Characterization of side population cells in human malignant mesothelioma cell lines

Our plain-English summary of Characterization of side population cells in human malignant mesothelioma cell lines, by K. Kai, S. D'Costa, B.I. Yoon, et al (Lung Cancer, 2010).

The study

• Authors: K. Kai, S. D'Costa, B.I. Yoon, A.R. Brody, R.C. Sills, Y. Kim
• Journal: Lung Cancer
• Published: 2010;70(2):146-51
• Cited by: 30 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Side population (SP) assay composed of Hoechst 33342 staining and subsequent flow cytometric analysis has been widely utilized for characterizing putative cancer stem cells (CSCs) in various human malignancies. The present study was designed to evaluate the SP assay as a research tool for mesothelial CSCs. A distinct fraction of SP cells was identified in various human malignant mesothelioma (HMM) cell lines, ranging from 0.05 to 1.32%. The sorted mesothelial SP cells exhibited enhanced proliferation potentials and higher expression of stem-cell genes, compared to non-SP (NSP) cells. Cisplatin treatment increased percentage of SP cells in the HMM cell lines. However, tumorigenic potential of SP cells in immunodeficient mice was similar to that of the NSP cells. These data indicated that SP assay may not be appropriate for enriching putative CSCs in HMM cell lines, and thus warrants the development of a novel tool for mesothelial CSC study.

Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

In plain English

This laboratory study used cells grown in dishes and injected into mice. It did not study people or asbestos exposure. The authors tested whether a 'side population' test, using a dye and a cell-sorting machine, could pick out possible cancer stem cells in human malignant mesothelioma cell lines. They examined three mesothelioma cell lines (NCI-H513, MS-1 and NCI-H2373) and one non-malignant mesothelial cell line (MeT-5A). All four contained side population cells, ranging from 0.27% (NCI-H2373) to 1.32% (NCI-H513). Sorted MS-1 cells were plated at 1,000 cells per well and grown for five days. Side population cells formed 215 colonies in total, compared with 63 from the other cells. In both NCI-H513 and MS-1, side population cells had higher mRNA levels of the stem-cell genes BMI1, OCT4 and NOTCH1. The cancer drug cisplatin raised the side population share in NCI-H513 from 0.71 to 8.37%. However, in immunodeficient mice, side population and other MS-1 cells formed tumors at rates that were not statistically different. The authors concluded that possible cancer stem cells were present in both groups and that their study underlines the limitations of the test.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) study of human mesothelioma cell lines, with a tumor-formation experiment in immunodeficient mice
• Who was studied: Three human malignant mesothelioma cell lines (NCI-H513, described as epithelioid; MS-1, biphasic; NCI-H2373, sarcomatoid) and one non-malignant SV40-transformed mesothelial cell line (MeT-5A). Cells from MS-1 were also injected under the skin of non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice.
• Exposure: No asbestos exposure was studied; asbestos appears only as background in the introduction. The cell lines were treated with the anticancer drug cisplatin and with LY294002, an inhibitor of PI3K signaling, and were compared with untreated or vehicle-treated cells.
• Main results:
  • All mesothelial cell lines contained side population cells. Mean side population fractions ranged from 0.27% (NCI-H2373) to 1.32% (NCI-H513) of gated cells; MS-1 was 0.88% and MeT-5A 0.36% in Table 1.
  • Cisplatin increased the side population fraction: in NCI-H513 (48 hours of treatment) from 0.71 to 8.37%.
  • In MS-1 cells, 20 hours of cisplatin treatment increased the side population fraction from 0.27 to 0.51%.
  • In both NCI-H513 and MS-1, side population cells had higher mRNA levels of the stem-cell genes BMI1, OCT4 and NOTCH1 than non-side population cells, measured by real-time RT-PCR.
  • Staining for the ABCG2 protein was markedly more frequent and more intense in side population cells than in non-side population cells.
  • In the colony test, sorted MS-1 side population and non-side population cells were plated at 1,000 cells per well and grown for 5 days. Side population cells produced 215 colonies in total, significantly more than the 63 from non-side population cells. Large colonies came only from side population cells.
  • In NOD/SCID mice injected with MS-1 cells, the frequency of tumor-bearing mice was not statistically different between unsorted, side population and non-side population cells. Tumors per injection (Table 2) were: unsorted 3/4 at each dose; side population 7/7 at 10^3 cells and 3/4 at 10^2 cells (not done at 5 × 10^4); non-side population 4/4 at 5 × 10^4, 4/4 at 10^3 and 3/4 at 10^2 cells.
  • The mouse tumors looked like biphasic mesothelioma under the microscope. Staining of 12 tumors (6 from side population cells, 6 from non-side population cells) showed no difference in pattern; tumor cells stained for both cytokeratin and vimentin.
  • The authors stated that possible cancer stem cells were present in both the side population and non-side population fractions of the MS-1 line.
• The authors' conclusion: The authors concluded that the side population test may not be appropriate for enriching possible cancer stem cells in mesothelioma cell lines. They said their study underlines the limitations of this widely used test and called for further studies using other mesothelioma cell lines and clinical samples.
• Limitations the authors state:
  • The authors suggested that the unexpectedly high tumor formation might be explained by differences in the strain of NOD/SCID mice.
  • The MS-1 line has been grown in the laboratory for decades, which the authors offered as another possible reason why most of its cells may form tumors.
  • The authors said further studies using cancer cells taken directly from patients' tumors may be necessary.
  • The authors noted that their explanation for the cisplatin effect (side population cells being more resistant to cisplatin) is not proven.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Mesothelioma.

Medical terms in this study, explained in our Lexicon: Malignant Mesothelioma, STEM, Cisplatin.

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