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


CONCLUSIONS
Tumor spheres in which enriched cancer stem cells can be generated under serum-free culture condition with EGF and bFGF.
Li
Ying-fei
YF
Institute for Digestive Diseases, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. liyingfei304@yahoo.com.cn
Xiao
Bing
B
Lai
Zhuo-sheng
ZS
Tu
San-fang
SF
Wang
Yuan-yuan
YY
Zhang
Xiao-lan
XL
D000071916:AC133 Antigen; D015703:Antigens, CD / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D016895:Culture Media, Serum-Free; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D014411:Neoplastic Stem Cells / Q000166:cytology* / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism; D018874:Spheroids, Cellular / Q000166:cytology*
D000071916:AC133 Antigen; D015703:Antigens, CD; D016895:Culture Media, Serum-Free; D006023:Glycoproteins; D018960:Hyaluronan Receptors; C491297:MSI1 protein, human; D009419:Nerve Tissue Proteins; C000606023:PROM1 protein, human; D010455:Peptides; D016601:RNA-Binding Proteins