肝素酶siRNA联合亚砷酸抑制胰腺癌侵袭性实验探究.docVIP

肝素酶siRNA联合亚砷酸抑制胰腺癌侵袭性实验探究 摘 要：目的：观察Hpa小干扰RNA（small-interfering RNA, siRNA）联合亚砷酸对胰腺癌侵袭力的影响，为Hpa-siRNA联合亚砷酸治疗胰腺癌提供实验依据。方法：将3条Hpa干扰靶序列Hpa1-siRNA，Hpa2-siRNA，Hpa3-siRNA稀释退火，分别与线性化pGenesil-1.1 质粒表达载体的连接制成pGenesil1.1/Hpa1-siRNA、pGenesil1.1/Hpa2-siRNA、pGenesil1.1/Hpa3-siRNA等重组质粒。然后将上述目的基因转染SW1990胰腺癌细胞，并设空白对照和阴性对照组。RT-PCR检测转染后Hpa-mRNA表达，Western blot检测转染后Hpa蛋白表达。采用Transwell小室试验检测转染后Hpa-siRNA和经亚砷酸处理SW1990细胞体外侵袭能力。结果：Hpa-mRNA和Hpa蛋白在基因转染前后的表达，NC和HK组无明显差别，Hpa1-siRNA，Hpa2-siRNA，Hpa3-siRNA三组均明显下降，Hpa2-siRNA抑制效率优于Hpa1-siRNA和Hpa3-siRNA（P0.05）。随着亚砷酸浓度从0.5、1.0、2.0mg/ml递增，侵袭抑制率也随之升高，分别为7.78%、46.8%及76.8%，差异有统计学意义（χ2=71.633，P0.05）。Hpa2-siRNA联合亚砷酸后均能显著抑制SW1990胰腺癌细胞侵袭力，抑制率均达100%。结论 ：Hpa-siRNA和亚砷酸对SW1990胰腺癌细胞侵袭力具有协同抑制作用。 关键词：乙酰肝素酶；小片段干扰RNA；亚砷酸；胰腺癌，侵袭力 中图分类号：R285.5文献标识码：A文章编号：1673-7717(2011)12-2657-05 Research on Heparanase siRNA Combined with Arsenous Acid Inhibiting Invasion of Pancreatic Cancer Cells in Vitro ZHENG Jiaping1,2,YANG JianMin3,Ru QingJing1 (1.Zhejiang University of Traditional Chinese Medicine,Hanghzou 310053,Zhejiang,China; 2.Zhejiang Tumor Hospital,Hangzhou 310022,Zhejiang,China; 3.Zhejiang People’s Hospital,Hangzhou 310014,Zhejiang,China) Abstract:Objective：To observe invasion inhibiting effect of Hpa small-interfering RNA (siRNA) combined with arsenous acid on pancreatic cancer cells, so as to provide the theoretical foundation for further clinical use in the cancer treatment. Methods：Target gene (Hpa1-siRNA，Hpa2-siRNA，Hpa3-siRNA) fragments were diluted, annealed and connected with linear plasmid expression vector pGenesil-1.1, then pGenesil1.1/Hpa1-siRNA, pGenesil1.1/Hpa2-siRNA and pGenesil1. 1/Hpa3-siRNA were reconstructed. The Hpa-siRNA was transfected into SW1990 pancreatic cancer cell by DOTAP Liposomes meanwhile the empty control and negative control group were designed. Those target genes were respectively transfected into Hpa-mRNA and Hpa protein expression was detected by RT-PCR and Western blot respectively. Transwell cab model was