[Spheres isolated from Colo205 cell line possess cancer stem-like cells under serum-free culture condition]

Ying-fei Li, Bing Xiao, Zhuo-sheng Lai, San-fang Tu, Yuan-yuan Wang, Xiao-lan Zhang
Nan Fang Yi Ke da Xue Xue Bao, Journal of Southern Medical University 2008, 28 (2): 236-40

OBJECTIVE: Isolation and expansion tumor spheres from colorectal cancer cell line Colo205 cultured in serum-free medium(SFM) supplemented with human recombinant EGF and bFGF.

METHODS: Colo205 cells were cultivated in SFM,while cells cultivated in serum-supplemented medium(SSM) served as the control. Cells morphology were observed by optical microscope, and expression of intestinal stem cells marker Musashi-1 was detected by immunocytochemical. To induce cell differentiation, tumour spheres were cultivated without EGF and bFGF in the presence of 10% serum. Then we analysed expressions of stem cell surface markers CD133 and CD44 among undifferentiated cell, post-differentiated cells and routine Colo205 cells under serum-supplemented culture condition by flow cytometry. At last we compared cell cycle and spectral karyotype between two groups.

RESULTS: In SFM consisting of EGF and bFGF, a minority of Colo205 cells could survive, proliferate and form the suspended tumor spheres. We detected high Musashi-1 expression in these cells. Compared with the SSM group and the post-differentiation SFM group, the expressions of CD133 and CD44 were significantly increased in the undifferentiated SFM group (P<0.05). There was no statistical difference in the expression of CD133 and CD44 between the post-differentiation SFM group and the SSM group (P>0.05). Cell cycle analysis indicated that tumor spheres were of a high proliferation state.We could not find any noticeable difference in the number of chromatosomes between the SFM group and the SSM group.

CONCLUSION: Tumor spheres in which enriched cancer stem cells can be generated under serum-free culture condition with EGF and bFGF.

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