硝基左旋精氨酸甲脂对碱烧伤后角膜新生血管作用的实验研究

目的:探讨碱烧伤后角膜新生血管形成过程中诱导型一氧化氮合成酶(iNOS)的表达及作用;观察局部应用硝基左旋精氨酸甲脂(LNAME)对角膜新生血管形成的影响。方法:采用碱烧伤诱导角膜新生血管模型,将50只SD大鼠随机分成5组:A组为治疗I组,B组为治疗Ⅱ组,C组为治疗Ⅲ组,D组为对照组和E组为正常组。治疗组(A、B、C组)碱烧伤后分别滴用0.3%,1%,3%的硝基左旋精氨酸甲脂滴眼液,对照组(D组)滴赋形剂,E组滴用30g·L-1的硝基左旋精氨酸甲脂滴眼液。采用裂隙灯照相,免疫组化染色,多媒体彩色图像病理分析系统分别计算8h、1,4,7,14,28d共6个不同时间点的角膜新生血管面积和诱导型一氧化氮合成酶免组染色的平均光密度值。并观察局部用药的毒副反应。结果:碱烧伤后大鼠角膜上诱导型一氧化氮合成酶主要表达于基质层中浸润的炎性细胞和新生血管内皮细胞。iNOS的表达与角膜新生血管面积在治疗组B、C组与治疗组A和对照组之间在统计学上的差异有显著意义(P<0.05)。重复局部给药良好耐受。结论:局部应用LNAME对炎性角膜新生血管有一定的治疗作用;iNOS表达水平与炎性角膜新生血管明显相关;局部应用LNAME无明显毒副作用,是安全有效的给药方式。

Effects of N G -Nitro-L-Arginine Methyl Ester on Corneal Neovascularization after Alkali Burn

Objective: To investigate the expression of inducible nitric oxide synthesis (iNOS) in cornea after alkaline burn, and to observe the effect of topically applied N G-Nitro-L-Arginine Methyl Ester(L-NAME) on corneal neovascularization (CNV) in rats. Methods: The corneal neovascularization model was established by putting the round filter paper with the diameter of 3 mm which were immersed in 1 mol·L -1 NaOH solution on the rat’s cornea for 90 seconds. Fifty Sprague-Dawley rats were randomly divided into 3 treated groups (group A, B and C), a control group and a normal one. The L-NAME drops with different concentrations were applied topically to rat corneas four times daily for four weeks on each group: 3 g·L -1 in group A, 10 g·L -1 in group B, 30 g·L -1 in group C, carrier solution in group D(controls) and 30 g·L -1 in group E(normal). Slit-lamp biomicroscopy photography, and immunohistochemical staining were done. The area of CNV was calculated and the average optic density of immunohistochemical staining of iNOS was measured after 8 hours and 1, 4, 7, 14 and 28 days by imaging analysis system. Meanwhile, the toxicity effects of topically applied L-NAME were tested. Results: The expression of iNOS mainly located into the inflammatory cells in corneal stroma layer and endothelial cells of neovascularization. The difference in the expression of iNOS and the area of neovascularization between group B, C and group A, D were statistically significant (P<0.05). All rats treated with repeatedly and topically applied L-NAME were well tolerated. Conclusion: Topical application of L-NAME is effective in the inhibition of corneal neovascularization induced by alkaline burn. The levels of iNOS were markedly related to inflammatory corneal neovascularization. In addition, it is a safe way to apply L-NAME topically to protect the cornea from obvious side effects.