Heterogeneous cell types in single-cell-derived clones of MCF7 and MDA-MB-231 cells

Gul Nabi Khan, Eun Jin Kim, Tae Seop Shin, Sang Ho Lee

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Background/Aim: Variations in cell phenotype in a single-cell-derived clone may result from asymmetric cell divisions that lead to different cell fate in a homogenous microenvironment. The aim of this study was to demonstrate the extent of cell variety in single-cell-derived clones and propose a different strategy in treating cancer by observed phenotypic heterogeneity in cellular types. Additionally, the role of metabolic enzyme and housekeeping gene, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), in cellular phenotype was evaluated in two breast cancer cell lines. Materials and Methods: Two GAPDH-overexpressing breast cancer cell lines, MCF7-GAPDH-RFP (MCF7-RFP) and MDA-MB-231-GAPDH-RFP (MDA-RFP), were established. Microscopic recordings were made at 12-h intervals in single-cell-derived clones of both cell lines up to 8-10 days. Crystal violet and Hoechst 33342 (Hoechst), as well as specific cell-type (epithelial and mesenchymal) antibodies, were used for cytochemical and immunohistochemical staining analyses, respectively, at 3, 6 and 9 days during cell growth. Results: Three types of clones with distinct morphologies were identified as holo-, mero- and paraclones. The rates of colony survival during cell growth were 8.3 and 41.7% in MCF7-RFP and MDA-RFP, respectively. Although no significant difference was found in the colony forming efficiency (CFE) of both MCF7 and MDAMD-231 wild-type cells, a markedly significant difference was seen in the CFE of MCF7-RFP and MDA-RFP cells (p=0.001). Wild-type cell-derived holoclones of both cell lines showed drug resistance to doxorubicin (Dox). However, levels of proliferating cell nuclear antigen (PCNA) and vimentin (VIM) marginally decreased in Dox-treated clones. Furthermore, high level of intraclonal heterogeneity was found for CD44, CD140a, VIM, fibronectin (FN), focal adhesion kinase (FAK), paxillin (PXN) and vinculin (VCL) in MCF7 and MDA clones during different stages of clonal development. Expressions of CD140a, FN, VIM and FAK were induced in GAPDH-red fluorescent protein (RFP)-tagged clones of both cell lines. Conclusion: The GAPDH-RFP recombinant protein played an important role in morphological heterogeneity detection in early stages of clonal development. Moreover, phenotypic heterogeneity in clones, caused by the cells expressing specific antigens, such as CD44, CD140a, FN, VIM, FAK and VCL, can be the right target for therapeutic drugs.

Original languageEnglish
Pages (from-to)2343-2354
Number of pages12
JournalAnticancer Research
Volume37
Issue number5
DOIs
Publication statusPublished - 2017 May 1

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Glyceraldehyde-3-Phosphate Dehydrogenases
Clone Cells
Cells
Vimentin
Focal Adhesion Protein-Tyrosine Kinases
Fibronectins
Cell Line
Vinculin
Cell growth
Doxorubicin
Paxillin
Gentian Violet
Asymmetric Cell Division
Breast Neoplasms
Phenotype
Proliferating Cell Nuclear Antigen
Recombinant Proteins
Essential Genes
Pharmaceutical Preparations
Growth

Keywords

  • Clonal analysis
  • GAPDH
  • Heterogeneity
  • MCF7
  • MDA-MB-231
  • Surface markers

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

Heterogeneous cell types in single-cell-derived clones of MCF7 and MDA-MB-231 cells. / Khan, Gul Nabi; Kim, Eun Jin; Shin, Tae Seop; Lee, Sang Ho.

In: Anticancer Research, Vol. 37, No. 5, 01.05.2017, p. 2343-2354.

