Research Techniques Made Simple: Analysis of Collective Cell Migration Using the Wound Healing Assay

ABSTRACTThe wound healing assay provides essential information about collective cell migration and cell-to-cell interactions. It is a simple, effective, and widely used tool for observing the effect of numerous chemical treatments on wound healing speed. To perform and analyze a wound healing assay, various imaging techniques have been utilized. However, image acquisition and analysis are often limited in two-dimensional space or require the use of exogenous labeling agents. Here, we present a method for imaging large-scale wound healing assays in a label-free and volumetric manner using optical diffraction tomography (ODT). We performed quantitative high-resolution three-dimensional (3D) analysis of cell migration over a long period without difficulties such as photobleaching or phototoxicity. ODT enables the reconstruction of the refractive index (RI) tomogram of unlabeled cells, which provides both structural and biochemical information about the individual cell at subcellular resolution. Stitching multiple RI tomograms enables long-term (24 h) and large field-of-view imaging (> 800 × 400 μm2) with a lateral resolution of 110 nm. We demonstrated the thickness changes of leading cells and studied the effects of cytochalasin D. The 3D RI tomogram also revealed increased RI values in leading cells compared to lagging cells, suggesting the formation of a highly concentrated subcellular structure.STATEMENT OF SIGNIFICANCEThe wound healing assay is a simple but effective tool for studying collective cell migration (CCM) that is widely used in biophysical studies and high-throughput screening. However, conventional imaging and analysis methods only address two-dimensional properties in a wound healing assay, such as gap closure rate. This is unfortunate because biological cells are complex 3D structures, and their dynamics provide significant information about cell physiology. Here, we presented three-dimensional (3D) label-free imaging for wound healing assays and investigated the 3D dynamics of CCM. High-resolution subcellular structures as well as their collective dynamics were imaged and analyzed quantitatively. Our label-free quantitative 3D analysis method provides a unique opportunity to study the behavior of migrating cells during the wound healing process.

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A Wound-Healing Assay Based on Ultraviolet Light Ablation

SLAS TECHNOLOGY Translating Life Sciences Innovation ◽

10.1177/2211068216646741 ◽

2016 ◽

Vol 22 (1) ◽

pp. 36-43 ◽

Cited By ~ 3

Author(s):

Shang-Ying Wu ◽

Yung-Shin Sun ◽

Kuan-Chen Cheng ◽

Kai-Yin Lo

Keyword(s):

Wound Healing ◽

Cell Migration ◽

Healing Process ◽

Cell Monolayer ◽

Wound Healing Process ◽

Collective Cell Migration ◽

Wound Healing Assay ◽

Physiological Processes ◽

Free Region ◽

A Cell

Collective cell migration plays important roles in many physiological processes such as embryonic development, tissue repair, and angiogenesis. A “wound” occurs when epithelial cells are lost and/or damaged due to some external factors, and collective cell migration takes place in the following wound-healing process. To study this cellular behavior, various kinds of wound-healing assays are developed. In these assays, a “wound,” or a “cell-free region,” is created in a cell monolayer mechanically, chemically, optically, or electrically. These assays are useful tools in studying the effects of certain physical or chemical stimuli on the wound-healing process. Most of these methods have disadvantages such as creating wounds of different sizes or shapes, yielding batch-to-batch variation, and damaging the coating of the cell culture surface. In this study, we used ultraviolet (UV) lights to selectively kill cells and create a wound out of a cell monolayer. A comparison between the current assay and the traditional scratch assay was made, indicating that these two methods resulted in similar wound-healing rates. The advantages of this UV-created wound-healing assay include fast and easy procedure, high throughput, and no direct contact to cells.

