An Evaluation of Matrix-Containing and Humanised Matrix-Free 3-Dimensional Cell Culture Systems for Studying Breast Cancer

Grace C Roberts, Paul G Morris, Marcus A Moss, Sarah L Maltby, Chelsea A Palmer, Claire E Nash, Emily Smart, Deborah L Holliday, Valerie Speirs

Research output: Contribution to journalArticle

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3 Downloads (Pure)

Abstract

BACKGROUND: 3D cell cultures are emerging as more physiologically meaningful alternatives to monolayer cultures for many biological applications. They are attractive because they more closely mimic in vivo morphology, especially when co-cultured with stromal fibroblasts.

METHODOLOGY/PRINCIPAL FINDINGS: We compared the efficacy of 3 different 3D cell culture systems; collagen I, low attachment culture vessels and a modification of Fibrolife®, a specialised humanised cell culture medium devoid of animal-derived components, using breast cancer cell lines representative of the different molecular subtypes of breast cancer, cultured alone or with human mammary fibroblasts with a view to developing matrix-free humanised systems. 3D collagen I culture supported the growth of a range of breast cancer cell lines. By modifying the composition of Fibrolife® to epiFL, matrix-free cell culture was possible. During sequential transfer to epiFL breast cancer cells gradually detached from the flask, growing progressively as spheroids. Phenotype was stable and reversible with cells remaining actively proliferating and easily accessible throughout culture. They could also be revived from frozen stocks. To achieve co-culture with fibroblasts in epiFL required use of low attachment culture vessels instead of standard plastic as fibroblasts remained adherent in epiFL. Here, cancer cell spheroids were allowed to form before adding fibroblasts. Immunohistochemical examination showed fibroblasts scattered throughout the epithelial spheroid, not dissimilar to the relationship of tumour stroma in human breast cancer.

CONCLUSIONS: Because of its ease of handling, matrix-free 3D cell culture may be a useful model to study the influence of fibroblasts on breast cancer epithelial cells with use of epiFL culture medium taking this a step further towards a fully humanised 3D model. This methodology could be applied to other types of cancer cell lines, making this a versatile technique for cancer researchers wishing to use in vitro systems that better reflect cancer in vivo.

Original languageEnglish
Article numbere0157004
JournalPloS ONE
Volume11
Issue number6
DOIs
Publication statusPublished - 14 Jun 2016

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Cell culture
breast neoplasms
Fibroblasts
fibroblasts
cell culture
Cell Culture Techniques
Breast Neoplasms
Cells
cell lines
Neoplasms
Cell Line
neoplasms
Culture Media
collagen
Collagen
culture media
coculture
Coculture Techniques
Plastics
breasts

Keywords

  • Biocompatible Materials
  • Breast
  • Breast Neoplasms
  • Cell Adhesion
  • Cell Line, Tumor
  • Cell Survival
  • Coculture Techniques
  • Collagen Type I
  • Female
  • Fibroblasts
  • Humans
  • Spheroids, Cellular
  • Tumor Cells, Cultured
  • Journal Article

Cite this

An Evaluation of Matrix-Containing and Humanised Matrix-Free 3-Dimensional Cell Culture Systems for Studying Breast Cancer. / Roberts, Grace C; Morris, Paul G; Moss, Marcus A; Maltby, Sarah L; Palmer, Chelsea A; Nash, Claire E; Smart, Emily; Holliday, Deborah L; Speirs, Valerie.

In: PloS ONE, Vol. 11, No. 6, e0157004, 14.06.2016.

Research output: Contribution to journalArticle

Roberts, GC, Morris, PG, Moss, MA, Maltby, SL, Palmer, CA, Nash, CE, Smart, E, Holliday, DL & Speirs, V 2016, 'An Evaluation of Matrix-Containing and Humanised Matrix-Free 3-Dimensional Cell Culture Systems for Studying Breast Cancer', PloS ONE, vol. 11, no. 6, e0157004. https://doi.org/10.1371/journal.pone.0157004
Roberts, Grace C ; Morris, Paul G ; Moss, Marcus A ; Maltby, Sarah L ; Palmer, Chelsea A ; Nash, Claire E ; Smart, Emily ; Holliday, Deborah L ; Speirs, Valerie. / An Evaluation of Matrix-Containing and Humanised Matrix-Free 3-Dimensional Cell Culture Systems for Studying Breast Cancer. In: PloS ONE. 2016 ; Vol. 11, No. 6.
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abstract = "BACKGROUND: 3D cell cultures are emerging as more physiologically meaningful alternatives to monolayer cultures for many biological applications. They are attractive because they more closely mimic in vivo morphology, especially when co-cultured with stromal fibroblasts.METHODOLOGY/PRINCIPAL FINDINGS: We compared the efficacy of 3 different 3D cell culture systems; collagen I, low attachment culture vessels and a modification of Fibrolife{\circledR}, a specialised humanised cell culture medium devoid of animal-derived components, using breast cancer cell lines representative of the different molecular subtypes of breast cancer, cultured alone or with human mammary fibroblasts with a view to developing matrix-free humanised systems. 3D collagen I culture supported the growth of a range of breast cancer cell lines. By modifying the composition of Fibrolife{\circledR} to epiFL, matrix-free cell culture was possible. During sequential transfer to epiFL breast cancer cells gradually detached from the flask, growing progressively as spheroids. Phenotype was stable and reversible with cells remaining actively proliferating and easily accessible throughout culture. They could also be revived from frozen stocks. To achieve co-culture with fibroblasts in epiFL required use of low attachment culture vessels instead of standard plastic as fibroblasts remained adherent in epiFL. Here, cancer cell spheroids were allowed to form before adding fibroblasts. Immunohistochemical examination showed fibroblasts scattered throughout the epithelial spheroid, not dissimilar to the relationship of tumour stroma in human breast cancer.CONCLUSIONS: Because of its ease of handling, matrix-free 3D cell culture may be a useful model to study the influence of fibroblasts on breast cancer epithelial cells with use of epiFL culture medium taking this a step further towards a fully humanised 3D model. This methodology could be applied to other types of cancer cell lines, making this a versatile technique for cancer researchers wishing to use in vitro systems that better reflect cancer in vivo.",
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AU - Roberts, Grace C

