biology. Cell reactions to applied stretch have recently been examined [11]. The.