Korean J Gastroenterol.  2011 Mar;57(3):150-157. 10.4166/kjg.2011.57.3.150.

Effects of Chromosomal Polyploidy on Survival of Colon Cancer Cells

• Park SU
• Choi ES
• Jang YS
• Hong SH
• Kim IH
• Chang DK

• KMID: 1128353
• DOI: http://doi.org/10.4166/kjg.2011.57.3.150

Abstract

BACKGROUND/AIMS
Tetraploid cells are frequently observed in the inflamed mucosal epithelial cells of the patients with Barrett's esophagus or chronic ulcerative colitis. Polyploidy often occurs during cell fusion, abortive cell cycle, and endoreplication. Most tetraploid cells are engaged to apoptotic pathway, but some remaining stable tetraploid cells consequently cause aneuploidization and chromosomal instability. We investigated whether tetraploid cells could acquire survival advantage and hold a dominant position for natural selection.
METHODS
We established tetraploid cell line (HCT116GH) from parental diploid colorectal cancer cell line (HCT116) via PEG-mediated cell fusion and compared its cell viability, cell cycle response and apoptotic fractions responded to H2O2 with diploid HCT116 and p53 suppressed HCT116/H6 cell lines.
RESULTS
Using MTT assay, plating efficiency and clonogenicity, we evaluated the survival of each cell line. Tetraploid cell line HCT116GH demonstrated an 83 fold greater resistance to 100 microM H2O2 than the parental diploid HCT116, and 6 fold greater than even the p53 negative diploid HCT116/E6. Cellular sensitivity, G2/M arrests, and apoptotic proportion were observed less in response to H2O2 in HCT116GH compared with HCT116 and HCT116/E6. HCT116GH expressed lower level of p53 and p21 than diploid HCT116.
CONCLUSIONS
Stable tetraploid cell lines showed enhanced viability in comparison to parental diploid cell lines. The enhanced viability observed in tetraploidization surpassed that from downregulation of p53. Frequent appearance of tetraploid cells in stressful condition can be caused by natural selection owing to their enhanced viability and may consequently contribute to cancer cell transformation.

Keyword

Tetraploidy; Chromosomal instability; Aneuploidy; Hydrogen peroxide

MeSH Terms

Apoptosis
Cell Division
Cell Line, Tumor
Cell Survival
Chromosomal Instability
Colonic Neoplasms/*genetics/metabolism
Cyclin-Dependent Kinase Inhibitor p21/metabolism
G2 Phase
Humans
Hydrogen Peroxide/toxicity
Oxidative Stress
*Polyploidy
Tumor Suppressor Protein p53/metabolism

Figure

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  Fig. 1. Establishment of the tetraploid cell line. Tetraploid cell line HCT116GH demonstrated 2N at G1 and 4N at G2 phase when the parental diploid cell line HCT116 demonstated N at G1 and 2N at G2 phase.

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  Fig. 2. Morphology of the tetraploid cells. Tetraploid cell line HCT116GH demonstrated that the size of cells was larger than that of parental diploid HCT116 cells.

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  Fig. 3. Confirmation of stable tetraploid cell line with the reference of the diploid HCT116YFP cell line. In the lower panel depicting the mixture of HCY116YFP and HCT116GH, 2N peak has increased and 4N peak newly appeared, which confirmed that HCT116GH was a stably established tetraploid cell line.

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  Fig. 4. Comparison of survival among tetraploid cell line, patental diploid cell line and p53(−) diploid cell line. MTT assay demonstrated that the tetraploid cell line HCT116GH showed higher survival at each time scale after H2O2 challenging (p＜0.05, ANOVA).

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  Fig. 5. Comparison of the plating efficiency between tetraploid cell line, patental diploid cell line and p53(−) diploid cell line. Plating efficiency demonstrated that tetraploid cell line HCT116GH was resistant to H2 O2 in comparison to the parental diploid HCT116 and even p53 negative diploid cell line HCT116/E6 (p＜0.05, ANOVA).

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  Fig. 6. Comparison of the apoptosis to oxyradical stress among tetraploid cell line, parental diploid cell line and p53(−) diploid cell line. Apoptotic fraction assessed by annexin-V staining was lowest in the tetraploid cell line HCT116GH.

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  Fig. 7. Comparison of the G2/M arrest status to oxyradical stress among tetraploid cell line, patental diploid cell line and p53(−) diploid cell line. The fraction of cells in G2/M arrest was lowest in the tetraploid cell line HCT116GH.

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  Fig. 8. Expression of p53 and p21 to oxyradical stress in tetraploid cell line, patental diploid cell line and p53(−) diploid cell line. Tetraploid cell line HCT116GH expressed lower level of p53 and p21 than diploid HCT116. p53 negative HCT116E6 expressed p21 less abundantly than HCT116, with p53 independent manner.

Reference

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