ARHI基因与HuR蛋白在胰腺癌PANC-1细胞中的表达
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摘要
目的观察ARHI基因转染胰腺癌PANC-1细胞后对HuR蛋白的影响。方法采用脂质体法将表达ARHI基因的质粒p LNCX2-ARHI-EGFP和空载体p LNCX2-EGFP转染胰腺癌PANC-1细胞,细胞分为3组:实验组(p LNCX2-ARHIEGFP)、空载体转染组(p LNCX2-EGFP)和PANC-1细胞空白对照组,用G418筛选稳定转染的细胞株;采用PCR和Western blot方法检测ARHI基因表达,采用Western blot方法检测HuR蛋白表达。结果稳定转染ARHI基因组可检测到ARHI基因mRNA和蛋白的表达。根据图像灰度计算蛋白条带相对表达量,稳定转染ARHI基因组HuR蛋白表达(0.2226±0.0644)较空载体对照转染组(0.4063±0.0925,P=0.029)和PANC-1细胞空白对照组(0.4178±0.1319,P=0.022)显著降低。结论 ARHI基因可以导致胰腺癌细胞株中HuR蛋白表达降低。
Objective To investigate the effect of ARHI transfection on the expression of HuR protein in PANC-1 cells. Methods Plasmids p LNCX2-ARHI-EGFP which express ARHI and empty plasmid p LNCX2-EGFP were used to transfect PANC-1 cells,which were divided into three groups: p LNCX2-ARHI-EGFP group,p LNCX2-EGFP group,and blank control. G418 was used to select stable transfected cell lines. ARHI gene expression was detected by PCR and Western blot,and the expression of HuR protein was detected by Western blot. Results PANC-1 cells transfected with p LNCX2-ARHI-EGFP,the mRNA and protein expression of ARHI were detected. HuR protein expression appeared significantly decreased in PANC- 1 cells transfected with p LNCX2-ARHI-EGFP( 0. 2226 ± 0. 0644) compared with the p LNCX2-EGFP-transfected cells( 0. 4063 ±0. 0925,P = 0. 029) and PANC-1 blank control cells( 0. 4178 ± 0. 1319,P = 0. 022). Conclusion ARHI gene in PANC-1 cells may reduce the expression of HuR protein.
Objective To investigate the effect of ARHI transfection on the expression of HuR protein in PANC-1 cells. Methods Plasmids p LNCX2-ARHI-EGFP which express ARHI and empty plasmid p LNCX2-EGFP were used to transfect PANC-1 cells,which were divided into three groups: p LNCX2-ARHI-EGFP group,p LNCX2-EGFP group,and blank control. G418 was used to select stable transfected cell lines. ARHI gene expression was detected by PCR and Western blot,and the expression of HuR protein was detected by Western blot. Results PANC-1 cells transfected with p LNCX2-ARHI-EGFP,the mRNA and protein expression of ARHI were detected. HuR protein expression appeared significantly decreased in PANC- 1 cells transfected with p LNCX2-ARHI-EGFP( 0. 2226 ± 0. 0644) compared with the p LNCX2-EGFP-transfected cells( 0. 4063 ±0. 0925,P = 0. 029) and PANC-1 blank control cells( 0. 4178 ± 0. 1319,P = 0. 022). Conclusion ARHI gene in PANC-1 cells may reduce the expression of HuR protein.
引文
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[2]Yang H,Lu X,Qian J,et al.Imprinted tumor suppressor gene ARHI induces apoptosis correlated with changes in DNA methylation in pancreatic cancer cells[J].Mol Med Rep,2010,3:581.
[3]Yang H,Rao JN,Wang JY.Posttranscriptional regulation of intestinal epithelial tight junction barrier by RNA-binding proteins and microRNAs[J].Tissue Barriers,2014,2:e28320.
[4]Liu L,Christodoulou-Vafeiadou E,Rao JN,et al.RNAbinding protein HuR promotes growth of small intestinal mucosa by activating the Wnt signaling pathway[J].Mol Biol Cell,2014,25:3308-3318.
[5]Blancoo FF,Jimbo M,Wulfkuhle J,et al.The mRNA-binding protein HuR promotes hypoxia-induced chemoresistance through posttranscriptional regulation of the proto-oncogene PIM1 in pancreatic cancer cells[J].Oncogene,2016,35:2529-2541.
[6]Jimbo M,Blanco FF,Huang YH,et al.Targeting the mRNA-binding protein HuR impairs malignant characteristics of pancreatic ductal adenocarcinoma cells[J].Oncotarget,2015,6:27312-27331.
[7]Costanitino CL,Witkiewica AK,Kuwano Y,et al.HuR's role in gemcitabine efficacy in pancreatic cancer:HuRupregulates the expression of the gemcitabine metabolizing enzyme,deoxycytidinekinase[J].Cancer Res,2009,69:4567-4572.
[8]Romeo C,Weber MC,Zarei M,et al.HuR contributes to TRAIL resistance by restricting death receptor expression in pancreatic cancer cells[J].Mol Cancer Res,2016,14:599-611.
[9]王敏,彭宣荣,朱彦丽,等.HuR VEGF和COX-2表达及临床病理因素与早期宫颈癌患者预后关系的分析[J].中国肿瘤临床,2010,37:984-990.
[10]胡跃云,吴宏菊,辛彦,等.HuR蛋白在胃癌组织中的表达及其意义[J].中华临床医师杂志,2011,5:91-95.