Most C6 Cells Are Cancer Stem Cells: Evidence from Clonal and Population Analyses

Abstract

Cancer stem cells have been isolated from human gliomas and many other parenchymal tumors. It was previously assumed that many established malignant cell lines also contain a rare subpopulation of stem cells. This study was designed to investigate the fraction of cancer stem cells in the C6 glioma cell line using clonal and population analyses, rather than isolating methods, which are based on specific markers. Interestingly, in the serum-containing medium, each of the 67 single C6 cells plated per miniwell was able to generate a clone and subclones, which subsequently gave rise to a xenograft glioma in the BALB/C-nude mouse. The CD133 À C6 cells also possessed clonogenic, self-renewal, and tumorigenic capacities. Moreover, our findings indicated that brief exposure to Hoechst 33342 was harmful to the clonogenicity and proliferation of individual C6 cells. Therefore, the non-sidepopulation cells may be deprived of their stem cell features in the process of Hoechst 33342 staining as a step in isolating a Hoechst-negative side population with flow cytometry. Thus, we concluded that the C6 line was mainly composed of cancer stem cells, although many of them were neither CD133 + nor side population. [Cancer Res 2007;67(8):3691-7]

Breast Tumor Heterogeneity: Cancer Stem Cells or Clonal Evolution?

“…Also, according to the cancer stem cell hypothesis, tumor progression is a result of the metastatic spread of these cells, and cancer recurrence is caused by their resistance to therapy. 10,[14][15][16][17][18][19][20][21][22][23][24][25][26] Supporting evidence. The idea that cancers originate from rare stem cells is a long-standing one, first proposed over a century ago by Cohnheim,27 that is supported by many general observations of biology.…”

“…Indeed, recent evidence has shown that all cells in the C6 glioma cell line have cancer stem cell properties but only under certain conditions. 26,76 PossibLe modeL And finAL remArKs Conclusions. Based on the information presented here, it seems that breast tumor heterogeneity is likely caused by a version of the clonal evolution model that incorporates some features of the cancer stem cell hypothesis (Fig.…”

“…37 Since then, this assay has become the accepted standard method for determining whether cell populations isolated from solid tumors are "cancer stem cells" or not. [20][21]26 Cells are purified from tumor samples based on their cell surface marker expression, ability to form spheres in culture, or exclusion of the dye Hoechst 33342 and are then tested for the ability to form tumors when injected into immunodeficient mice. Cancer stem cells have been reported in many tumor types such as lung, brain, skin, prostate, and colon.…”

“…These approaches can harm certain cells and thereby artificially select for others. 19,26 Also, cell recovery from solid tumors is usually 10% or less, so samples analyzed may not be representative of the original lesions. 19 Therefore, actual cancer stem cells may be missed, while other cells may be inaccurately identified as cancer stem cells.…”

The cancer stem cell hypothesis: in search of definitions, markers, and relevance

“…This possibility is well illustrated by the existence of highly tumorigenic cell lines that can efficiently form xenograft tumors without any obvious stem cell-like subpopulations. 22 The recent finding that, depending on the tumor analyzed, glioblastoma cancer stem cells can be CD133 þ or CD133 À cells, also emphasizes that either we do not have good markers for cancer stem cells or that all tumor cells are tumorigenic just at varying degree. 23 Finally, the occasional genetic divergence of CD24 À/low /CD44 þ and CD24 þ /CD44 þ /À breast cancer cells re-emphasizes the notion that tumors are genetically heterogeneous and there is a continuous selection for the survival of the fittest.…”