Screening glioma stem cells in U251 cells based on the P1 promoter of the CD133 gene

Wang,Xiaofeng; Chen,Lu; Xiao,Zhongdi; Wang,Yali; Liu,Tiemei; Zhang,Tianfu; Zhang,Yucheng

doi:10.3892/ol.2016.4966

Screening glioma stem cells in U251 cells based on the P1 promoter of the CD133 gene

Authors:
- Xiaofeng Wang
- Lu Chen
- Zhongdi Xiao
- Yali Wang
- Tiemei Liu
- Tianfu Zhang
- Yucheng Zhang
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Affiliations: Department of Stomatology, China‑Japan Union Hospital, Jilin University, Changchun, Jilin 130033, P.R. China, Department of Tumor and Blood Disease, The Affiliated Hospital to Changchun University of Chinese Medicine, Changchun, Jilin 130021, P.R. China, Department of General Surgery, General Hospital of Daqing Oil Field, Daqing, Heilongjiang 163001, P.R. China, Department of Blood Transfusion, China‑Japan Union Hospital, Jilin University, Changchun, Jilin 130033, P.R. China, Scientific Research Center, China‑Japan Union Hospital, Jilin University, Changchun, Jilin 130033, P.R. China
Published online on: August 5, 2016 https://doi.org/10.3892/ol.2016.4966
Pages: 2457-2462
Copyright: © Wang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Cluster of differentiation (CD)133 is an important cell surface marker of glioma stem cells (GSCs). The transcription of the CD133 gene is controlled by five alternative promoters (P1, P2, P3, P4 and P5), which are expressed in a tissue‑specific manner. In the present study, gene recombination technology was used to construct two types of gene expression vectors that contained the P1 promoter of the CD133 gene, which regulated either the neomycin‑resistance gene or the herpes simplex virus thymidine kinase (HSV-TK) gene. Following the stable transfection of U251 glioblastoma cells with these two gene vectors, the cells expressing the P1 promoter that regulated the neomycin‑resistance gene were named CD133 (+) cells, while the cells expressing the P1 promoter regulating the HSV‑TK gene were called CD133 (‑) cells. The expression of CD133 was detected by flow cytometry and reverse transcription‑quantitative polymerase chain reaction. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was used to assess cell proliferation ability, while the cell cycle was analyzed by flow cytometry, and a clone formation test was performed to evaluate the invasive capability of the cells. The results demonstrated that, due to CD133 expression, the cell proliferation ability and the invasive capability of CD133 (+) cells were significantly higher than those of CD133 (‑) cells. In conclusion, the present study successfully established a novel method of screening GSCs in U251 cells based on the P1 promoter of the CD133 gene.

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