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Xiaohua M.,Soochow University of China | Xiaohua M.,Key Laboratory for Stem Cell Research of Jiangsu Province | Yan G.,Soochow University of China | Yan G.,Key Laboratory for Stem Cell Research of Jiangsu Province | And 7 more authors.
Chinese Journal of Cancer Biotherapy | Year: 2013

Objective:To study the impact of CD133 on biological characteristics of human glioma U251 cells by construction a U251 cell line stably overexpression human CD133 gene. Methods: The full length human CD133 cDNA was sub-cloned into retroviral expressing vector pEGZ-Term to obtain recombinant pEGZ-Term-CD133 retrovirus. Then, U251 cells were infected by pEGZ-Term-CD133 retrovirus. The expression of CD133 on infected U251 cells was detected by flow cytometry and real-time PCR. By cell counting and neurosphere formation analysis, the impact of CD133 overexpression on the proliferation and neurosphere formation of U251 cells in vitro was determined. Tumorigenicity was evaluated by subcutaneous injection of CD133 infected U251 cells into the nude mice. Results: The pEGZ-Term-CD133 retrovirus expression vector was constructed successfully and a CD133 stably infected U251 cell line was obtained. Compared with U251-mock and U251 cells, the expressions of CD133 mRNA ([7 400.2 ± 5 003. 4] vs [2. 0 ± 1. 1, 1.0 ±2.2]; all P= 0.0007) and CD133 protein were significantly increased in U251-CD133 cells. pEGZ-Term-CD133 retrovirus infection showed no effect on the proliferation of U251 cells (P >0. 05). However, the neurosphere formation of U251-CD133 cells was obviously higher than U251-mock and U251 cells in the presence of serum free neurosphere medium (I 34. 0 ±7.5 I vs [14. 6 ± 2. 3 ], [11. 5 ± 1. 3 ]; all P < 0. 01). Compared with the U251 -mock cells, the tumor formation time of U251 -CD133 cells was shorter (32 d vs 38 d), the tumorigenesis ratio was higher (100% vs 30%) and the tumor volume was significantly larger ([ 180. 3 ± 146. 8 ] vs [ 4. 0 ±0. 0 ] mm3, P =0. 003) at 41 d when subcutaneous inoculated 1 × 10 5cells. Conclusion: CD133 has no influence on proliferation of glioma U251 cells, but could enhance neurosphere formation and tumorigenicity of U251 cells.

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