Interaction between CD133 and phosphatidylinositol 3-kinase p85 promote the chemoresistance capacity of gastric cancer cells
2017
Objective
By building stable CD133 knock-down and overexpression cell lines, we will study its effects on the ability of multiple chemoresistance and the possible relevant mechanism.
Methods
Firstly, we established KATOⅢ-shCD133 and MKN45-CD133 cell lines, and then tested the interference and overexpression efficiency of CD133 by Western blotting. Secondly, we detected the cells sensibility to 5-fluorouracil (5-Fu) and cisplatin (DDP) by cell counting kit-8 (CCK-8), and measured the protein level of chemoresistance related factors: P-gp, B cell lymphoma/leukemia-2 (bcl-2), B cell lymphoma/leukemia-2 associated X protein (bax) and the activation level of protein kinase B (Akt) by Western blotting. Finally, we examined the interaction of CD133 and p85 by immunoprecipitation.
Results
By Western blotting, we found the protein level of CD133 of the knock-down or overexpression cell lines were significantly lower or higher than the control group. Under the same drug concentrations, 5-Fu and DDP showed higher inhibition rate in the CD133 konck-down group than that of the vector group with the half maximal inhibitory concentration (IC50) value [(54.030 0±0.647 9) μmol/L vs. (133.300 0±4.918 0) μmol/L, P=0.005] and [(2.312 0±0.223 6) μmol/L vs. (24.590 0±2.159 0) μmol/L, P=0.001], while the overexpression group took the opposite sides with the IC50 value [(8.102 0±0.433 1) μmol/L vs. (2.217 0±0.132 4) μmol/L, P=0.010] and [(24.487 0±2.132 0) μmol/L vs. (11.620 0±1.534 0) μmol/L, P=0.030]. After stablyknock-down the expression of CD133, the protein levels of chemoresistance related factor: P-gp, bcl-2 and p-Akt were significantly lower, and the promoting apoptosis factor bax was significantly increased; the overexpression group was still the opposite.
Conclusion
CD133 could obviously increase the chemoresistance capacity of gastric cancer cells, which might be realized through the activation of phosphoinositide 3-kinase (PI3K)/Akt pathway by CD133-p85 interaction.
Key words:
Gastric cancer; CD133; Chemoresistance; Phosphatidylinositol 3-kinase p85
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