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Hierarchical modeling of mechano-chemical dynamics of epithelial sheets across cells and tissue

Scientific Reports ◽

10.1038/s41598-021-83396-6 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Yoshifumi Asakura ◽

Yohei Kondo ◽

Kazuhiro Aoki ◽

Honda Naoki

Keyword(s):

Wound Healing ◽

Cell Migration ◽

Continuum Model ◽

Tissue Level ◽

Chemical Signals ◽

Cellular Level ◽

Mathematical Framework ◽

Collective Cell Migration ◽

The Continuum ◽

Cell Cell

AbstractCollective cell migration is a fundamental process in embryonic development and tissue homeostasis. This is a macroscopic population-level phenomenon that emerges across hierarchy from microscopic cell-cell interactions; however, the underlying mechanism remains unclear. Here, we addressed this issue by focusing on epithelial collective cell migration, driven by the mechanical force regulated by chemical signals of traveling ERK activation waves, observed in wound healing. We propose a hierarchical mathematical framework for understanding how cells are orchestrated through mechanochemical cell-cell interaction. In this framework, we mathematically transformed a particle-based model at the cellular level into a continuum model at the tissue level. The continuum model described relationships between cell migration and mechanochemical variables, namely, ERK activity gradients, cell density, and velocity field, which could be compared with live-cell imaging data. Through numerical simulations, the continuum model recapitulated the ERK wave-induced collective cell migration in wound healing. We also numerically confirmed a consistency between these two models. Thus, our hierarchical approach offers a new theoretical platform to reveal a causality between macroscopic tissue-level and microscopic cellular-level phenomena. Furthermore, our model is also capable of deriving a theoretical insight on both of mechanical and chemical signals, in the causality of tissue and cellular dynamics.

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Equation-Based Models of Wound Healing and Collective Cell Migration

Complex Systems and Computational Biology Approaches to Acute Inflammation ◽

10.1007/978-3-030-56510-7_11 ◽

2020 ◽

pp. 199-221

Author(s):

Julia Arciero ◽

David Swigon

Keyword(s):

Wound Healing ◽

Cell Migration ◽

Collective Cell Migration

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A Method for Quantitative Analysis of the Kinematics of Fibroblast Migration in a Monolayer Wound Model

ASME 2011 Summer Bioengineering Conference, Parts A and B ◽

10.1115/sbc2011-53070 ◽

2011 ◽

Author(s):

Gil Topman ◽

Orna Sharabani-Yosef ◽

Amit Gefen

Keyword(s):

Wound Healing ◽

Cell Migration ◽

Wound Healing Assay ◽

Complete Coverage ◽

Time Intervals ◽

Fibroblast Migration ◽

Wound Model ◽

Regular Time

A wound healing assay is simple but effective method to study cell migration in vitro. Cell migration in vitro was found to mimic migration in vivo to some extent [1,2]. In wound healing assays, a “wound” is created by either scraping or mechanically crushing cells in a monolayer, thereby forming a denuded area. Cells migrate into the denuded area to complete coverage, and thereby “heal” the wound. Micrographs at regular time intervals are captured during such experiments for analysis of the process of migration.

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Viscoelasticity and cell jamming state transition

10.1101/2021.03.19.436195 ◽

2021 ◽

Author(s):

Ivana Pajic-Lijakovic ◽

Milan Milivojevic

Keyword(s):

Residual Stress ◽

Wound Healing ◽

Cell Migration ◽

State Transition ◽

State Transitions ◽

Collective Cell Migration ◽

Biological Processes ◽

Cell Mobility ◽

Controversial Topic ◽

The Impact

Although collective cell migration (CCM) is a highly coordinated migratory mode, perturbations in the form of jamming state transitions and vice versa often occur even in 2D. These perturbations are involved in various biological processes, such as embryogenesis, wound healing and cancer invasion. CCM induces accumulation of cell residual stress which has a feedback impact to cell packing density. Density-mediated change of cell mobility influences the state of viscoelasticity of multicellular systems and on that base the jamming state transition. Although a good comprehension of how cells collectively migrate by following molecular rules has been generated, the impact of cellular rearrangements on cell viscoelasticity remains less understood. Thus, considering the density driven evolution of viscoelasticity caused by reduction of cell mobility could result in a powerful tool in order to address the contribution of cell jamming state transition in CCM and help to understand this important but still controversial topic. In addition, five viscoelastic states gained within three regimes: (1) convective regime, (2) conductive regime, and (3) damped-conductive regime was discussed based on the modeling consideration with special emphasis of jamming and unjamming states.

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