血管内皮生长因子靶向抗体对大鼠角膜碱烧伤后新生血管抑制作用的研究

Inhibitory effects of anti-vascular endothelial growth factor( VEGF) antibody MIL60 on alkali-induced corneal angiogenesis in rats

目的通过对角膜碱烧伤大鼠模型结膜下注射VEGF靶向抗体MIL60,观察其对角膜新生血管(corneal neovascularization,CoNV)生长的影响,初步探讨MIL60抑制大鼠CoNV生长的基本作用机制。方法建立碱烧伤诱导的SD大鼠CoNV模型(右眼造模),根据碱烧伤后第1天结膜下注射的药物不同将72只大鼠随机分为:25 mg·m L^(-1) MIL60组、地塞米松组、MIL60溶剂组和NaCl组。观察并记录CoNV生长情况及角膜的形态变化,通过软件分析Co NV的长度及累及面积。碱烧伤后第3天、第7天、第14天、第21天和第28天分别处死动物,取角膜组织行HE染色和免疫组织化学染色,同时检测角膜中血管内皮生长因子(vascular endothelial growth factor,VEGF)、VEGF受体-1(VEGF receptor-1,VEGFR-1)、VEGFR-2和基质金属蛋白酶-9(matrix metallopeptidase-9,MMP-9)蛋白的表达。结果在各个时间点,与MIL60溶剂组和NaCl组比较,地塞米松组、25 mg·m L^(-1) MIL60组CoNV面积和长度显著减少,差异均具有统计学意义(均为P<0.01);25 mg·m L^(-1)MIL60组的CoNV长度和面积与地塞米松组相当(均为P>0.05)。同时,MIL60还能有效降低角膜组织中VEGF、VEGFR-1、VEGFR-2以及MMP-9蛋白的表达。结论 MIL60能显著抑制角膜碱烧伤后CoNV的生长,并且还能减轻碱烧伤引起的炎症反应。

Objective To investigate the efficacy of an anti-vascular endothelial growth factor （VEGF） antibody,MIL60,in inhibiting corneal neovascularization （CoNV） formation in a rat model of alkali cauterization and its involved mechanisms.Methods Rat CoNV model induced by alkali burn was founded in the right eyes,and then 72 cases were randomly divided into four groups according to the subconjunctival administration of medicine next day after the successful establishment of this model：25 mg·mL^-1 MIL60 group,dexamethasone group,MIL60 solvent group and NaCl group.Then CoNV was observed for recording the its length and the involved area using digital photograph.Next the rats were sacrificed on day 7,14,21 and 28,followed by the collection of rats’ cornea for HE and immunohistochemical staining to analyze the protein expression of VEGF,VEGF receptor-1 （VEGFR-1）,VEGFR -2 and matrix metallopeptidase-9 （MMP-9）.Results At each time point,the area and length of CoNV in the 25 mg·mL^-1 MIL60 and dexamethasone group were significantly less than those in the MIL60 solvent and NaCl group,and the differences were statistically significant （all P〈0.01）,and 25 mg·mL^-1 MIL60 group had the similar CoNV area and length with the dexamethasone group （all P〉0.05）.Moreover,HE and immunohistochemical staining showed that MIL60 could inhibit the protein expression of VEGF,VEGFR-1,VEGFR-2 and MMP-9,which could explain its effective anti-angiogenic activity.Conclusion Subconjunctival administration of MIL60 can significantly inhibit corneal neovascularization formation and alleviate the inflammation in rats suffered from alkali burn.