血管内皮生长因子基因转染对犬肾动脉内膜损伤后的作用

目的:观察血管内皮生长因子(VEGF)基因转染肾动脉内膜后的表达,以及对肾动脉内膜损伤后的作用。方法:利用分子生物学技术,将pcD2/VEGF121真核表达质粒及pcD2空载质粒转化大肠杆菌XL1-Blue,大量提取质粒。试验犬14只采用自身对照方法将左右肾动脉随机分为治疗侧和对照侧。行双侧肾动脉造影,将PTCA球囊送入肾动脉中段加压并拉伤动脉内膜,再将涂布pcD2/VEGF121质粒及pcD2空载质粒的球囊分别导入治疗侧及对照侧损伤段动脉加压维持5min,以使基因转染动脉内膜。分别于术后1、6及10周行肾动脉造影后将犬处死,取损伤段肾动脉,制成病理切片进行HE染色及免疫组织化学染色,观察肾动脉内膜的变化和VEGF蛋白的表达,在电镜下观察动脉内膜的超微结构变化,以判断VEGF基因对犬肾动脉损伤后的作用。结果:肾动脉造影结果显示术后6周双侧肾动脉较术前均轻度狭窄,术后10周狭窄程度更明显,但治疗侧和对照侧之间狭窄程度差异无统计学意义;HE染色显示术后6周双侧肾动脉内膜较术前均轻度增生,10周时增生程度更明显,但治疗侧和对照侧肾动脉内膜增生程度均差异无统计学意义。免疫组织化学染色显示1周时,治疗侧肾动脉内膜及平滑肌的VEGF蛋白表达量明显高于对照侧,6周时仍有差异,但其差异减小;电镜显示:与对照侧比较,术后10周治疗侧动脉内膜覆盖较完全。结论:VEGF基因转染动脉壁后可以促进损伤动脉的内膜化,但不能阻止动脉内膜的过度增生及动脉损伤后形成的狭窄。

Effects of vascular endothelial growth factor gene transfection on injured renal artery intima of dogs

Objective:To study the expression of vascular endothelial growth factor(VEGF) gene transfection in renal artery intima of dogs and its efficacy to injured renal artery intima. Method:We produced an eukaryotic expression plasmid of pcD2/VEGF121 and pcD2 plasmid through transferring colibacillus XL1-Blue as a large scale by the technique of molecular biology.The 2 sides of renal arteries of 14 dogs were randomly divided into 2 groups: therapy sides and control sides.Renal arteriography of 14 dogs were performed and then the PTCA balloons was sent into renal arteries and injured the renal artery intima by compression of balloon.The balloons coated pcD2/VEGF121 plasmid and pcD2 plasmid was respectively sent into therapy sides,and control sides and transferred the gene to renal artery intima by compression of balloon for 5 minutes.Repeated renal arteriography were performed at 1st week,6th week and 10th week respectively after operation and then killed dogs to take the injured renal arteries for HE staining and immunohistochemistry after making pathological section.We observed the changes of renal artery intima,the expression of VEGF protein and ultrastructure of renal artery by HE staining,immunohistochemistry and electron microscope to identify the efficacy of VEGF gene to the injured renal artery of dogs. Result:Light stenosis was observed in 2 sides of renal artery at 6th week after operation,and stenosis became significant at 10th week.But there was no significant difference between therapy sides and control sides.Proliferation was observed in 2 sides at 6th week after operation and became significant at 10th week,there was no significant difference between therapy sides and control sides.Expression of VEGF protein in therapy sides was significantly higher in 1st week after operation than control sides,and the difference became weak at 6th week.There was better intima covering in therapy sides than control sides at 10th week. Conclusion:VEGF gene local delivery could accelerate formation of intima but could not prevent the excessive proliferation of intima and stenosis after injury.