Adenovirus-mediated siRNA targeting Bcl-xL inhibits proliferation, reduces invasion and enhances radiosensitivity of human colorectal cancer cells

Wei De,Ming Sun,Jinsong Yang,Aiping Zhang,Zhaoxia Wang,Chengyu Lv

doi:10.1186/1477-7819-9-117

Abstract

IntroductionBcl-xL, an important member of anti-apoptotic Bcl-2 family, plays critical roles in tumor progression and development. Previously, we have reported that overexpression of Bcl-xL was correlated with prognosis of colorectal cancer (CRC) patients. The aim of this study was to investigate the association of Bcl-xL expression with invasion and radiosensitivity of human CRC cells.MethodsRT-PCR and Western blot assays were performed to determine the expression of Bcl-xL mRNA and protein in CRC cells and normal human intestinal epithelial cell line. Then, adenovirus-mediated RNA interference technique was employed to inhibit the expression of Bcl-xL gene in CRC cells. The proliferation of CRC cells was analyzed by MTT and colony formation assay. The migration and invasion of CRC cells was determined by wound-healing and tranwell invasion assays. Additionally, the in vitro and in vivo radiosensitivity of CRC cells was determined by clonogenic cell survival assay and murine xnograft model, respectively.ResultsThe levels of Bcl-xL mRNA and protein expression were significantly higher in human CRC cells than in normal human intestinal epithelial cell line. Ad/shBcl-xL could significantly reduce the expression of Bcl-xL protein in CRC cells. Also, we showed that adenovirus-mediated siRNA targeting Bcl-xL could significantly inhibit proliferation and colony formation of CRC cells. Ad/shBcl-xL could significantly suppress migration and invasion of CRC cells. Moreover, Ad/shBcl-xL could enhance in vitro and in vivo radiosensitivity of CRC cells by increasing caspase-dependent apoptosis.ConclusionsTargeting Bcl-xL will be a promising strategy to inhibit the metastatic potential and reverse the radioresistance of human CRC.

Highlights

Bcl-xL, an important member of anti-apoptotic Bcl-2 family, plays critical roles in tumor progression and development
The expression level of Bcl-xL is higher in colorectal cancer (CRC) cell lines than in human intestinal epithelial cell line (HIEC) Semi-quantitative Reverse Transcription (RT)-PCR and Western blot assays were done to determine the expression of Bcl-xL mRNA and protein in four human CRC cell lines (SW480, HT-29, Colo320 and LoVo) and a human intestinal epithelial cell line (HIEC)
The expression levels of Bcl-xL mRNA and protein were obviously higher in human CRC cell lines than in human intestinal epithelial cell line (Figure 1A and 1B)

Summary

Introduction

Bcl-xL, an important member of anti-apoptotic Bcl-2 family, plays critical roles in tumor progression and development. Bcl-xL, an anti-apoptotic member, plays important roles in tumor progression and development [2]. Guichard’ et al showed that short hairpin RNAs targeting Bcl-xL modulated senescence and apoptosis following SN-38 and irinotecan exposure in a colon cancer model [11] Zhu and his colleagues found that the combination of Bcl-XL-specific small interfering RNA and 5-FU had additive effect on the inhibition of 5-FU-resistant cells [12]. Nita’et al showed that the suppression of Bcl-X(L) expression by the specific antisense ODNs could increase the sensitivity of CRC cells to 5-FU [13] From these experimental data, it was concluded that Bcl-xL might play important roles in the chemoresistance of human CRC. To the best of our knowledge, there have been no reports about the correlation between Bcl-xL expression and metastasis or radioresistance of CRC cells

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