Non-invasive single-cell biomechanical analysis using live-imaging datasets

Abstract

The physiological state of a cell is governed by a multitude of processes and can be described by a combination of mechanical, spatial and temporal properties. Quantifying cell dynamics at multiple scales is essential for comprehensive studies of cellular function, and remains a challenge for traditional end-point assays. We introduce an efficient, non-invasive computational tool that takes time-lapse images as input to automatically detect, segment and analyze unlabeled live cells; the program then outputs kinematic cellular shape and migration parameters, while simultaneously measuring cellular stiffness and viscosity. We demonstrate the capabilities of the program by testing it on human mesenchymal stem cells (huMSCs) induced to differentiate towards the osteoblastic (huOB) lineage, and T-lymphocyte cells (T cells) of naïve and stimulated phenotypes. The program detected relative cellular stiffness differences in huMSCs and huOBs that were comparable to those obtained with studies that utilize atomic force microscopy; it further distinguished naïve from stimulated T cells, based on characteristics necessary to invoke an immune response. In summary, we introduce an integrated tool to decipher spatiotemporal and intracellular dynamics of cells, providing a new and alternative approach for cell characterization.

Deconstructing Immune Microenvironments of Lymphoid Tissues for Reverse Engineering

“…The widely used combination of a biocompatible polydimethylsiloxane (PDMS) microfluidic chip on a glass base also allows long‐term live imaging of cells without the loss of migration information. In fact, through novel quantification methods more information can be gathered by applying fluorescent molecular probes or by using genetically modified cells expressing proteins with fluorescent tags, during live cell imaging.…”

“…Transwells assays are also an end‐point method whereby cells are quantified only after they have migrated over a given amount of time . In addition to losing much data during the migration, including persistence of direction, velocity of migration, and pausing behavior, time and consumable resources have to be a consideration for optimization of time points of biological relevance . Polymerization time of hydrogels within Transwells is also an issue, especially for larger well formats.…”

“…The effects of glycation of collagen are fascinating and present a nontoxic method to increase compressive modulus of collagen matrices . With microfluidics, single cell analysis is possible and therefore able to capture immune cell heterogeneity, contrasting bulk population evaluation when using Transwells . There has been an explosion of different microfluidics platforms for tissue‐based bioassays; here, we focus on platforms pertaining to interfacing immune cells to an in vivo‐like cellular microenvironment.…”

Shaping the Cell and the Future: Recent Advancements in Biophysical Aspects Relevant to Regenerative Medicine

“…For example, eccentricity and filopodia were shown to robust classifiers of myogenic differentiation [27]. Pearson et al combined a unique approach which simultaneously measured cell morphology parameters, migration trajectories, as well as viscoelasticity parameters and cell stiffness for characterizing MSC differentiated phenotypes and naïve vs. stimulated T cells [28].…”

Quantification of sterol-specific response in human macrophages using automated imaged-based analysis

“…To address these issues, we present and discuss the output of a newly developed, customized image-processing method, designed to identify individual cells via boundary detection (from populations and clusters) by applying watershed image segmentation, implemented in MATLAB software (Mathworks Inc.) (see also reference [ 23 ]), to quickly extract and analyze data from bright-field and fluorescence images to provide single-cell properties, such as area, extent of staining and intracellular heterogeneity as well as eccentricity, for over one thousand cells in 25 different chemical environments.…”