Research output: Contribution to journalArticle

Khan, Gul Nabi ; Kim, Eun Jin ; Shin, Tae Seop ; Lee, Sang Ho. / Heterogeneous cell types in single-cell-derived clones of MCF7 and MDA-MB-231 cells. In: Anticancer Research. 2017 ; Vol. 37, No. 5. pp. 2343-2354.
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N2 - Background/Aim: Variations in cell phenotype in a single-cell-derived clone may result from asymmetric cell divisions that lead to different cell fate in a homogenous microenvironment. The aim of this study was to demonstrate the extent of cell variety in single-cell-derived clones and propose a different strategy in treating cancer by observed phenotypic heterogeneity in cellular types. Additionally, the role of metabolic enzyme and housekeeping gene, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), in cellular phenotype was evaluated in two breast cancer cell lines. Materials and Methods: Two GAPDH-overexpressing breast cancer cell lines, MCF7-GAPDH-RFP (MCF7-RFP) and MDA-MB-231-GAPDH-RFP (MDA-RFP), were established. Microscopic recordings were made at 12-h intervals in single-cell-derived clones of both cell lines up to 8-10 days. Crystal violet and Hoechst 33342 (Hoechst), as well as specific cell-type (epithelial and mesenchymal) antibodies, were used for cytochemical and immunohistochemical staining analyses, respectively, at 3, 6 and 9 days during cell growth. Results: Three types of clones with distinct morphologies were identified as holo-, mero- and paraclones. The rates of colony survival during cell growth were 8.3 and 41.7% in MCF7-RFP and MDA-RFP, respectively. Although no significant difference was found in the colony forming efficiency (CFE) of both MCF7 and MDAMD-231 wild-type cells, a markedly significant difference was seen in the CFE of MCF7-RFP and MDA-RFP cells (p=0.001). Wild-type cell-derived holoclones of both cell lines showed drug resistance to doxorubicin (Dox). However, levels of proliferating cell nuclear antigen (PCNA) and vimentin (VIM) marginally decreased in Dox-treated clones. Furthermore, high level of intraclonal heterogeneity was found for CD44, CD140a, VIM, fibronectin (FN), focal adhesion kinase (FAK), paxillin (PXN) and vinculin (VCL) in MCF7 and MDA clones during different stages of clonal development. Expressions of CD140a, FN, VIM and FAK were induced in GAPDH-red fluorescent protein (RFP)-tagged clones of both cell lines. Conclusion: The GAPDH-RFP recombinant protein played an important role in morphological heterogeneity detection in early stages of clonal development. Moreover, phenotypic heterogeneity in clones, caused by the cells expressing specific antigens, such as CD44, CD140a, FN, VIM, FAK and VCL, can be the right target for therapeutic drugs.

AB - Background/Aim: Variations in cell phenotype in a single-cell-derived clone may result from asymmetric cell divisions that lead to different cell fate in a homogenous microenvironment. The aim of this study was to demonstrate the extent of cell variety in single-cell-derived clones and propose a different strategy in treating cancer by observed phenotypic heterogeneity in cellular types. Additionally, the role of metabolic enzyme and housekeeping gene, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), in cellular phenotype was evaluated in two breast cancer cell lines. Materials and Methods: Two GAPDH-overexpressing breast cancer cell lines, MCF7-GAPDH-RFP (MCF7-RFP) and MDA-MB-231-GAPDH-RFP (MDA-RFP), were established. Microscopic recordings were made at 12-h intervals in single-cell-derived clones of both cell lines up to 8-10 days. Crystal violet and Hoechst 33342 (Hoechst), as well as specific cell-type (epithelial and mesenchymal) antibodies, were used for cytochemical and immunohistochemical staining analyses, respectively, at 3, 6 and 9 days during cell growth. Results: Three types of clones with distinct morphologies were identified as holo-, mero- and paraclones. The rates of colony survival during cell growth were 8.3 and 41.7% in MCF7-RFP and MDA-RFP, respectively. Although no significant difference was found in the colony forming efficiency (CFE) of both MCF7 and MDAMD-231 wild-type cells, a markedly significant difference was seen in the CFE of MCF7-RFP and MDA-RFP cells (p=0.001). Wild-type cell-derived holoclones of both cell lines showed drug resistance to doxorubicin (Dox). However, levels of proliferating cell nuclear antigen (PCNA) and vimentin (VIM) marginally decreased in Dox-treated clones. Furthermore, high level of intraclonal heterogeneity was found for CD44, CD140a, VIM, fibronectin (FN), focal adhesion kinase (FAK), paxillin (PXN) and vinculin (VCL) in MCF7 and MDA clones during different stages of clonal development. Expressions of CD140a, FN, VIM and FAK were induced in GAPDH-red fluorescent protein (RFP)-tagged clones of both cell lines. Conclusion: The GAPDH-RFP recombinant protein played an important role in morphological heterogeneity detection in early stages of clonal development. Moreover, phenotypic heterogeneity in clones, caused by the cells expressing specific antigens, such as CD44, CD140a, FN, VIM, FAK and VCL, can be the right target for therapeutic drugs.

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KW - Heterogeneity

KW - MCF7

KW - MDA-MB-231

KW - Surface markers

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