角膜交联对真菌性角膜溃疡的治疗作用

Management of fungal corneal ulcer by corneal collagen cross-linking technique

背景 真菌性角膜溃疡是一种严重威胁视力的炎症性疾病,严重者需行角膜移植术或眼球摘除术.角膜交联术（CXL）是近年来治疗一些角膜疾病的有效方法,但其用于真菌性角膜溃疡的疗效方面少有研究.目的 观察CXL对真菌性角膜溃疡的治疗作用和效果.方法 选取8周龄健康新西兰白兔15只,其中5只作为正常对照组,另取10只兔刮除右侧角膜上皮行角膜划痕,涂抹镰刀菌液,然后行异种脱细胞角膜片覆盖,制备真菌性角膜溃疡动物模型.按照随机数字表法将模型兔随机分为非治疗组和CXL治疗组,每日行裂隙灯显微镜检查,测量角膜病灶的直径,并观察角膜水肿和炎性细胞浸润情况.于CXL治疗后第3、7、14、21、28天分别对非治疗组和CXL治疗组兔眼行眼前节照相和激光扫描共焦显微镜检查.治疗后4周,收集15只实验兔角膜组织,于扫描电子显微镜下检测正常对照组、非治疗组和CXL治疗组角膜胶原纤维的超微结构.结果 实验兔右眼造模后3d即可见角膜病灶区灰白色溃疡灶,激光扫描共焦显微镜下浅基质层局部见豆荚样菌丝.造模后l周角膜溃疡灶加深,范围扩大,激光扫描共焦显微镜下浅基质层见大量真菌孢子和短棒样菌丝,可见角膜内皮细胞层有炎性细胞及前房内渗出.CXL治疗后3、7、14、21 d,CXL治疗组兔角膜上皮缺失范围均小于非治疗组,差异均有统计学意义（P＜0.05）.治疗后28 d,正常对照组、非治疗组和CXL治疗组兔角膜胶原纤维束平均直径分别为（24.6±1.8）、（24.9±1.9）和（43.0±7.4）nm,3个组间差异有统计学意义（F=27.05,P=0.00）,其中CXL治疗组胶原纤维直径较非治疗组和正常对照组增粗,差异均有统计学意义（t =-5.30、5.40,P＜0.05）,胶原纤维间见较多成纤维细胞;而非治疗组兔角膜胶原纤维直径与正常对照组间差异无统计学意义（t=0.25,P＞0.05）,正常对照组胶原纤维间少见成纤维细胞.结论 CXL治疗镰刀菌性角膜溃疡疗效显著,安全性好.CXL可使角膜胶原增粗并刺激成纤维细胞增生,抑制真菌生长和炎症反应,加速角膜修复.

Background Fungal corneal ulcer is a visual-threatening eye disease,and drug therapy has a limiting efficacy.Corneal transplantation or eye enucleation sometimes is necessary to the severe patients.Corneal collagen cross-linking （CXL） is an effective method for some corneal diseases,but the study on CXL for fungal corneal ulcer is lack.Objective This study was to evaluate the clinical effectiveness and safety CXL for fungal corneal ulcer.Methods Fifteen 8-week-old healthy New Zealand white rabbits were used in this study and other 5 rabbits served as normal controls.Fungal corneal ulcer models were established in the right eyes of other 10 rabbits by infecting sickle bacteria liquid after corneal scratching and removing corneal epithelium,then decellularized ostrich corneal patch covered the defected cornea.The models were randomly divided into the non-treatment group and the CXL treatment group.Corneal lesions were examined under the slit lamp microscope every day,and cornea was pictured by laser scanning confocal microscope on the 3rd,7th,14th,21st and 28th day individually after CXL.All rabbits were sacrificed and corneal tissues were obtained 4 weeks after treatment,and the collagen fiber diameter and fibrocytes were observed under the scanning electron microscope.Results Fungal corneal ulcer models were successfully established by corneal scratching and decellularized ostrich cornea covering.The gray ulcer lesions and hypbae like bean pod were seen by slit lamp microscope and laser scanning confocal microscope 3 days after modeling.Corneal ulcer deepened and expanded 1 week later,and there were a large number of spore and hyphae criss-crossing as short rod in shallow stroma.Inflammatory cells were observed in corneal endothelial cells and ocular anterior chamber.In the CXL treatment group,the range of corneal epithelial deficiency was less than that in the nontreatment group on the 3rd,7th,14th,and 21st （all at P〈 0.05）.The diameters of collagen fibers were （24.6± 1.8） nm,（24.9 ± 1.9） nm and （43.0 ± 7.4） nm in the normal control group,non-treatment group and CXL treatment group,showing a significant difference among the 3 groups （F =27.05,P =0.00）,and the collagen diameters were thicker in the CXL treatment group than those in the normal control group and non-treatment group （t =5.40,-5.30,both at P〈0.05）,and fibrocytes were seen among the collagen fibers.No significant difference was found in the collagen diameters between the non-treatment group and normal control group,and the fibrocytes were less in the non-treatment group.Conclusions CXL therapy can treat fungal corneal ulcer by enhancing collagen,promoting fibrocytes proliferation,suppressing fungus and inflammatory response and accelerating tissue repair.