The inhibitory effect of silencing RAGE gene expression by shRNA on the proliferation of prostate cancer cells
10.3781/j.issn.1000-7431.2010.03.006
- VernacularTitle:shRNA沉默RAGE基因表达对前列腺癌细胞增殖的抑制作用
- Author:
Xinping SUN
;
Chunyan MA
;
Yulian JIAO
;
Yunyun QU
;
Min ZHU
;
Xiaowen LIU
;
Jie XU
;
Yueran ZHAO
- Publication Type:Journal Article
- Keywords:
Prostatic neoplasms;
Glycosylation end products,advanced;
RNA,small interfering;
Cell,DU145
- From:
Tumor
2010;(3):199-204
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To construct a specific small hairpin RNA (shRNA) expressing vectors against human receptor for advanced glycation end product (RAGE) gene and study its inhibitory effect on the proliferation of androgen-independent prostate cancer cells DU145. Methods:Four RAGE specific oligonucleotides were designed and synthesized. These oligonucleotides were annealed to forill double strand DNA fragments and this fragment was cloned into psi-U6 plasmid. The recombinants were transfected into RAGE-overexpressing sub DU145-2C1 cells. Cellular morphology and transfection efficiency were observed under fluorescence microscope. The inhibitory effect of RAGE shRNA construct on RAGE mRNA and protein expression was examined with semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting, respectively. The cellular proliferation was detected with cell counting kit-8 (CCK-8). Scratch test was used to observe the migration of DU145 cells.Results:RAGE shRNA expression plasmids were successfully constructed and transfected into sub DU145-2C1 cells. It can effectively inhibit the expression of RAGE mRNA (P<0.05). The inhibitory effects of shRNA RAGE-1 (R1) was the most stronger. The RAGE mRNA expression was inhibited by 84% and RAGE protein expression was inhibited by 27%. Compared with negative control, the proliferation potential was significantly decreased in shRNA RAGE-transfected cells. The cell migration capability had no significant changes. Conclusion:RAGE shRNA effectively inhibited the expression of RAGE mRNA and protein and suppressed the proliferation of DU145 cells in vitro.
