Cancer stem cells have been isolated from human lung cancer and cancer cell lines. This study was designed to investigate the fraction of cancer stem cells in the A549 and H446 cell lines by cloning and tumorigenic analyses. We found that >45% of A549 and H446 cells formed large clones that are able to generate subclones and subsequently give rise to xenograft tumors in the BALB/C-nude mouse. Both CD133+ and CD133− cells of A549 and H446 possessed stem cell characteristics and exposure to Hoechst 33342 affected the clonogenicity and proliferation of single A549 and H446 cell. Moreover, based on cell cycle phase distribution and growth curve, most A549 and H446 cells may not be stem cells, but they acquired stem cell characteristics when they were alone. Therefore, these cells that formed large clones should be called cancer initiating cells, not stem cells. Thus, we concluded that >45% of A549 and H446 cells were cancer initiating cells, although many of them were neither CD133+ cells nor side population cells.

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