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背景:角膜移植后40%～60%的患者发生了角膜新生血管,移植后的角膜新生血管加速了角膜免疫排斥反应的发生。抑制角膜移植后的角膜新生血管有利于延长植片的存活时间,提高角膜移植手术的成功率。目的:探讨强力霉素对角膜移植后角膜新生血管的抑制作用。设计、时间及单位:随机对照,动物实验观察,于2007-03/08在中山大学中山眼科中心(眼科学国家重点实验室,2006DA105054)完成。材料:健康清洁级SD大鼠48只作为角膜移植受体,Wistar大鼠24只作为供体,受体眼为右眼,供体眼为双眼。CD31-PE荧光抗体由美国Sigma公司提供,VEGFELISA试剂盒由美国RapidBio公司提供。方法:建立大鼠同种异体角膜移植模型,48只受体鼠随机分为2组:盐水对照组和强力霉素用药组,24只/组。术前20min以1%阿托品滴眼液散瞳,植片直径为2.75mm,植床直径为2.5mm。术后盐水对照组右眼结膜囊内滴用生理眼水,3次/d;强力霉素用药组右眼结膜囊内滴用1%强力霉素眼药水,3次/d;直至术后30d。主要观察指标:全角膜免疫荧光法检测角膜移植后角膜新生血管的发展,ELISA动态检测角膜组织中VEGF蛋白的表达。结果:①强力霉素用药组植片存活(20.67±3.01)d,比盐水对照组(9.67±2.73)d明显延长(P<0.01)。②角膜移植后3,7,14d盐水对照组新生血管面积相对百分比分别为(4.00±1.00)%,(14.33±4.04)%,(31.33±3.51)%;强力霉素用药组新生血管面积相对百分比分别为(1.67±1.15)%,(4.67±1.53)%,(18.33±1.53)%。其中7,14d时两组比较差异显著(P<0.05)。③角膜移植后强力霉素用药组血管内皮生长因子蛋白水平均低于盐水对照组(P<0.01)。角膜移植后3,7,14d,盐水对照组血管内皮生长因子蛋白水平为(541.00±75.44),(960.00±90.14),(976.00±130.41)ng/g;强力霉素用药组为(115.33±9.29),(239.00±41.62),(361.00±65.20)ng/g,各时间段比较差异显著(P<0.05)。结论:强力霉素能抑制角膜移植后角膜新生血管的生长,延长植片的生存时间。
Background: Corneal neovascularization occurs in 40% -60% of patients after corneal transplantation. Corneal neovascularization after transplantation accelerates the occurrence of corneal immune rejection. Inhibition of corneal neovascularization after corneal transplantation is beneficial to prolong the survival time of the implants and improve the success rate of corneal transplantation operations. Objective: To investigate the inhibitory effect of doxycycline on corneal neovascularization after corneal transplantation. DESIGN, TIME AND UNITS: A randomized controlled and animal experiment was performed at the Zhongshan Eye Center of Sun Yat-sen University (State Key Laboratory of Ophthalmology, 2006DA105054) from March to August 2007. MATERIALS: Forty-eight healthy SD rats were used as corneal transplant recipients. Wistar rats were used as donors, recipient eyes as right eyes and donor eyes as eyes. CD31-PE fluorescent antibody provided by the United States Sigma, VEGF ELISA kit provided by the United States RapidBio company. Methods: Rat models of allograft keratoplasty were established. Forty eight recipients were randomly divided into two groups: saline control group and doxycycline group, 24 rats / group. Preoperative 20min with 1% atropine eye dilatation, diameter of 2.75mm graft, implantation diameter of 2.5mm. The rats in the saline control group received drops of physiological eye water 3 times / d in the conjunctival sac of the right eye and 1% doxycycline eye drops in the right conjunctival sac of the doxycycline group 3 times / d until 30 days after operation . MAIN OUTCOME MEASURES: Whole corneal immunofluorescence was used to detect the development of corneal neovascularization after corneal transplantation. ELISA was used to detect the expression of VEGF protein in corneal tissue. Results: ① Dicomycin treatment group survival (20.67 ± 3.01) d, compared with saline control group (9.67 ± 2.73) d significantly prolonged (P <0.01). ② The relative percentages of neovascularization area in the saline control group at 3, 7 and 14 days after corneal transplantation were (4.00 ± 1.00)%, (14.33 ± 4.04)% and (31.33 ± 3.51)%, respectively. The relative percentage of neovascularization area (1.67 ± 1.15)%, (4.67 ± 1.53)% and (18.33 ± 1.53)%, respectively. The difference between the two groups was significant at 7 and 14 days (P <0.05). ③The protein level of vascular endothelial growth factor in the doxycycline group after corneal transplantation was lower than that in the saline control group (P <0.01). At 3, 7 and 14 days after corneal transplantation, the levels of VEGF protein in the saline control group were (541.00 ± 75.44) and (960.00 ± 90.14) and (976.00 ± 130.41) ng / g, respectively, and those in the doxycycline group were (115.33 ± 9.29) ), (239.00 ± 41.62) and (361.00 ± 65.20) ng / g, respectively. There was significant difference between each time period (P <0.05). Conclusion: Doxycycline can inhibit corneal neovascularization after corneal transplantation and prolong the survival time of the implants.