ABSTRACT
Objective To investigate the effects of over-expression of homo sapiens eukaryotic translation elongation factor 1 alpha 2 (EEF1A2) on in vitro invasion and lung metastasis of human pancreatic cancer SW1990 cells.Methods Letivirus-mediated delivery of EEF1A2 was used to enhance the expression of EEF1A2 gene in human pancreatic cancer SW1990 cells,the SW1990 cells stably over-expressing EEF1A2 protein (SW1990/EEF1A2 cells) were obtained,and the parent SW1990 cells and SW1990/GFP cells were used as the control,and the expressions of EEF1 A2 mRNA and protein were determined by Real-time PCR and Western blotting.The invasion ability of cells was determined by Transwell assay.The lung metastasis model was established by injection of SW1990 cells into the tail vein.Whole lung tissues were harvested,and visible nodules on tung surface were counted macroscopically 8 weeks later.Results The EEF1 A2 mRNA expression of SW1990/EEF1A2 was 3.252 ± 0.344,which was significantly higher than those in SW1990/GFP cells (1.000 ±0.060) and SW1990 cells (0.944 ±0.041,t =2.255,2.305,P＜0.01) ; the EEF1A2 protein expression was 0.833 ± 0.050,which was significantly higher than those in SW1990/GFP cells (0.247 ± 0.035) and SW1990 cells (0.273± 0.041,t=0.572,0.559,P＜0.01).The ability of invasion of SW1990/EEF1A2 cells was (60 ±4) cells,which was sigmificantly higher than (33 ±4) cells in SW1990/GFP group and (26 ± 3) cells in SW1990 group (t =31.33,34.78,P ＜ 0.01).Furthernore,SW1990/EEF1 A2 cells had a much higher incidence of lung metastasis in nude mice than SW1990/GFP cells and SW1990 cells in vivo (100％ vs.20％,20％,P ＜ 0.05).Conclusions EEF1 A2 over-expression can obviously increase the in vitro invasion and lung metastasis of pancreatic cancer SW1990 cells.
ABSTRACT
Objective: To investigate the effect of targeted gene silencing of homo sapiens EEF1A2 (eukaryotic translation elongation factor 1 alpha 2) on migration and invasion of human pancreatic cancer cell line BxPC-3 in vitro , and to explore its possible molecular mechanism. Methods: The mRNA and protein expression levels of EEF1A2 in four strains of human pancreatic cancer cells were detected by RT-PCR and Western blotting, respectively. EEF1A2-siRNA (small interference RNA) and negative control siRNA were transfected into BxPC-3 cells, respectively. Then the mRNA and protein expressions of EEF1A2 were determined by RT-PCR and Western blotting, respectively. The abilities of migration and invasion of BxPC-3 cells were determined by wound healing assay and Transwell invasion assay, respectively. The changes of expressions of p-Akt and total Akt proteins were detected by Western blotting. Results: Of the four strains of human pancreatic cancer cells, SW1990 cells displayed a low level of EEF1A2, and the remaining three strains of human pancreatic cancer cells including BxPC-3, Patu8988 and Panc-1 all displayed high expressions of EEF1A2. At 48 h post-transfection, the expression levels of EEF1A2 mRNA and protein in BxPC-3 cells transfected with EEF1A2-siRNA were significantly decreased as compared with those transfected with negative control siRNA or without any transfection (P 0.05). Conclusion: Targeted silencing of EEF 1A 2 by siRNA can obviously inhibit the migration and invasion abilities of human pancreatic cancer BxPC-3 cells in vitro . This effect may be associated with Akt signaling pathway regulated by EEF1A2. Copyright © 2012 by TUMOR.
ABSTRACT
Objective To elucidate whether down-regulation of homo sapiens eukaryotic translation elongation factor 1 alpha 2 (EEF1A2) expression induces apoptosis in pancreatic cancer cells and its possible mechanisms. Methods Two siRNAs targeting human EEF1A2 were synthesized and the siRNA/liposome complexes were transfected into the pancreatic cancer cell line BxPC-3. RTPCR and Western blot were used to analyze the change of EEF1A2 expression and the apoptosis rate of BxPC-3 cells was studied using Annexin-V/PI assay. To identify the mechanisms involved, the apoptosis associated proteins such as caspase-3, caspase-8, caspase-9, PARP, cytochrome C and Bid were detected by Western blotting. Results Both EEF1A2-targeting siRNAs reduced the EEF1A2expression, and the No. 2 siRNA inhibited EEF1A2 expression to less than 25 % in mRNA and protein levels. Down-regulation of EEF1A2 expression in BxPC-3 cells enhanced cell apoptosis (15.28% ±3.65%) at a greater level than negative siRNA-expressing cells (10. 11% ± 3. 05%) or mock cells (9.41 % ±4.14 %). Furthermore, reduction of EEF1A2 activated the pro-caspase-8, pro-caspase-3,pro-caspase-9,PARP and Bid to their active forms, and increased the expression of cytochrome C.Conclusions These data suggest that EEF1A2 down-regulation could significantly induce apoptosis of pancreatic cancer cell line BxPC-3, which is likely mediated by the death receptor and mitochondrial apoptotic pathways.