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Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-xenotransplantation

BACKGROUND: Ovarian tissue cryopreservation has a wide range of cancerous indications. Avoiding relapse becomes a specific concern that clinicians frequently encounter. The data about the comparative viability of cancer cells after cryopreservation are limited. This study aimed to evaluate the effec...

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Autores principales: Du, Xinxin, Todorov, Plamen, Isachenko, Evgenia, Rahimi, Gohar, Mallmann, Peter, Meng, Yuanguang, Isachenko, Vladimir
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7425039/
https://www.ncbi.nlm.nih.gov/pubmed/32787800
http://dx.doi.org/10.1186/s12885-020-07227-z
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author Du, Xinxin
Todorov, Plamen
Isachenko, Evgenia
Rahimi, Gohar
Mallmann, Peter
Meng, Yuanguang
Isachenko, Vladimir
author_facet Du, Xinxin
Todorov, Plamen
Isachenko, Evgenia
Rahimi, Gohar
Mallmann, Peter
Meng, Yuanguang
Isachenko, Vladimir
author_sort Du, Xinxin
collection PubMed
description BACKGROUND: Ovarian tissue cryopreservation has a wide range of cancerous indications. Avoiding relapse becomes a specific concern that clinicians frequently encounter. The data about the comparative viability of cancer cells after cryopreservation are limited. This study aimed to evaluate the effect of cryopreservation on breast cancer cells. METHODS: We used in-vitro cultured ZR-75-1 and MDA-MB-231 cell lines. Cell samples of each lineage were distributed into the non-intervened and cryopreserved groups. The cryopreservation procedures comprised programmed slow freezing followed by thawing at 100 °C, 60 s. Biological phenotypes and the related protein markers were compared between the two groups. The EVOS FL Auto 2 Cell Image System was used to monitor cell morphology. Cell proliferation, motility, and penetration were characterized by CCK-8, wound-healing, and transmembrane assay, respectively. The expression of Ki-67, P53, GATA3, E-cadherin, Vimentin, and F-Actin was captured by immunofluorescent staining and western blotting as the proxy measurements of the related properties. The chorioallantoic membrane (CAM) xenotransplantation was conducted to explore angiogenesis induced by cancer cells. RESULTS: After 5 days in vitro culture, the cell concentration of cryopreserved and non-intervened groups was 15.7 × 10(4) vs. 14.4 × 10(4)cells/ml, (ZR-75-1, p > 0.05), and 25.1 × 10(4) vs. 26.6 × 10(4) cells/ml (MDA-MB-231, p > 0.05). Some cryopreserved ZR-75-1 cells presented spindle shape with filopodia and lamellipodia and dissociated from the cell cluster after cryopreservation. Both cell lines demonstrated increased cell migrating capability and invasion after cryopreservation. The expression of Ki-67 and P53 did not differ between the cryopreserved and non-intervened groups. E-cadherin and GATA3 expression downregulated in the cryopreserved ZR-75-1 cells. Vimentin and F-actin exhibited an upregulated level in cryopreserved ZR-75-1 and MDA-MB-231 cells. The cryopreserved MDA-MB-231 cells induced significant angiogenesis around the grafts on CAM with the vascular density 0.313 ± 0.03 and 0.342 ± 0.04, compared with that of non-intervened cells of 0.238 ± 0.05 and 0.244 ± 0.03, p < 0.0001. CONCLUSIONS: Cryopreservation promotes breast cancer cells in terms of epithelial-mesenchymal transition and angiogenesis induction, thus increasing metastasis risk.
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spelling pubmed-74250392020-08-16 Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-xenotransplantation Du, Xinxin Todorov, Plamen Isachenko, Evgenia Rahimi, Gohar Mallmann, Peter Meng, Yuanguang Isachenko, Vladimir BMC Cancer Research Article BACKGROUND: Ovarian tissue cryopreservation has a wide range of cancerous indications. Avoiding relapse becomes a specific concern that clinicians frequently encounter. The data about the comparative viability of cancer cells after cryopreservation are limited. This study aimed to evaluate the effect of cryopreservation on breast cancer cells. METHODS: We used in-vitro cultured ZR-75-1 and MDA-MB-231 cell lines. Cell samples of each lineage were distributed into the non-intervened and cryopreserved groups. The cryopreservation procedures comprised programmed slow freezing followed by thawing at 100 °C, 60 s. Biological phenotypes and the related protein markers were compared between the two groups. The EVOS FL Auto 2 Cell Image System was used to monitor cell morphology. Cell proliferation, motility, and penetration were characterized by CCK-8, wound-healing, and transmembrane assay, respectively. The expression of Ki-67, P53, GATA3, E-cadherin, Vimentin, and F-Actin was captured by immunofluorescent staining and western blotting as the proxy measurements of the related properties. The chorioallantoic membrane (CAM) xenotransplantation was conducted to explore angiogenesis induced by cancer cells. RESULTS: After 5 days in vitro culture, the cell concentration of cryopreserved and non-intervened groups was 15.7 × 10(4) vs. 14.4 × 10(4)cells/ml, (ZR-75-1, p > 0.05), and 25.1 × 10(4) vs. 26.6 × 10(4) cells/ml (MDA-MB-231, p > 0.05). Some cryopreserved ZR-75-1 cells presented spindle shape with filopodia and lamellipodia and dissociated from the cell cluster after cryopreservation. Both cell lines demonstrated increased cell migrating capability and invasion after cryopreservation. The expression of Ki-67 and P53 did not differ between the cryopreserved and non-intervened groups. E-cadherin and GATA3 expression downregulated in the cryopreserved ZR-75-1 cells. Vimentin and F-actin exhibited an upregulated level in cryopreserved ZR-75-1 and MDA-MB-231 cells. The cryopreserved MDA-MB-231 cells induced significant angiogenesis around the grafts on CAM with the vascular density 0.313 ± 0.03 and 0.342 ± 0.04, compared with that of non-intervened cells of 0.238 ± 0.05 and 0.244 ± 0.03, p < 0.0001. CONCLUSIONS: Cryopreservation promotes breast cancer cells in terms of epithelial-mesenchymal transition and angiogenesis induction, thus increasing metastasis risk. BioMed Central 2020-08-12 /pmc/articles/PMC7425039/ /pubmed/32787800 http://dx.doi.org/10.1186/s12885-020-07227-z Text en © The Author(s) 2020 Open AccessThis 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/. 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 in a credit line to the data.
spellingShingle Research Article
Du, Xinxin
Todorov, Plamen
Isachenko, Evgenia
Rahimi, Gohar
Mallmann, Peter
Meng, Yuanguang
Isachenko, Vladimir
Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-xenotransplantation
title Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-xenotransplantation
title_full Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-xenotransplantation
title_fullStr Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-xenotransplantation
title_full_unstemmed Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-xenotransplantation
title_short Increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after CAM-xenotransplantation
title_sort increasing of malignancy of breast cancer cells after cryopreservation: molecular detection and activation of angiogenesis after cam-xenotransplantation
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7425039/
https://www.ncbi.nlm.nih.gov/pubmed/32787800
http://dx.doi.org/10.1186/s12885-020-07227-z
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