Chinese Journal of Tissue Engineering Research ›› 2015, Vol. 19 ›› Issue (45): 7286-7291.doi: 10.3969/j.issn.2095-4344.2015.45.013

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Protein expression and biological characteristics of colon cancer stem cells: culture and screening

Zhang Dong-xing1, Yan Deng-guo 2   

  1. 1Department of General Surgery, Guizhou Provincial People’s Hospital, Guiyang 550002, Guizhou Province, China; 2Department of Anorectal Surgery, Affiliated Hospital of Guizhou Medical University, Guiyang 550001, Guizhou Province, China
  • Received:2015-09-28 Online:2015-11-05 Published:2015-11-05


BACKGROUND: EpCAMhighCD44+ colon cancer stem cells have been isolated in recent years, and most of them are related to the study of isolating tumor stem cells in colorectal cancer cell lines. There are less reports on the function of colon cancer stem cells.
OBJECTIVE: To isolate, screen and culture colon cancer stem cells from colorectal carcinoma tissues and to explore the protein expression and biological characteristics of colon cancer stem cells.
METHODS: The primary cells were isolated from the colon cancer tissues. EpCAMhighCD44+ cells were detected and sorted by flow cytometry. Then, these cells were cultured in serum-free medium. After the cells were 
replanted subcutaneously into the mice, we observed the daily performance and tumor growth in the mice. When the tumor diameter was above 1 cm, tumor tissues were taken for immunohistochemical testing. The growth of EpCAMhighCD44+ cells cultured in the serum-free medium was observed; expressions of neutral epithelial mucin and CK-20 in EpCAMhighCD44+ cells were detected.
RESULTS AND CONCLUSION: EpCAMhighCD44+ colon cancer stem cells were detected in the colon cancer tissues, and the proportion of EpCAMhighCD44+ cells in primary colon cancer cells was 1.7%-38%. After 24 hours of serum-free culture, there were a few of small suspended cell spheres which were incompact. After 21 days, dense cell spheres with uniform size were found. The formation of transplanted tumor was found in the nude mice. Hematoxylin-eosin staining showed the transplanted tumor had the typical characteristics of colon cancer cells. The expression of neutral epithelial mucin and CK-20 was observed in all the transplanted tumors. Additionally, the proliferation of EpCAMhighCD44+ cells could be promoted by 5-fluorouracil (10, 1, and 0.1 PPC) with time. These findings indicate that EpCAMhighCD44+ colon cancer stem cells in the colon cancer have a certain ability of proliferation, regeneration, differentiation, tumor formation and chemotherapy resistance.

Key words: Stem Cells, Tumor Stem Cells, Neoplastic Stem Cells, Colonic Neoplasms, Cell Culture Techniques, Tissue Engineering