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Improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with ZFtool
BACKGROUND: Zebrafish (Danio rerio) is a model organism that has emerged as a tool for cancer research, cancer being the second most common cause of death after cardiovascular disease for humans in the developed world. Zebrafish is a useful model for xenotransplantation of human cancer cells and tox...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5748948/ https://www.ncbi.nlm.nih.gov/pubmed/29291719 http://dx.doi.org/10.1186/s12885-017-3919-8 |
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author | Cabezas-Sainz, Pablo Guerra-Varela, Jorge Carreira, María J. Mariscal, Javier Roel, María Rubiolo, Juan A. Sciara, Andrés A. Abal, Miguel Botana, Luis M. López, Rafael Sánchez, Laura |
author_facet | Cabezas-Sainz, Pablo Guerra-Varela, Jorge Carreira, María J. Mariscal, Javier Roel, María Rubiolo, Juan A. Sciara, Andrés A. Abal, Miguel Botana, Luis M. López, Rafael Sánchez, Laura |
author_sort | Cabezas-Sainz, Pablo |
collection | PubMed |
description | BACKGROUND: Zebrafish (Danio rerio) is a model organism that has emerged as a tool for cancer research, cancer being the second most common cause of death after cardiovascular disease for humans in the developed world. Zebrafish is a useful model for xenotransplantation of human cancer cells and toxicity studies of different chemotherapeutic compounds in vivo. Compared to the murine model, the zebrafish model is faster, can be screened using high-throughput methods and has a lower maintenance cost, making it possible and affordable to create personalized therapies. While several methods for cell proliferation determination based on image acquisition and quantification have been developed, some drawbacks still remain. In the xenotransplantation technique, quantification of cellular proliferation in vivo is critical to standardize the process for future preclinical applications of the model. METHODS: This study improved the conditions of the xenotransplantation technique – quantification of cellular proliferation in vivo was performed through image processing with our ZFtool software and optimization of temperature in order to standardize the process for a future preclinical applications. ZFtool was developed to establish a base threshold that eliminates embryo auto-fluorescence and measures the area of marked cells (GFP) and the intensity of those cells to define a ‘proliferation index’. RESULTS: The analysis of tumor cell proliferation at different temperatures (34 °C and 36 °C) in comparison to in vitro cell proliferation provides of a better proliferation rate, achieved as expected at 36°, a maintenance temperature not demonstrated up to now. The mortality of the embryos remained between 5% and 15%. 5- Fluorouracil was tested for 2 days, dissolved in the incubation medium, in order to quantify the reduction of the tumor mass injected. In almost all of the embryos incubated at 36 °C and incubated with 5-Fluorouracil, there was a significant tumor cell reduction compared with the control group. This was not the case at 34 °C. CONCLUSIONS: Our results demonstrate that the proliferation of the injected cells is better at 36 °C and that this temperature is the most suitable for testing chemotherapeutic drugs like the 5-Fluorouracil. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12885-017-3919-8) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-5748948 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-57489482018-01-05 Improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with ZFtool Cabezas-Sainz, Pablo Guerra-Varela, Jorge Carreira, María J. Mariscal, Javier Roel, María Rubiolo, Juan A. Sciara, Andrés A. Abal, Miguel Botana, Luis M. López, Rafael Sánchez, Laura BMC Cancer Research Article BACKGROUND: Zebrafish (Danio rerio) is a model organism that has emerged as a tool for cancer research, cancer being the second most common cause of death after cardiovascular disease for humans in the developed world. Zebrafish is a useful model for xenotransplantation of human cancer cells and toxicity studies of different chemotherapeutic compounds in vivo. Compared to the murine model, the zebrafish model is faster, can be screened using high-throughput methods and has a lower maintenance cost, making it possible and affordable to create personalized therapies. While several methods for cell proliferation determination based on image acquisition and quantification have been developed, some drawbacks still remain. In the xenotransplantation technique, quantification of cellular proliferation in vivo is critical to standardize the process for future preclinical applications of the model. METHODS: This study improved the conditions of the xenotransplantation technique – quantification of cellular proliferation in vivo was performed through image processing with our ZFtool software and optimization of temperature in order to standardize the process for a future preclinical applications. ZFtool was developed to establish a base threshold that eliminates embryo auto-fluorescence and measures the area of marked cells (GFP) and the intensity of those cells to define a ‘proliferation index’. RESULTS: The analysis of tumor cell proliferation at different temperatures (34 °C and 36 °C) in comparison to in vitro cell proliferation provides of a better proliferation rate, achieved as expected at 36°, a maintenance temperature not demonstrated up to now. The mortality of the embryos remained between 5% and 15%. 5- Fluorouracil was tested for 2 days, dissolved in the incubation medium, in order to quantify the reduction of the tumor mass injected. In almost all of the embryos incubated at 36 °C and incubated with 5-Fluorouracil, there was a significant tumor cell reduction compared with the control group. This was not the case at 34 °C. CONCLUSIONS: Our results demonstrate that the proliferation of the injected cells is better at 36 °C and that this temperature is the most suitable for testing chemotherapeutic drugs like the 5-Fluorouracil. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12885-017-3919-8) contains supplementary material, which is available to authorized users. BioMed Central 2018-01-02 /pmc/articles/PMC5748948/ /pubmed/29291719 http://dx.doi.org/10.1186/s12885-017-3919-8 Text en © The Author(s). 2017 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Article Cabezas-Sainz, Pablo Guerra-Varela, Jorge Carreira, María J. Mariscal, Javier Roel, María Rubiolo, Juan A. Sciara, Andrés A. Abal, Miguel Botana, Luis M. López, Rafael Sánchez, Laura Improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with ZFtool |
title | Improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with ZFtool |
title_full | Improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with ZFtool |
title_fullStr | Improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with ZFtool |
title_full_unstemmed | Improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with ZFtool |
title_short | Improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with ZFtool |
title_sort | improving zebrafish embryo xenotransplantation conditions by increasing incubation temperature and establishing a proliferation index with zftool |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5748948/ https://www.ncbi.nlm.nih.gov/pubmed/29291719 http://dx.doi.org/10.1186/s12885-017-3919-8 |
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