Label‐free high temporal resolution assessment of cell proliferation using digital holographic microscopy

Janicke, Birgit; Kårsnäs, Andreas; Egelberg, Peter; Alm, Kersti

doi:10.1002/cyto.a.23108

Cited by 28 publications

(23 citation statements)

References 67 publications

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“…With its ability to detect single cells, DHM is well qualified to measure cell proliferation 41,51 . We additionally analyzed cell proliferation using the same images and found that there was a visually lower proliferation rate for cells in 50% MG compared to uncoated surface and 1% Matrigel (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…When these waves are rejoined with the reference beam, an interference pattern, a hologram, is detected by the digital image sensor. Based on the hologram, a cell image or holographic image is reconstructed using a computer algorithm 51 . This quantitative phase imaging method is used for analysis and quantification of various cellular features, like cell movement and parameters for cell morphology 24 .…”

Section: Methodsmentioning

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: Methodsmentioning

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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show abstract

“…Since the cell numbers in each cell culture differed at time-point 0, the cell numbers at time-point t were normalized to those of the control sample (NG group) at time-point 0 as follows: where N t,g is the total number of imaged cells at time-point t , g represents different groups, and cell confluence was calculated and normalized similarly (Janicke et al 2017 ).…”

Section: Methodsmentioning

confidence: 99%

“…where N t,g is the total number of imaged cells at time-point t, g represents different groups, and cell confluence was calculated and normalized similarly (Janicke et al 2017).…”

Section: Statistical Analysis Statistical Analysis Was Performed Usingmentioning

confidence: 99%

Upregulation of miRNA-23a-3p rescues high glucose-induced cell apoptosis and proliferation inhibition in cardiomyocytes

Wang

Yang

et al. 2020

In Vitro Cell.Dev.Biol.-Animal

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Maternal hyperglycemia potentially inhibits the development of the fetal heart by suppressing cardiomyocyte proliferation and promoting apoptosis. Different studies have indicated that miRNAs are key regulators of cardiomyocyte proliferation, differentiation, and apoptosis and play a protective role in a variety of cardiovascular diseases. However, the biological function of miRNA-23a in hyperglycemia-related cardiomyocyte injury is not fully understood. The present study investigated the effect of miRNA-23a-3p on cell proliferation and apoptosis in a myocardial injury model induced by high glucose. H9c2 cardiomyocytes were exposed to high glucose to establish an in vitro myocardial injury model and then transfected with miRNA-23a-3p mimics. After miRNA-23a-3p transfection, lens-free microscopy was used to dynamically monitor cell numbers and confluence and calculate the cell cycle duration. CCK-8 and EdU incorporation assays were performed to detect cell proliferation. Flow cytometry was used to measured cell apoptosis. Upregulation of miRNA-23a-3p significantly alleviated high glucose-induced cell apoptosis and cell proliferation inhibition (p < 0.01 and p < 0.0001, respectively). The cell cycle of the miRNA-23a-3p mimics group was significantly shorter than that of the negative control group (p < 0.01). The expression of cell cycle–activating and apoptosis inhibition-associated factors Ccna2, Ccne1, and Bcl-2 was downregulated by high glucose and upregulated by miRNA-23a-3p overexpression in high glucose-injured H9c2 cells. miRNA-23a-3p mimics transfection before high glucose treatment had a significantly greater benefit than transfection after high glucose treatment (p < 0.0001), and the rescue effect of miRNA-23a-3p increased as the concentration increased. This study suggests that miRNA-23a-3p exerted a dose- and time-dependent protective effect on high glucose-induced H9c2 cardiomyocyte injury.

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“…This technique applied to live cells is noninvasive, label-free, and sensitive to minute alterations in cell shape and subcellular features (24,25) and movement (26,27). Parameter maps utilizing reconstructed optical phase contain information related to phase height and pixel intensity variation, which reveals fruitful information related to cell structure and function (28,29). Quantitative cell phase height maps recorded by DHM have been used to monitor and characterize alterations in cell phase height and shape parameters during the cell cycle related to division, apoptosis, and substrate adhesion (30)(31)(32)(33).…”

mentioning

confidence: 99%

Morphology, Motility, and Cytoskeletal Architecture of Breast Cancer Cells Depend on Keratin 19 and Substrate

et al. 2020

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Cancer cells gain motility through events that accompany modulation of cell shape and include altered expression of keratins. However, the role of keratins in change of cancer cell architecture is not well understood. Therefore, we ablated the expression of keratin 19 (K19) in breast cancer cells of the MDA-MB-231 cell line and found that cells lacking K19 become more elongated in culture, with morphological reversion toward the parental phenotype upon transduction of KRT19. Also, the number of actin stress fibers and focal adhesions were significantly reduced in KRT19 knockout (KO) cells. The altered morphology of KRT19 KO cells was then characterized quantitatively using digital holographic microscopy (DHM), which not only confirmed the phenotypic change of KRT19 KO cells but also identified that the K19-dependent morphological change is dependent on the substrate type. A new quantitative method of single cell analysis from DHM, via average phase difference maps, facilitated evaluation of K19-substrate interactive effects on cell morphology. When plated on collagen substrate, KRT19 KO cells were less elongated and resembled parental cells. Assessing single cell motility further showed that while KRT19 KO cells moved faster than parental cells on a rigid surface, this increase in motility became abrogated when cells were plated on collagen. Overall, our study suggests that K19 inhibits cell motility by regulating cell shape in a substrate-dependent manner. Thus, this study provides a potential basis for the altered expression of keratins associated with change in cell shape and motility of cancer cells.

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Label‐free high temporal resolution assessment of cell proliferation using digital holographic microscopy

Cited by 28 publications

References 67 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

Upregulation of miRNA-23a-3p rescues high glucose-induced cell apoptosis and proliferation inhibition in cardiomyocytes

Morphology, Motility, and Cytoskeletal Architecture of Breast Cancer Cells Depend on Keratin 19 and Substrate

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