转录因子SP1诱骗性寡核苷酸对猪血管内皮细胞α半乳糖苷链表达的影响  

The effect of transcription factor SP1 decoy oligodeoxynucleotides on expression of α Gal in SV-40-PED cells

作　　者：黄亚冰[1] 王璐[1] 谢林[1] 潘桃[1] 郭晖[1] 李荣[1] 陈松[1] 陈实[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院器官移植研究院教育部/卫生部器官移植国家重点实验室,湖北武汉430030

出　　处：《医学研究生学报》2007年第5期486-489,I0004,共5页Journal of Medical Postgraduates

基　　金：国家863高技术研究发展计划基金资助项目(批准号:2003AA205009);高等学校博士学科点专项基金资助项目(批准号:20040487077)

摘　　要：目的:探讨转录因子SP1诱骗性寡核苷酸(decoy ODN)对猪血管内皮细胞系(SV-40-PED)膜蛋白α半乳糖苷链(αGal)表达的影响。方法:SP1诱骗性寡核苷酸转染猪血管内皮细胞系,细胞爬片后荧光显微镜下观察αGal抗原表位的表达;蛋白印迹法(Western blot)检测蛋白表达情况;RT-PCR法检测SV-40-PED转染前后α1,3半乳糖转移酶(α1,3GT)mRNA。结果:荧光显微镜下细胞爬片可见转染组(decoy ODN组)细胞膜荧光表达明显减弱,部分部位可见点状荧光缺损。Western blot显示decoy ODN/PED的αGal相对吸光度值为48.2±0.9,仅为空转染组(只含有转染试剂oligofectamine)91.6±1.7的52.6%(P<0.05)。转染组α1,3GT基因异构体的mRNA表达量(异构体1为0.42±0.20;异构体2为0.27±0.12)显著低于空转染组(异构体1为0.72±0.17;异构体2为0.50±0.19;P<0.05),但错配组(错义decoy ODN组)及空转染组相比差异无统计学意义(P>0.05)。结论:SP1 decoy ODN可以靶向性抑制猪血管内皮细胞系αGal基因的表达。Objective：To investigate the role of transcription factor SP1 decoy oligodeoxynucleotides （ODN） on expression of α Gal in SV-40-PED cells. Methods：Immortalized porcine aortic endothelial cells of the PED line were cultured and transfected with α1, 3galactosyhransferase （α1 ,3GT） specific decoy ODN. Cells transfected with mismatch ODN was used as negative controls. Twenty-six hours later the cells were collected. The expression of α Gal was determined with fluorescence microscope and Western blot. The expression of α1 ,3GT mRNA was examined by RT-PCR. Results：Fluorescence microscopy observed the decreased fluorescence of α Gal after decoy ODN transfection. Western blot showed that the average absorbance of the PED cells transfected with decoy ODNs was （48.2 ± 0.9）. It is 52.6% of the mock group（P 〈0.05）. The expression of α1 ,3GT mRNA in the PED cells transfected with decoy ODN was （ isoform 1,0.42 -0.20;isoform 2,0. 27 ±0. 12） lower in comparison with the mock group（ isoform 1, 0.72± 0.17 ; isoform 2,0.50 ± 0.19 ; both P 〈 0.05）. The expression of α1, 3GT in the mismatch group was not different from those in the mock group （P 〉 0.05 ）. Conclusion：α1,3GT gene reduce actually occurs following transfection of decoy ODN. Porcine endothelial cells can be the targets of decoy ODN.

关 键 词：转录因子 SP1诱骗性寡核苷酸 血管内皮细胞 器官移植 异种 

分 类 号：R392.4[医药卫生—免疫学]