Abstract
Stem cells have been isolated by their ability to efflux Hoechst 33342 dye and are referred to as the "side population" (SP). In this study, we used flow cytometry and Hoechst 33342 dye efflux assay to isolate and characterize SP cells from six human lung cancer cell lines (H460, H23, HTB-58, A549, H441, and H2170). Nonobese diabetic/severe combined immunodeficiency xenograft experiments showed that SP cells were enriched in tumor-initiating capability compared with non-SP cells. Matrigel invasion assay showed that SP cells also have higher potential for invasiveness. Further characterization of this SP phenotype revealed several stem cell properties. We found evidence for repopulating ability by SP to regenerate a population resembling the original population. SP displayed elevated expression of ABCG2 as well as other ATP-binding cassette transporters and showed resistance to multiple chemotherapeutic drugs. Human telomerase reverse transcriptase expression was higher in the SP, suggesting that this fraction may represent a reservoir with unlimited proliferative potential for generating cancer cells. mRNA levels of minichromosome maintenance (MCM) 7, a member of the MCM family of proteins critical to the DNA replication complex, were lower in SP cells, suggesting that a majority of the SP fraction was in the G(0) quiescent state. Sixteen clinical lung cancer samples also displayed a smaller but persistent SP population. These findings indicate that SP is an enriched source of lung tumor-initiating cells with stem cell properties and may be an important target for effective therapy and a useful tool to investigate the tumorigenic process.
Highlights
Lung cancer is the most common cause of cancer death in the world today [1]
Human tumor cell lines A549, H23, H460, HTB-58, H2170, and H441 were obtained from the American Type Culture Collection (ATCC) and maintained in culture medium recommended by ATCC
We examined the existence of side population’’ (SP) in six human lung cancer cell lines by staining them with Hoechst 33342 dye to generate a Hoechst blue-red profile
Summary
Lung cancer is the most common cause of cancer death in the world today [1]. Lung cancer survival is poor with only 15% of patients surviving 5 years after diagnosis. New chemotherapy agents and radiotherapy have improved survival and quality of life of patients, the overall effect in the last decade has been mainly on palliation rather than reduction in mortality. Evidence is accumulating that solid tumors, such as brain and breast cancer, contain a minor population of ‘‘cancer stem cells’’. Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). M.M. Ho and A.V. Ng contributed to this work and are considered cofirst authors
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