AU - Morris, Paul G

AU - Moss, Marcus A

AU - Maltby, Sarah L

AU - Palmer, Chelsea A

AU - Nash, Claire E

AU - Smart, Emily

AU - Holliday, Deborah L

AU - Speirs, Valerie

N1 - Funding: This work was supported by Breast Cancer Now (formerly Breast Cancer Campaign; http://breastcancernow.org/) Grant number 2008NovPr04; Lord Dowding Fund (http://www.ldf.org.uk/) No specific grant number; Pathological Society of Great Britain & Ireland (http://www.pathsoc.org/) Grant number PHD2011/13; CRUK (http://www.cancerresearchuk.org/) Grant number C16708/A18080; Dr Hadwen Trust (GB) (http://www.drhadwentrust.org). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

PY - 2016/6/14

Y1 - 2016/6/14

N2 - BACKGROUND: 3D cell cultures are emerging as more physiologically meaningful alternatives to monolayer cultures for many biological applications. They are attractive because they more closely mimic in vivo morphology, especially when co-cultured with stromal fibroblasts.METHODOLOGY/PRINCIPAL FINDINGS: We compared the efficacy of 3 different 3D cell culture systems; collagen I, low attachment culture vessels and a modification of Fibrolife®, a specialised humanised cell culture medium devoid of animal-derived components, using breast cancer cell lines representative of the different molecular subtypes of breast cancer, cultured alone or with human mammary fibroblasts with a view to developing matrix-free humanised systems. 3D collagen I culture supported the growth of a range of breast cancer cell lines. By modifying the composition of Fibrolife® to epiFL, matrix-free cell culture was possible. During sequential transfer to epiFL breast cancer cells gradually detached from the flask, growing progressively as spheroids. Phenotype was stable and reversible with cells remaining actively proliferating and easily accessible throughout culture. They could also be revived from frozen stocks. To achieve co-culture with fibroblasts in epiFL required use of low attachment culture vessels instead of standard plastic as fibroblasts remained adherent in epiFL. Here, cancer cell spheroids were allowed to form before adding fibroblasts. Immunohistochemical examination showed fibroblasts scattered throughout the epithelial spheroid, not dissimilar to the relationship of tumour stroma in human breast cancer.CONCLUSIONS: Because of its ease of handling, matrix-free 3D cell culture may be a useful model to study the influence of fibroblasts on breast cancer epithelial cells with use of epiFL culture medium taking this a step further towards a fully humanised 3D model. This methodology could be applied to other types of cancer cell lines, making this a versatile technique for cancer researchers wishing to use in vitro systems that better reflect cancer in vivo.

AB - BACKGROUND: 3D cell cultures are emerging as more physiologically meaningful alternatives to monolayer cultures for many biological applications. They are attractive because they more closely mimic in vivo morphology, especially when co-cultured with stromal fibroblasts.METHODOLOGY/PRINCIPAL FINDINGS: We compared the efficacy of 3 different 3D cell culture systems; collagen I, low attachment culture vessels and a modification of Fibrolife®, a specialised humanised cell culture medium devoid of animal-derived components, using breast cancer cell lines representative of the different molecular subtypes of breast cancer, cultured alone or with human mammary fibroblasts with a view to developing matrix-free humanised systems. 3D collagen I culture supported the growth of a range of breast cancer cell lines. By modifying the composition of Fibrolife® to epiFL, matrix-free cell culture was possible. During sequential transfer to epiFL breast cancer cells gradually detached from the flask, growing progressively as spheroids. Phenotype was stable and reversible with cells remaining actively proliferating and easily accessible throughout culture. They could also be revived from frozen stocks. To achieve co-culture with fibroblasts in epiFL required use of low attachment culture vessels instead of standard plastic as fibroblasts remained adherent in epiFL. Here, cancer cell spheroids were allowed to form before adding fibroblasts. Immunohistochemical examination showed fibroblasts scattered throughout the epithelial spheroid, not dissimilar to the relationship of tumour stroma in human breast cancer.CONCLUSIONS: Because of its ease of handling, matrix-free 3D cell culture may be a useful model to study the influence of fibroblasts on breast cancer epithelial cells with use of epiFL culture medium taking this a step further towards a fully humanised 3D model. This methodology could be applied to other types of cancer cell lines, making this a versatile technique for cancer researchers wishing to use in vitro systems that better reflect cancer in vivo.

KW - Biocompatible Materials

KW - Breast

KW - Breast Neoplasms

KW - Cell Adhesion

KW - Cell Line, Tumor

KW - Cell Survival

KW - Coculture Techniques

KW - Collagen Type I

KW - Female

KW - Fibroblasts

KW - Humans

KW - Spheroids, Cellular

KW - Tumor Cells, Cultured

KW - Journal Article

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DO - 10.1371/journal.pone.0157004

M3 - Article

C2 - 27300768

VL - 11

JO - PloS ONE

JF - PloS ONE

SN - 1932-6203

IS - 6

M1 - e0157004

ER -