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A Machine Learning-Based Image Segmentation Method to Quantify In Vitro Osteoclast Culture Endpoints
Quantification of in vitro osteoclast cultures (e.g. cell number) often relies on manual counting methods. These approaches are labour intensive, time consuming and result in substantial inter- and intra-user variability. This study aimed to develop and validate an automated workflow to robustly qua...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Springer US
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10516805/ https://www.ncbi.nlm.nih.gov/pubmed/37566229 http://dx.doi.org/10.1007/s00223-023-01121-z |
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author | Davies, Bethan K. Hibbert, Andrew P. Roberts, Scott J. Roberts, Helen C. Tickner, Jennifer C. Holdsworth, Gill Arnett, Timothy R. Orriss, Isabel R. |
author_facet | Davies, Bethan K. Hibbert, Andrew P. Roberts, Scott J. Roberts, Helen C. Tickner, Jennifer C. Holdsworth, Gill Arnett, Timothy R. Orriss, Isabel R. |
author_sort | Davies, Bethan K. |
collection | PubMed |
description | Quantification of in vitro osteoclast cultures (e.g. cell number) often relies on manual counting methods. These approaches are labour intensive, time consuming and result in substantial inter- and intra-user variability. This study aimed to develop and validate an automated workflow to robustly quantify in vitro osteoclast cultures. Using ilastik, a machine learning-based image analysis software, images of tartrate resistant acid phosphatase-stained mouse osteoclasts cultured on dentine discs were used to train the ilastik-based algorithm. Assessment of algorithm training showed that osteoclast numbers strongly correlated between manual- and automatically quantified values (r = 0.87). Osteoclasts were consistently faithfully segmented by the model when visually compared to the original reflective light images. The ability of this method to detect changes in osteoclast number in response to different treatments was validated using zoledronate, ticagrelor, and co-culture with MCF7 breast cancer cells. Manual and automated counting methods detected a 70% reduction (p < 0.05) in osteoclast number, when cultured with 10 nM zoledronate and a dose-dependent decrease with 1–10 μM ticagrelor (p < 0.05). Co-culture with MCF7 cells increased osteoclast number by ≥ 50% irrespective of quantification method. Overall, an automated image segmentation and analysis workflow, which consistently and sensitively identified in vitro osteoclasts, was developed. Advantages of this workflow are (1) significantly reduction in user variability of endpoint measurements (93%) and analysis time (80%); (2) detection of osteoclasts cultured on different substrates from different species; and (3) easy to use and freely available to use along with tutorial resources. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00223-023-01121-z. |
format | Online Article Text |
id | pubmed-10516805 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Springer US |
record_format | MEDLINE/PubMed |
spelling | pubmed-105168052023-09-24 A Machine Learning-Based Image Segmentation Method to Quantify In Vitro Osteoclast Culture Endpoints Davies, Bethan K. Hibbert, Andrew P. Roberts, Scott J. Roberts, Helen C. Tickner, Jennifer C. Holdsworth, Gill Arnett, Timothy R. Orriss, Isabel R. Calcif Tissue Int Original Research Quantification of in vitro osteoclast cultures (e.g. cell number) often relies on manual counting methods. These approaches are labour intensive, time consuming and result in substantial inter- and intra-user variability. This study aimed to develop and validate an automated workflow to robustly quantify in vitro osteoclast cultures. Using ilastik, a machine learning-based image analysis software, images of tartrate resistant acid phosphatase-stained mouse osteoclasts cultured on dentine discs were used to train the ilastik-based algorithm. Assessment of algorithm training showed that osteoclast numbers strongly correlated between manual- and automatically quantified values (r = 0.87). Osteoclasts were consistently faithfully segmented by the model when visually compared to the original reflective light images. The ability of this method to detect changes in osteoclast number in response to different treatments was validated using zoledronate, ticagrelor, and co-culture with MCF7 breast cancer cells. Manual and automated counting methods detected a 70% reduction (p < 0.05) in osteoclast number, when cultured with 10 nM zoledronate and a dose-dependent decrease with 1–10 μM ticagrelor (p < 0.05). Co-culture with MCF7 cells increased osteoclast number by ≥ 50% irrespective of quantification method. Overall, an automated image segmentation and analysis workflow, which consistently and sensitively identified in vitro osteoclasts, was developed. Advantages of this workflow are (1) significantly reduction in user variability of endpoint measurements (93%) and analysis time (80%); (2) detection of osteoclasts cultured on different substrates from different species; and (3) easy to use and freely available to use along with tutorial resources. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00223-023-01121-z. Springer US 2023-08-11 2023 /pmc/articles/PMC10516805/ /pubmed/37566229 http://dx.doi.org/10.1007/s00223-023-01121-z Text en © The Author(s) 2023 https://creativecommons.org/licenses/by/4.0/Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . |
spellingShingle | Original Research Davies, Bethan K. Hibbert, Andrew P. Roberts, Scott J. Roberts, Helen C. Tickner, Jennifer C. Holdsworth, Gill Arnett, Timothy R. Orriss, Isabel R. A Machine Learning-Based Image Segmentation Method to Quantify In Vitro Osteoclast Culture Endpoints |
title | A Machine Learning-Based Image Segmentation Method to Quantify In Vitro Osteoclast Culture Endpoints |
title_full | A Machine Learning-Based Image Segmentation Method to Quantify In Vitro Osteoclast Culture Endpoints |
title_fullStr | A Machine Learning-Based Image Segmentation Method to Quantify In Vitro Osteoclast Culture Endpoints |
title_full_unstemmed | A Machine Learning-Based Image Segmentation Method to Quantify In Vitro Osteoclast Culture Endpoints |
title_short | A Machine Learning-Based Image Segmentation Method to Quantify In Vitro Osteoclast Culture Endpoints |
title_sort | machine learning-based image segmentation method to quantify in vitro osteoclast culture endpoints |
topic | Original Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10516805/ https://www.ncbi.nlm.nih.gov/pubmed/37566229 http://dx.doi.org/10.1007/s00223-023-01121-z |
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