关键词: X射线, 肿瘤坏死因子相关的凋亡诱导配体, 条件复制型腺病毒, MDA-MB-231细胞

Abstract:

Abstract:Objective To construct the conditionally replicative adenovirus vector pAd-Egr1-TRAIL-hTERT-E1A-E1Bp-E1B55Kcarrying early growth response gene-1（Egr1） promoter and tumor necrosis factor related apoptosisinducing ligand (TRAIL) gene,and to observe the effects of the vector combined with 2 Gy irradiationon the TRAIL expression in MDA-MB-231 cells.Methods Egr-1 promotor sequence was cloned from pMD18 T-Egr1,TRAIL was constructed the downstream of Egr1 promoter,pShuttle-Egr1-TRAIL-hTERT-E1A-E1Bp-E1B55K (CRAd.pEgr1-TRAIL) was constructed,after the adenovirus vector was packaged successfully,MDA-MB-231 cells were infected with them and irradiated with X-rays.Real time PCR method and ELISA wereused to detect the expression levels of TRAIL mRNA and protein,respectively.Six groups in theexperiment were set up: control,2 Gy,CRAd.p,CRAd.pEgr1-TRAIL,CRAd.p + 2 Gy and CRAd.pEgr1-TRAIL + 2Gy. Results The recombinant adenovirus vector pAd-Egr1-TRAIL-hTERT-E1A-E1Bp-E1B55K was constructedand packaged successfully. The expression level of TRAIL mRNA in MDA-MB-231 cells transfected withthe vector of 5 MOI for 24 h following 2.0 Gy X-rays irradiation began to increase and arrived tothe top 8 h later in various groups,then declined.The expression level of TRAIL protein in MDA-MB-231 cells began to increase 6 h after irradiation and reached to the peak 24 h later,then declined48 h later.There were significant differences in the expression levels of TRAIL protein betweenCRAd.pEgr1-TRAIL + 2.0 Gy and other groups at the same time point (P<0.01).Conclusion Therecombinant adenovirus vector is obtained successfully,and the TRAIL mRNA and protein expressionlevels in MDA-MB-231 cells can be increased significantly by the vector combined with 2.0 Gy X-raysirradiation.

Key words: X-rays, tumor necrosis factor related apoptosis inducing ligand, conditionallyreplicative adenovirus, MDA-MB-231 cells