摘要
目的:探讨转染内皮抑制素(Endostatin)基因对酸烧伤引起的大鼠角膜新生血管的基因治疗。方法:通过限制性内切酶酶切反应、聚合酶链式反应、DNA测序及用NCBIBLAST软件与基因库序列比较的方法进行鉴定,鉴定后在大肠杆菌中扩增,用质粒纯化试剂盒抽提纯化;用750g/L硝酸银和250g/L硝酸钾混合烧灼液制作大鼠角膜新生血管模型,用结膜下注射脂质体包裹的质粒pBlast- hEndostatin来进行体内基因治疗。结果:实验证实质粒pBlast-hEndostatin含有人endostatin基因。结膜下注射脂质体包裹的质粒Tel押029-82245172Email押IJO.2000@163.compBlast-hEndostatin对酸烧伤引起的炎症性角膜新生血管有明显抑制作用,术后6,10,15d对角膜新生血管面积的抑制率分别为37%,40%,43%;对角膜新生血管密度也有明显的抑制作用,抑制率达40%。对角膜新生血管长度和角膜炎症细胞没有明显抑制作用。角膜新生血管面积与角膜水肿、角膜混浊呈正相关。结论:用结膜下注射脂质体包裹的内皮抑制素基因可以部分抑制酸烧伤引起的大鼠角膜新生血管。其作用机制是转基因产生的内皮抑制素蛋白直接抑制角膜新生血管的形成,而不是通过抑制炎症反应来抑制角膜新生血管的形成。
AIM: To study the endostatin gene therapy for rat corneal neovascularization induced by acid cauterization. · METHODS: PBlast-hEndostatin and pBlast-Mcs were identified by digesting them with Nhe Ⅰand Sal Ⅰ, PCR reaction, sequence, and Alignments of PCR product with genebank using NCBIblast software. Then, they were purified with Qiagen endofree plasmid maxi kit. Rat corneal neovascularization models were made with 750g/L AgNO3 and 250g/L KNO3 cauterization. The treatment was subconjunctival injection of the pBlast-hEndostatin with the control of pBlast-Mcs. · RESULTS: PBlast-hEndostatin contained human endostatin gene. The rat corneal neovascularization induced by acid cauterization was significantly suppressed by subconjunctival injection of the pBlast-hEndostatin. The inhibition rates were 37%, 40%, 43% respectively on the sixth, tenth and fifteenth day after treatment.PBlast-hEndostation also showed inhibitory effect on the density of corneal neovascularization, and the inhibition rate was 40%. But it showed no inhibitory effect on the length of the newly-formed vessels and corneal inflammatory cells. Area of corneal neovascularization was positively related to corneal edema and corneal opacity.· CONCLUSION: The plasmid of pBlast-hEndostatin contains human endostatin gene. The rat corneal neovascularization induced by acid cauterization can be partly inhibited by subconjunctival injection of the pBlast-hEndostatin mediated by liposome. The mechanism is that the corneal neovascularization is inhibited directly by the endostatin produced by the transfected fibroblast cells, but not by inflammatory reaction.·
出处
《国际眼科杂志》
CAS
2004年第1期60-65,共6页
International Eye Science
基金
中国广东省卫生厅基金资助(No:B200051)~~
