摘要
目的研究ARHI基因对人胰腺癌细胞株PANC-1和MiaPaCa-2凋亡的影响,探讨其诱导凋亡的作用机制。方法分别应用反转录聚合酶链反应(RT-PCR)和免疫印迹法(Western blot)检测稳定转染的胰腺癌细胞中ARHI和死亡相关蛋白激酶1(DAPK-1)的mRNA和蛋白表达,应用流式细胞仪检测稳定转染的胰腺癌细胞中发生细胞凋亡的百分率。结果转染ARHI和空载体质粒后,无转染组和空载体组均无ARHI mRNA和蛋白的表达,ARHI组可见ARHI mRNA和蛋白的表达;ARHI组PANC-1、MiaPaCa-2细胞凋亡的百分率分别为(32.9±6.7)%和(39.5±6.0)%,均高于空载体组的(13.8±2.9)%和(15.2±1.8)%(P<0.01);在稳定转染的PANC-1和MiaPaCa-2细胞中,ARHI组DAPK-1基因和蛋白表达均较空载体组和无转染组上调。结论 ARHI可明显的诱导胰腺癌细胞发生凋亡,DAPK-1参与了ARHI引起凋亡的机制。ARHI表达下调可能在胰腺癌的发生发展中起了重要的作用。
Objective To study the effects of apoptosis of human pancreatic cancer cell line PANC-1 and MiaPaCa-2 cells induced by ARHI gene and to investigate its mechanism. Methods The mRNA and protein expression of ARHI and death associated protein kinase 1 （DAPK-1） were analyzed by reverse transcriptase polymerase chain reaction（RT-PCR） and Western blot methods respectively in stable transfected pancreatic cells. Flow cytometry was used to analyze the effect of ARHI on the apoptosis of stable transfected cell line. Results The ARHI stable transfected pancreatic cancer cells produced obvious expression of ARHI protein and mRNA, but failed to induce detectable ARHI expression in empty plasmid transfected cells. In ARHI group, PANC-1, MiaPaCa-2 cells apoptosis rate were （32.9±6.7）% and （39.5±6.0）% respectively,and in vector group they were （13.8±2.9）0/00 and （15.2± 1.8）%. ARHI increased the percentage of apoptosis of cell and induced apoptosis of stable transfected cell line, compared to vector（ P〈0.01）. ARHI up-regulated expression of DAPK-1 protein and mRNA in stable transfected pancreatic cell,compared with vector controls. Conclusion Up-regulation of DAPK-1 by ARHI might be associated with the suppression of apoptosis. ARHI may play an important role in the pancreatic carcinogenesis.
出处
《临床荟萃》
CAS
2011年第24期2145-2148,共4页
Clinical Focus
基金
河北省高等学校科学技术研究青年基金项目(2010118)
河北省医学科学研究重点课题计划(20110156)
关键词
胰腺肿瘤
基因
细胞凋亡
细胞培养技术
逆转录聚合酶链反应
pancreatic neoplasms
genes
apoptosis
cell culture techniques
reverse transcriptase polymerase chain reaction
