Isolation and characterization of cancer stem-like side population cells in human oral cancer cells

Abstract

Recent studies suggest that cancer stem cells may be responsible for tumorigenesis and contribute to some individuals' resistance to cancer therapy. Some studies demonstrate that side population (SP) cells isolated from diverse cancer cell lines harbor stem cell-like properties; however, there are few reports examining the role of SP cells in human oral cancer. To determine whether human oral cancer cell lines contain a SP cell fraction, we first isolated SP cells by fluorescence activated cell sorting, followed by culturing in serum-free medium (SFM) using the SCC25 tongue cancer cell line, so that SP cells were able to be propagated to maintain the CSC property. Differential expression profile of stem cell markers (ABCG2, Oct-4 and EpCAM) was examined by RT-PCR in either SP cells or non-SP cells. Growth inhibition by 5-FU was determined by the MTT assay. Clonogenic ability was evaluated by colony formation assay. SCC25 cells contained 0.23% SP cells. The fraction of SP cells was available to grow in SFM cultures. SP cells showed higher mRNA expression of stem cell markers (ABCG2, Oct-4 and EpCAM) as compared with non-SP cells. Moreover, SP cells demonstrated more drug resistance to 5-FU, as compared with non-SP cells. The clone formation efficiency of SP cells was significantly higher than non-SP cells at an equal cell number (P<0.01). We isolated cancer stem-like SP cells from an oral cancer cell line. SP cells possessed the characteristics of cancer stem cells, chemoresistance, and high proliferation ability. Further characterization of cancer stem-like SP cells may provide new insights for novel therapeutic targets.

Expression of the cancer stem cell markers CD44v6 and ABCG2 in tongue cancer: Effect of neoadjuvant chemotherapy on local recurrence

“…proposed to explain tumor cell heterogeneity, some research has suggested that current therapies fail to prevent cancer relapse and metastasis because of a small, surviving population of CSC (37)(38)(39)(40)(41). We previously demonstrated that the side population cells isolated from an oral cell line possessed the characteristics of cancer stem cells, chemoresistance, and high proliferation ability (24). Moreover, in another study, we estimated that not only the mechanism for the closed margin but also residual cancer cells exhibiting the characteristic of CSC after NAC are involved in the mode of local recurrence (13).…”

“…Elevated expression of ABCG2 has been observed in a number of putative CSCs from various cancers (24,34,35). AGCG2 is a well-known marker useful for identifying and isolating CSCs (22)(23)(24)34,35,47).…”

“…CD44 is a single-pass type I transmembrane glycoprotein and functions as a cellular adhesion molecule for hyaluronic acid, a major component of the extracellular matrix (16,17 (22). This phenotype has been observed in OSCC (23,24). Increasing evidence therefore suggests that ABCG2 is recognized as a CSC marker.…”

Expression of SOX2 in oral squamous cell carcinoma and the association with lymph node metastasis

“…The most frequently applied method for identifying CSCs is flow cytometry, which detects cells with the ability to excrete the vital DNA dye Hoechst 33342 (34)(35)(36)(37)(38). A distinctive small non-dyed population of cells, termed the side population (SP), has been detected in numerous tumors, and is highly tumorigenic (39)(40)(41)(42)(43)(44).…”

Significance of ATP-binding cassette transporter proteins in multidrug resistance of head and neck squamous cell carcinoma

“…The sphere formation assay was conducted as previously described (16). First, the sorted SP and non-SP cells were placed at a density of 1000 cells/ml resuspended in a tumor sphere medium consisting of a serum-free 1:1 mixture of Ham's F-12/DMEM and N2 supplement, 10 ng/ml human recombinant bFGF and 10 ng/ml EGF.…”