Investigation of self-renewal mechanism about CD133+ cancer stem cells in human laryngeal carcinoma Hep-2 cell line.
- Author:
Xudong WEI
;
Jian HE
;
Jiangxia GAO
;
Jing CHEN
;
Jingyu WANG
- Publication Type:Journal Article
- MeSH:
AC133 Antigen;
Antigens, CD;
Apoptosis;
Cell Physiological Phenomena;
physiology;
Down-Regulation;
Flow Cytometry;
Glycoproteins;
Humans;
Laryngeal Neoplasms;
Neoplastic Stem Cells;
physiology;
Patched Receptors;
Patched-1 Receptor;
Peptides;
Receptors, Cell Surface;
genetics;
metabolism;
Signal Transduction;
beta Catenin;
genetics
- From:
Journal of Clinical Otorhinolaryngology Head and Neck Surgery
2014;28(21):1636-1641
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To investigate the self-renewal mechanism of CD133+ cancer stem cells from Hep-2 cell line.
METHOD:The CD133+ cells were sorted by flow cytometry from Hep-2 cell line. Then the sorted CD133+ cells were cultured in RPMI1640. The ability of self-renewal of CD133+ cells were tested by MTT assay. mRNA and protein expression of self-renewal related genes were detected by western blot and RT- PCR.
RESULT:(3.10 ± 0.21)% of Hep-2 cells expressed the membrane antigen CD133. CD133+ fraction was raised to (90.20 ± 5.51)% by flow cytometry. In vitro culture and growth curve showed CD133+ cells had more active proliferation ability than CD133- cells, which showed statistically significant difference between these two group (P < 0.01). RT- PCR and western blot results showed upregulated mRNA and protein expression of Fas, c-myc, survivin in CD133+ group (P < 0.01). In the same time, the ratio of Bcl-2/Bax gene expression was obviously increased in CD133+ group. Self-renewal related gene such as β-catenin, SHH, SMOH and Bmi-1,Gli-1 were all up-regulated in CD133+ group both in mRNA and protein. On the contrary, PTCH gene was down-regulated.
CONCLUSION:CD133 positive cells are a small proportion of a Hep-2 cell line. The results of this experiment verified that CD133 positive cells owned the properties of cancer stem cells. Upregulated anti-apoptotic gene is the foundatiom of self-renewal mechanism of CD133+ cells. Cancer stem cells related signal pathways such as Hedgehog, Wnt and Bmi-1 pathway are in state of activation. The identification of self-renewal mechanism about cancer stem cell provides a powerful tool to investigate the tumorigenic process in the larynx and to develop therapies targeting to these signal pathways.
