康柏西普对兔眼准分子激光角膜切削术术后角膜上皮下雾状混浊的影响

Effects of conbercept on haze formation after photorefractive keratectomy in rabbit eyes

目的探讨康柏西普(conbercept)对兔眼准分子激光角膜切削术术后角膜上皮下雾状混浊(haze)的影响。方法采用随机数字表法将64只纯种新西兰大白兔分为单纯手术组、生理盐水组、conbercept0.5mg组和conbercept1.0mg组,每组16只,均行右眼准分子激光角膜切削术(PRK),术后单纯手术组不做任何处理,其余3个组分别给予术眼结膜下注射生理盐水0.05ml、conbercept0.5mg(0.05ml)和conbercept1.0mg(0.10ml),另选取8只兔作为正常对照组。术后裂隙灯显微镜下观察角膜上皮愈合情况及haze程度并进行Fantes分级,各手术组术后1周和4周分别空气栓塞法处死8只,正常对照组处死4只,取其角膜组织行苏木精-伊红染色,并通过免疫组织化学法检测角膜组织中转化生长因子-β1(TGF-β1)、α-平滑肌肌动蛋白(α-SMA)及基质金属蛋白酶-2(MMP-2)的表达。结果术后兔眼角膜上皮均于3~5d完全愈合,各组兔眼术后角膜上皮愈合时间比较,差异无统计学意义(F=0.37,P=0.77)。在观察期间,各手术组haze均于术后4周时最明显,conbercept1.0mg组haze最严重,其次为单纯手术组和生理盐水组,conbercept0.5mg组haze较其他手术组明显减轻,各组兔眼不同时间点haze分级比较,差异均有统计学意义(F组别=20.114,P=0.000;F时间=8.084,P=0.006)。苏木精-伊红染色示PRK术后1周,术区角膜上皮细胞增生,细胞排列紊乱、形态各异,浅基质层成纤维细胞数量增多,胶原纤维排列紊乱。角膜上皮及浅基质层的增生程度由强到弱依次为conbercept1.0mg组、单纯手术组和生理盐水组、conbercept0.5mg组。术后4周各手术组术区角膜组织结构较术前1周紊乱,各组增生程度次序同术前1周。免疫组织化学法检测显示,术后1周conbercept1.0mg组术区角膜组织中TGF-β1、α-SMA和MMP-2的表达最强,其次为单纯手术组和生理盐水组,conbercept0.5mg组次之,各手术组3种因子的表达较正常对照组明显增强,差异均有统计学意义(均P<0.05)。结论Conbercept适量结膜下注射可抑制haze的形成,抑制角膜上皮和浅基质层的增生,过量则促进haze的形成,促进角膜上皮和浅基质层的增生。

Objective To investigate the effect of conbercept on rabbit's haze after photorefractive keratectomy(PRK).Methods Sixty-four pedigree New Zealand white rabbits were randomly divided into PRK group,normal saline solution group,conbercept 0.5 mg group,conbercept 1.0 mg group,with 16 rabbits in each group.PRK was performed on right eyes,PRK group only received PRK,the other three surgery groups were given postoperative subconjunctival injection of 0.05 ml normal saline solution,0.5 mg(0.05 ml)conbercept and 1.0 mg(0.10 ml)conbercept,respectively.In addition,another 8 rabbits were randomly chosen as normal control group.The healing of postoperative corneal epithelial was observed by slit lamp biomicroscope,and the degrees of haze were graded based on Fantes.Eight rabbits in the surgery groups and 4 rabbits in the normal control group were killed in the first week and the fourth week.The corneal tissue was stained by hematoxylin-eosin,and the expressions of transforming growth factor-β1(TGF-β1),α-smooth muscle actin(α-SMA)and matrix metalloproteinase-2(MMP-2)were detected by immunochemistry.The use and feeding of experimental animals followed the Relevant Regulations of the Animal Management Committee of Binzhou Medical University.This study protocol was approved by Ethic Committee of the Affiliated Hospital of Binzhou Medical University(No.201701-08).Results Corneal epithelium of all operative rabbits healed completely at 3-5 days and no significant difference in healing time between the groups after operation(F=0.37,P=0.77).The degree of haze in each surgery group reached the highest value at about 4 weeks after operation,and haze in the conbercept 1.0 mg group was the most serious,followed by PRK group and normal saline solution group,haze in conbercept 0.5 mg group was significantly alleviated(F group=20.114,P=0.000;F time=8.084,P=0.006).Hematoxylin-eosin staining showed that corneal epithelial cells and fibroblasts in superficial stroma proliferated in one week after PRK,which lead to the disorder of cells and collagen fibers,and the extent of hyperplasia was the same as that of haze.Immunohistochemistry showed that at one week after operation,the expressions of factors in PRK group and normal saline solution group were apparently lower than that of conbercept 1.0 mg group,but were apparently higher than that of conbercept 0.5 mg group,and the expressions of the factors were the weakest in normal control group,with significant differences between them(all at P<0.05).Conclusions Subconjunctival injection of appropriate conbercept can inhibit the formation of haze and the proliferation of corneal epithelium and superficial stroma,but overdose of conbercept leads to opposite effects.