Influence on Biological Behavior of Colon Cancer Stem Cells after RNA Interfering CD133

Abstract

OBJECTIVE To observe the effects of blocking CD133 gene

expression and activation on biological characteristic of the colon cancer stem cells.

METHODS CD133+ colon cancer stem cells (CCSCs) were separated from EpCAMhigh CD44+CCSCs through fluorescence-activated cell sorting (FACS). The proliferation, the capability of spherical cell formation, neoplasia, and the expression of ABCG2 mRNA of CD133+ CCSCs were observed a er the CD133+ CCSCs were infected with LV-CD133shRNA. CD133 negative cells were isolated from EpCAMhigh CD44+ CCSCs with FACS, and the CD133 proteins in CD133- cells were detected with Western blot.

RESULTS CD133+ CCSCs were isolated from EpCAMhigh CD44+ CCSCs using FACS, and they accounted for 89.2% in the stem cells. In the experimental group, after the CD133+ CCSCs were knocked down by LV-CD133shRNA RNAi, the growth pa ern of the cells in the stem cell culture changed into adherent growth from suspended growth, and couldn’t generate spherical cells. Results of MTT assay showed that the CD133+ CCSCs infected with LV-CD133shRNA grew slowly, compared to the cells in the control groups. There was a decrease in the cloning efficiency. The infected cells were transplanted into the BALB/c nude mice. During the observation, no neoplasia was found in the CD133+ cells infected with LV-CD133shRNA. The level of ABCG2 mRNA expression was lowered greatly (P < 0.01). CD133- cells were obtained from the EpCAMhigh CD44+ CCSCs using FACS, in which the expression of CD133 protein was positive.

CONCLUSION CD133 retains the biological characteristics of the colon cancer stem cells.