VEGF siRNA对兔眼碱烧伤后角膜VEGF表达的作用

Modulation of expression of VEGF in cornea by VEGF siRNA after alkali burn in rabbits

目的研究血管内皮生长因子(VEGF)小干扰RNA(VEGF siRNA)对兔眼碱烧伤后角膜VEGF表达的调控作用。方法25只普通家兔以碱烧伤法(1mol/LNaOH溶液)诱导角膜新生血管(CNV)。碱烧伤后立即以脂质体(LF2000)为载体,右眼球结膜下注射VEGF siRNA重组质粒,左眼球结膜下注射pSilencer2.1-U6 hygro空白质粒作为对照。碱烧伤后1、3、5、7、14d,RT-PCR法检测角膜组织中VEGF mRNA的表达,免疫组织化学法检测角膜组织中VEGF蛋白的表达。结果与对照眼相比,实验组在各时间段RT-PCR结果显示角膜组织中VEGF mRNA的表达明显下调,免疫组织化学结果显示角膜组织中VEGF蛋白的表达明显下调,差异有统计学意义(P<0.05)。结论VEGF siRNA能有效地抑制兔眼碱烧伤后角膜组织中VEGF mRNA及蛋白的表达。

Objective The pathogenesis of corneal neovascularization remains unclear. It has been verified that vascular endothelial growth factor （VEGF） is one of its fundamental regulatory factors. The aim of this study was to explore the modulation of the expression of VEGF in cornea by VEGF small interfering RNA（ VEGF siRNA） after alkali burn in rabbits. Methods Corneal neovascularization （CNV） was induced in vivo by alkaline burn （ 1 mol/L NaOH） in the central cornea in the right eyes of 25 rabbits, and the fellow eyes were control eyes, and the normal corneas from another normal rabbit were as blank control. Using liposome（LF2000）as vector,VEGF siRNA recombinant plasmid was injected into the subconjunctiva of the right eyes in experimental group,and pSilencer 2. 1-U6 hygro was injected into the subconjunctiva of the left eyes in the control group. The expression of VEGF mRNA in rabbit cornea was evaluated by reverse transcription-polymerase chain reaction （RT-PCR） ,and the expression of VEGF protein in rabbit cornea was detected by irnmunohistochemical analysis 1 day,3,5,7,14 days after alkaline burn. Results A small amount of VEGF mRNA was expressed in the normal corneal tissue. The expression of VEGF mRNA and protein in cornea in the control group began to rise on the 1st day after alkali burn,peaked on the 5th day and reduced gradually 7 days later. The expression of VEGF mRNA and protein in corneas in experimental group was similar to the control groups. However,compared with the control eyes,the expression of VEGF mRNA and protein in the experimental eyes was significantly down regulated in different time points after alkaline burn（ P 〈 0.05 ）. Conclusion VEGF siRNA can effectively inhibit the expression of VEGF mRNA and VEGF protein in rabbits cornea after alkali burn.