摘要
目的 实体瘤的缺氧微环境限制了放射 基因治疗的效果。本研究利用缺氧反应元件 (hypoxiaresponseelements,HREs)实现放射诱导及缺氧增强肿瘤坏死因子α(TNF α)表达 ,以提高乏氧实体瘤的放射 基因治疗效果。方法 将HREs与放射敏感性Egr 1启动子串联 ,构建缺氧 /辐射双敏感性HRE Egr启动子及其调控的TNF α表达载体 ,用脂质体介导该载体转染肺癌A5 4 9细胞 ,予以照射 (6Gy)和 /或缺氧 (1 %氧浓度 )处理后 ,ELISA法检测转染细胞TNF α表达水平。通过细胞剂量 存活曲线及其参数D0 值 ,计算氧增比 (OER)及增敏比 (SER)。建立裸鼠A5 4 9皮下移植瘤模型 ,计算 5 0 %肿瘤控制所需照射剂量 (TCD50 )及SER。结果 HREs可使缺氧下转染细胞照射后的TNF α表达水平较常氧条件下增加到 3.4倍 ,且转染组D0 (1 .341Gy)和OER(0 .81 )均显著低于对照组 (6 .1 72Gy ,2 .6 5 ) ,SER(4 .6 0 )则显著升高 (P <0 .0 1 )。两组皮下移植瘤亦有相似的改变。 结论 成功构建了缺氧 /辐射双敏感性HRE Egr启动子及其调控的TNF α表达载体 ,该载体转染的A5 4 9细胞及其移植瘤 ,均有显著的放射乏氧增敏作用。
Objective To construct an expression vector of tumor necrosis factorα(TNFα) driven by a hypoxia/radiation dual sensitive promoter and to evaluate the improved efficacy of TNFα expression in lung adenocarcinoma cell line A549 transfected with the vector and its subcutaneous xenografts in nude mice.Methods The gene of hypoxia response elements(HREs) was inserted upstreamly into the promoter of early growth response gene 1(Egr 1) to generate a chimeric promoter HRE Egr.TNF α expression vector driven by hypoxia/radiation dual sensitive promoter HRE Egr was then transfected into A549 cells with lipofectant.After exposure to irradiation and/or hypoxia,expression of TNF α in the transfected cells was analyzed by ELISA,and sensitization enhancement rates(SER) of the constructed vectors were evaluated via curve simulating of the relative survival rates of transfected cells.The 50% tumor control dose(TCD 50 ) was determined in 40 nude mice bearing subcutaneous xenograft with the transfected cells.Results Expression of TNF α gene in the transfected cells which exposed to 6Gy irradiation was markedly increased under hypoxia.The survival rate of transfected cells which were radiated under hypoxia decreased significantly with comparison of that under normoxia( P <0.01).In in vivo study,much smaller value of TCD 50 (10.4Gy vs control 17.8Gy) and significant regression of xenografts were found in HRE Egr promoter transfected group( P <0.01).Conclusion Hypoxia/radiation dual sensitive promoter HRE Egr can be used to enhance the anti tumor effects of radiation on lung adenocarcinoma cell line A549 and its subcutaneous xenografts in nude mice.
出处
《重庆医学》
CAS
CSCD
2004年第7期962-964,共3页
Chongqing medicine
基金
国家自然科学基金资助项目 (30 30 0 0 97)
关键词
肺肿瘤
放射—基因治疗
缺氧反应元件
lung neoplasm
radio genetic therapy
hypoxia response elements
A549 cell line
