本文選題：ATRX + RNA干擾　； 參考：《吉林大學(xué)學(xué)報(bào)(醫(yī)學(xué)版)》2017年03期

【摘要】：目的:以RNA干擾(RNAi)沉默肺癌H460細(xì)胞ATRX基因,探討電離輻射對(duì)肺癌H460細(xì)胞凋亡的影響及其作用機(jī)制。方法:以靶向ATRX基因的慢病毒表達(dá)載體轉(zhuǎn)染293T細(xì)胞,所得慢病毒2次感染肺癌細(xì)胞H460,通過(guò)puromycin陽(yáng)性篩選獲得ATRX沉默的細(xì)胞模型,命名為sh-ATRX1-H460、sh-ATRX2-H460和sh-ATRX3-H460細(xì)胞,以sh-control-H460細(xì)胞作為對(duì)照。根據(jù)沉默結(jié)果分為sh-control-H460組和shATRX3-H460組,分別給予0、2和8Gy X射線(xiàn)照射。Western blotting法檢測(cè)細(xì)胞中ATRX、多聚腺苷酸二磷酸核糖基聚合酶1(PARP1)和caspase-3蛋白表達(dá)量;AnnexinⅤ-FITC/PI試劑盒和流式細(xì)胞術(shù)檢測(cè)各組細(xì)胞凋亡率。結(jié)果:成功獲得ATRX沉默的肺癌sh-ATRX3-H460細(xì)胞模型。與照射前比較,2和8Gy X射線(xiàn)照射后sh-control-H460組細(xì)胞中ATRX蛋白表達(dá)量增加,sh-ATRX3-H460組細(xì)胞中無(wú)ATRX蛋白表達(dá);與照射前比較,照射后sh-control-H460組和sh-ATRX3-H460組細(xì)胞凋亡率均明顯升高(P0.05或P0.01);8Gy X線(xiàn)照射后sh-ATRX3-H460組細(xì)胞凋亡率明顯高于sh-control-H460組(P0.01);2組細(xì)胞中cleaved PARP1蛋白表達(dá)量均增加且規(guī)律相似;sh-control-H460組細(xì)胞中procaspase-3蛋白表達(dá)量無(wú)明顯變化,8Gy X射線(xiàn)照射后sh-ATRX3-H460組細(xì)胞中procaspase-3蛋白表達(dá)量明顯增加。結(jié)論:RNAi可以實(shí)現(xiàn)ATRX沉默,ATRX沉默能夠增加輻射誘導(dǎo)的細(xì)胞凋亡,其機(jī)制可能與PARP1-caspase-3通路有關(guān)聯(lián)。
[Abstract]:Aim: to study the effect of ionizing radiation on apoptosis of H460 cells and its mechanism by silencing the ATRX gene of H460 cells by RNA interference RNAi. Methods: the lentivirus expression vector targeting ATRX gene was transfected into 293T cells. The cell model of ATRX silencing was obtained by puromycin positive screening. The cells were named sh-ATRX1-H460h-ATRX2-H460 and sh-ATRX3-H460 cells, and sh-control-H460 cells were used as control. According to the silencing results, they were divided into sh-control-H460 group and shATRX3-H460 group. ATRX, polyadenylate ribonucleotidyl polymerase 1 (PARP1), caspase-3 protein expression and Annexin 鈪，

本文編號(hào)：1965158