蛋白涂层支架携带一氧化氮合酶基因转染小型猪冠状动脉的可行性研究

The Feasibility of Local Plasmid-Mediated Human Inducible Nitric Oxide Synthase Gene Transfer to Coronary Artery with Protein-Coated Metallic Stents in Mini-Swine Model

目的 :探索蛋白涂层支架携带质粒介导人肝脏诱导型一氧化氮合酶 (i NOS)基因转染小型猪冠状动脉可行性。 方法 :使用蛋白支架吸附去内毒素纯化质粒 ,以常规支架置入技术置入小型猪冠状动脉前降支中段。置入后第 7天取出前降支置入段 ,分别提取总核糖核酸 (RAN)并进行逆向多聚酶链反应 (RT- PCR) ,免疫组化染色检测导入人肝脏i NOS蛋白的表达。 结果 :小型猪前降支置入支架处显示人 i NOS基因信使核糖核酸 (m RNA)转录 ,免疫组化染色显示中膜、内膜人i NOS基因表达人 i NOS蛋白的颗粒 ,以平滑肌细胞最明显。 结论 :蛋白涂层支架吸附去内毒素携带人 i NOS基因质粒植入小型猪前降支冠状动脉 ,RT- PCR显示人 i NOS基因的 m RNA转录 ,免疫组化显示人 i NOS蛋白的表达。

Objective:To assess the feasibility of plasmid medicated local transfer of inducible nitric oxide synthase (iNOS) gene to coronary artery using protein coated metallic stents in mini swine medel. Methods:The metallic stent was coated by cross linked gelatin and mounted on 3 0 mm Percutaneous transluminal coronary angioplasty (PTCA) balloon,then Endotoxin free ultrapure Endotoxin free ultrapure Plasmid pcDNA 3hepiNOS under the control of the comegalovirus (CMV) promoter/enhancer was absorbed on the stent.Protein coated stainless steel stents were used as controls.All stents were implanted into the middle segment of left anterior descending artery through 7F large luman guiding catheter.(The ratio of balloon to vessel diameter was 1 1 1 3∶1).Total RNA of stented segment of coronary artery was extracted,its reverse transeription polymerase chain reaction (RT PCR) and immunohistochemical staining were performed through routine methods. Results:At the 7 th day after stenting,RT PCR and immunohistochemical staining confirmed the expression of plasmid pcDNA3hepiNOS mRNA and presence of its protein at gene transered vessels ( n =2) but there was no expression in remote organs.Endothelialization of the vessel was observed in all animals through scanning electromicroscopy. Conclusions:Local plasmid mediated human inducible nitric oxide synthase gene transfer to coronary artery with protein coated metallic stents is feasible in mini swine model.