Quantitative phase imaging unravels new insight into dynamics of mesenchymal and amoeboid cancer cell invasion

Tolde, Ondřej; Gandalovičová, Aneta; Křížová, Aneta; Veselý, Pavel; Chmelík, Radim; Rösel, Daniel; Brábek, Jan

doi:10.1038/s41598-018-30408-7

Cited by 45 publications

(32 citation statements)

References 61 publications

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“…However, detailed analysis of fine protrusions is not possible with the 20 × objective used here. Protrusion-analysis was indeed previously performed on Matrigel/collagen embedded cells, there imaged with 40 × using a QPI-technique called CCHM (coherens-controlled holographic microscopy) 47 .…”

Section: Resultsmentioning

confidence: 99%

“…Some limitations of the DHM system should be noted. As a cell population-based instrument with a 20 × objective, this DHM system has limitations regarding detection and quantification of fine protrusions as possible with other systems 47 . Nor is it possible to distinguish intracellular organelles (besides the nucleus to some degree), something which has been done with other QPI systems 59 .…”

Section: Conclusion and Future Perspectivesmentioning

confidence: 99%

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Exploiting the potential of commercial digital holographic microscopy by combining it with 3D matrix cell culture assays

Hellesvik

Øye

Aksnes

2020

Sci Rep

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3D cell culture assays are becoming increasingly popular due to their higher resemblance to tissue environment. These provide an increased complexity compared to the growth on 2D surface and therefore allow studies of advanced cellular properties such as invasion. We report here on the use of 3D Matrigel cell preparations combined with a particular gentle and informative type of live-cell microscopy: quantitative digital holographic microscopy (DHM), here performed by a commercial software-integrated system, currently mostly used for 2D cell culture preparations. By demonstrating this compatibility, we highlight the possible time-efficient quantitative analysis obtained by using a commercial software-integrated DHM system, also for cells in a more advanced 3D culture environment. Further, we demonstrate two very different examples making use of this advantage by performing quantitative DHM analysis of: (1) wound closure cell monolayer Matrigel invasion assay and (2) Matrigel-trapped single and clumps of suspension cells. For both these, we benefited from the autofocus functionality of digital phase holographic imaging to obtain 3D information for cells migrating in a 3D environment. For the latter, we demonstrate that it is possible to quantitatively measure tumourigenic properties like growth of cell clump (or spheroid) over time, as well as single-cell invasion out of cell clump and into the surrounding extracellular matrix. Overall, our findings highlight several possibilities for 3D digital holographic microscopy applications combined with 3D cell preparations, therein studies of drug response or genetic alterations on invasion capacity as well as on tumour growth and metastasis.

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Section: Resultsmentioning

confidence: 99%

Section: Conclusion and Future Perspectivesmentioning

confidence: 99%

Exploiting the potential of commercial digital holographic microscopy by combining it with 3D matrix cell culture assays

Hellesvik

Øye

Aksnes

2020

Sci Rep

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“…el. indicate that changes in cell mass distribution may play a role in cell motility [47]. On the contrary, A2780 cells showed a significant decrease in cell mass even though the number of cells decreased only slightly.…”

Section: Discussionmentioning

confidence: 89%

Quantitative Phase Dynamics of Cancer Cell Populations Affected by Blue Light

et al. 2020

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Increased exposition to blue light may induce many changes in cell behavior and significantly affect the critical characteristics of cells. Here we show that multimodal holographic microscopy (MHM) within advanced image analysis is capable of correctly distinguishing between changes in cell motility, cell dry mass, cell density, and cell death induced by blue light. We focused on the effect of blue light with a wavelength of 485 nm on morphological and dynamical parameters of four cell lines, malignant PC-3, A2780, G361 cell lines, and the benign PNT1A cell line. We used MHM with blue light doses 24 mJ/cm2, 208 mJ/cm2 and two kinds of expositions (500 and 1000 ms) to acquire real-time quantitative phase information about cellular parameters. It has been shown that specific doses of the blue light significantly influence cell motility, cell dry mass and cell density. These changes were often specific for the malignant status of tested cells. Blue light dose 208 mJ/cm2 × 1000 ms affected malignant cell motility but did not change the motility of benign cell line PNT1A. This light dose also significantly decreased proliferation activity in all tested cell lines but was not so deleterious for benign cell line PNT1A as for malignant cells. Light dose 208 mJ/cm2 × 1000 ms oppositely affected cell mass in A2780 and PC-3 cells and induced different types of cell death in A2780 and G361 cell lines. Cells obtained the least damage on lower doses of light with shorter time of exposition.

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“…Thus far, 2D QPI techniques, such as digital holographic microscopy, have been applied to the study of cell migration or wound healing assays (39)(40)(41)(42). Although the use of 2D QPI techniques has included label-free and quantitative imaging capabilities, only limited 2D optical thickness information can be measured, and the 3D morphology of individual cells and subcellular structures is not accessible.…”

Section: Resultsmentioning

confidence: 99%

Three-Dimensional Label-Free Imaging and Quantification of Migrating Cells during Wound Healing

Lee

Hugonnet

Park

et al. 2020

Preprint

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The 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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Quantitative phase imaging unravels new insight into dynamics of mesenchymal and amoeboid cancer cell invasion

Cited by 45 publications

References 61 publications

Exploiting the potential of commercial digital holographic microscopy by combining it with 3D matrix cell culture assays

Exploiting the potential of commercial digital holographic microscopy by combining it with 3D matrix cell culture assays

Quantitative Phase Dynamics of Cancer Cell Populations Affected by Blue Light

Three-Dimensional Label-Free Imaging and Quantification of Migrating Cells during Wound Healing

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