Suppression of wingless‑type MMTV integration site family, member 1 expression by small interfering RNA inhibits U251 glioma cell growth in vitro

Dong,Lun; Duan,Xiao‑Chun; Han,Chong‑Xu; Zhang,Hengzhu; Wu,Yongkang

doi:10.3892/ol.2014.2647

Suppression of wingless‑type MMTV integration site family, member 1 expression by small interfering RNA inhibits U251 glioma cell growth in vitro

Authors:
- Lun Dong
- Xiao‑Chun Duan
- Chong‑Xu Han
- Hengzhu Zhang
- Yongkang Wu
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Affiliations: Department of Neurosurgery, Clinical Medical College of Yangzhou University, Yangzhou, Jiangsu 225001, P.R. China, Central Laboratory, Clinical Medical College of Yangzhou University, Yangzhou, Jiangsu 225001, P.R. China
Published online on: October 30, 2014 https://doi.org/10.3892/ol.2014.2647
Pages: 81-85

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Abstract

A Wingless‑type MMTV integration site family, member 1 (Wnt‑1) RNA interference expression vector was constructed during the present study, which was used to transfect the glioma U251 cell line and investigate its effect on glioma. Two 21‑base oligonucleotides complementary to the coding sequence that was flanking the loop sequence were designed to form a DNA hairpin template for the target small interfering RNA (siRNA). The siRNA templates were cloned into the siRNA expression vector, pGPU6/green fluorescent protein (GFP)/Neo and the sequence was confirmed by DNA sequencing. The pGPU6/GFP/Neo‑short hairpin RNA (shRNA)‑Wnt‑1 vector was subsequently transfected into U251 cells, and reverse transcription polymerase chain reaction and western blot analysis were used to evaluate the Wnt‑1 gene silencing effect on U251 cell growth by MTT assay and flow cytometry. The Wnt‑1 protein expression was significantly reduced following transfection with the recombinant plasmid, as determined by western blot analysis of the transfected U251 cells. This transfection exhibited a significantly higher death rate, as shown by MTT. Thus, the present study demonstrated that the pGPU6/GFP/Neo‑shRNA‑Wnt‑1 vector inhibited Wnt‑1 protein expression. However, further investigations regarding the Wnt signaling pathway in glioma pathogenesis are required.

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