新型抗体偶联胶原包被质粒DNA血管支架的研制  

A Novel Anti-DNA Antibody Modified Coronary Stent for Site-specific Plasmid DNA Delivery

作　　者：金旭[1] 冷希岗[1] 刘兰霞[1] 谷海刚[1] 张超[1] 王海[1] 宋存先[1] 

机构地区：[1]中国医学科学院中国协和医科大学生物医学工程研究所生物医学材料重点实验室,天津300192

出　　处：《中国医学科学院学报》2006年第5期665-669,I0002,共6页Acta Academiae Medicinae Sinicae

基　　金：国家自然科学基金(50473059);天津科学技术委员会重点基金(043803011);教育部博士点基金(20030023004)~~

摘　　要：目的研制一种以支架为基础的高效、局部定位的新型基因投递体系。方法利用双官能偶联剂N-琥珀酰亚胺基3-(2-吡啶二硫基)丙酸酯(SPDP)将支架表面涂布的胶原与抗DNA抗体化学键合,该抗体再与质粒DNA(pEGFP-C1)免疫结合,并加入阳离子脂质体提高细胞转染效率。用125I标记抗DNA抗体,检测支架表面偶联的抗体量。用32P标记pEGFP-C1检测通过上述方法结合在支架上的DNA量,同时以物理吸附组作为对照,通过细胞培养和兔体内实验验证其效果。结果实验组胶原基质携带抗体量约为对照组的15倍(P<0.005)。实验组胶原基质携带质粒DNA量显著高于对照组(P<0.01),释放时间达13d以上。细胞转染实验结果表明,抗DNA抗体连结pEGFP-C1质粒组支架胶原涂层表面有较多GFP转染阳性细胞生长,而未与支架接触的周边细胞几乎无转染。动物实验结果显示约有(2.8±0.7)%血管壁细胞被转染,新生内膜转染率最高,达7%,远隔器官未见载体扩散。结论抗体偶联胶原包被质粒DNA血管支架是一种新型高效、局部定位的基因转染体系。该体系适用于多种基因转染,且基因释放在一定条件下可控,为开发心血管内靶向基因投递体系提供了新思路。cient and site-specifi Objective To explore the feasibility of using an endovascular metal stent as a highly effic gene delivery system. Methods Stents were formulated with a collagen coating. Anti- DNA monoclonal antibodies were covalently bound to the co cinimidyl 3 - （ 2-pyridyldithio DNA on stents were quantifi surface by a cross linking reagent of N-succinimidy1 3-（2-pyridyldithio） propionate （SPDP）. Binding capacity and stability of antibody and plasmid ed by radioactive labeling. The gene transduction efficiency was evaluated in cell culture and in rabbits. Results The amount of antibodies binding on collagen matrix through SPDP reaction was 15 times higher than that of through physical absorption （ P 〈 0. 005 ）. The binding stability of plasmid was significantly better than the control groups （ P 〈 0.01 ）. There was no harmful effect on cell growth with the anti-DNA antibody modified stents. The stents retrieved from cell culture after 72 hours of incubation in A10 cells showed numerous transducted cells only infiltrating the surface coating indicating a highly localized and efficient gene delivery pattern. Results of in vivo gene transfer by this modified stent revealed （2. 8 ± 0. 7 ） % of total cells transduction and the higher transduction location was neointimal layer （ about 7% ）. No distal spread of vector was detectable in the anti-DNA antibody modified stent implantation animals. Conclusions Anti-DNA antibody modified stents represent a novel highly efficient and site-specific gene delivery system which can deliver various kinds of plasmi some conditions. This novel d vectors. The release of plasmid DNA tethered on the stents could be controlled in system provided a novel platform for cardiovascular site-specific gene therapy.

关 键 词：基因治疗 载体 抗体 支架 质粒 胶原 

分 类 号：R318[医药卫生—生物医学工程]