Quantification of dynamic morphological drug responses in 3D organotypic cell cultures by automated image analysis

Ville Härmä, Hannu-Pekka Schukov, Antti Happonen, Ilmari Ahonen, Johannes Virtanen, Harri Siitari, Malin Åkerfelt, Jyrki Lötjönen, Matthias Nees (Corresponding Author)

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39 Citations (Scopus)

Abstract

Glandular epithelial cells differentiate into complex multicellular or acinar structures, when embedded in three-dimensional (3D) extracellular matrix. The spectrum of different multicellular morphologies formed in 3D is a sensitive indicator for the differentiation potential of normal, non-transformed cells compared to different stages of malignant progression. In addition, single cells or cell aggregates may actively invade the matrix, utilizing epithelial, mesenchymal or mixed modes of motility. Dynamic phenotypic changes involved in 3D tumor cell invasion are sensitive to specific small-molecule inhibitors that target the actin cytoskeleton. We have used a panel of inhibitors to demonstrate the power of automated image analysis as a phenotypic or morphometric readout in cell-based assays. We introduce a streamlined stand-alone software solution that supports large-scale high-content screens, based on complex and organotypic cultures. AMIDA (Automated Morphometric Image Data Analysis) allows quantitative measurements of large numbers of images and structures, with a multitude of different spheroid shapes, sizes, and textures. AMIDA supports an automated workflow, and can be combined with quality control and statistical tools for data interpretation and visualization. We have used a representative panel of 12 prostate and breast cancer lines that display a broad spectrum of different spheroid morphologies and modes of invasion, challenged by a library of 19 direct or indirect modulators of the actin cytoskeleton which induce systematic changes in spheroid morphology and differentiation versus invasion. These results were independently validated by 2D proliferation, apoptosis and cell motility assays. We identified three drugs that primarily attenuated the invasion and formation of invasive processes in 3D, without affecting proliferation or apoptosis. Two of these compounds block Rac signalling, one affects cellular cAMP/cGMP accumulation. Our approach supports the growing needs for user-friendly, straightforward solutions that facilitate large-scale, cell-based 3D assays in basic research, drug discovery, and target validation
Original languageEnglish
Article numbere96426
JournalPLoS ONE
Volume9
Issue number5
DOIs
Publication statusPublished - 2014
MoE publication typeA1 Journal article-refereed

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Cell culture
Image analysis
Assays
cell culture
Cell Culture Techniques
image analysis
drugs
Actins
microfilaments
Pharmaceutical Preparations
Apoptosis
data analysis
assays
apoptosis
cells
Actin Cytoskeleton
Modulators
cell invasion
Quality control
Tumors

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Härmä, V., Schukov, H-P., Happonen, A., Ahonen, I., Virtanen, J., Siitari, H., ... Nees, M. (2014). Quantification of dynamic morphological drug responses in 3D organotypic cell cultures by automated image analysis. PLoS ONE, 9(5), [e96426]. https://doi.org/10.1371/journal.pone.0096426
Härmä, Ville ; Schukov, Hannu-Pekka ; Happonen, Antti ; Ahonen, Ilmari ; Virtanen, Johannes ; Siitari, Harri ; Åkerfelt, Malin ; Lötjönen, Jyrki ; Nees, Matthias. / Quantification of dynamic morphological drug responses in 3D organotypic cell cultures by automated image analysis. In: PLoS ONE. 2014 ; Vol. 9, No. 5.
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Härmä, V, Schukov, H-P, Happonen, A, Ahonen, I, Virtanen, J, Siitari, H, Åkerfelt, M, Lötjönen, J & Nees, M 2014, 'Quantification of dynamic morphological drug responses in 3D organotypic cell cultures by automated image analysis', PLoS ONE, vol. 9, no. 5, e96426. https://doi.org/10.1371/journal.pone.0096426

Quantification of dynamic morphological drug responses in 3D organotypic cell cultures by automated image analysis. / Härmä, Ville; Schukov, Hannu-Pekka; Happonen, Antti; Ahonen, Ilmari; Virtanen, Johannes; Siitari, Harri; Åkerfelt, Malin; Lötjönen, Jyrki; Nees, Matthias (Corresponding Author).

In: PLoS ONE, Vol. 9, No. 5, e96426, 2014.

Research output: Contribution to journalArticleScientificpeer-review

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T1 - Quantification of dynamic morphological drug responses in 3D organotypic cell cultures by automated image analysis

AU - Härmä, Ville

AU - Schukov, Hannu-Pekka

AU - Happonen, Antti

AU - Ahonen, Ilmari

AU - Virtanen, Johannes

AU - Siitari, Harri

AU - Åkerfelt, Malin

AU - Lötjönen, Jyrki

AU - Nees, Matthias

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AB - Glandular epithelial cells differentiate into complex multicellular or acinar structures, when embedded in three-dimensional (3D) extracellular matrix. The spectrum of different multicellular morphologies formed in 3D is a sensitive indicator for the differentiation potential of normal, non-transformed cells compared to different stages of malignant progression. In addition, single cells or cell aggregates may actively invade the matrix, utilizing epithelial, mesenchymal or mixed modes of motility. Dynamic phenotypic changes involved in 3D tumor cell invasion are sensitive to specific small-molecule inhibitors that target the actin cytoskeleton. We have used a panel of inhibitors to demonstrate the power of automated image analysis as a phenotypic or morphometric readout in cell-based assays. We introduce a streamlined stand-alone software solution that supports large-scale high-content screens, based on complex and organotypic cultures. AMIDA (Automated Morphometric Image Data Analysis) allows quantitative measurements of large numbers of images and structures, with a multitude of different spheroid shapes, sizes, and textures. AMIDA supports an automated workflow, and can be combined with quality control and statistical tools for data interpretation and visualization. We have used a representative panel of 12 prostate and breast cancer lines that display a broad spectrum of different spheroid morphologies and modes of invasion, challenged by a library of 19 direct or indirect modulators of the actin cytoskeleton which induce systematic changes in spheroid morphology and differentiation versus invasion. These results were independently validated by 2D proliferation, apoptosis and cell motility assays. We identified three drugs that primarily attenuated the invasion and formation of invasive processes in 3D, without affecting proliferation or apoptosis. Two of these compounds block Rac signalling, one affects cellular cAMP/cGMP accumulation. Our approach supports the growing needs for user-friendly, straightforward solutions that facilitate large-scale, cell-based 3D assays in basic research, drug discovery, and target validation

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