准分子激光角膜切削术后转化生长因子β及Ⅰ,Ⅲ型胶原 …

目的 观察准分子角膜切削术术后,猴角膜愈合过程中角膜转化生长因子－β的表达,以及Ⅰ、Ⅲ型胶原的表达情况,探讨ＴＧＦ－β与ＰＲＫ术后角膜愈合过程中Ⅰ、Ⅲ开明交原合成的关系。方法 对３只（６只眼）恒河猴行ＰＲＫ治疗。输入－１０．００Ｄ治疗程序,切削深度为１０３μｍ,分别于术后１、３及６个月行光镜、电镜及免疫组化检查。结果 术后１个月,ＴＧＦ－β在角膜上皮层染以阳性染色,上皮下也可见少量成纤维细胞染色阳     

准分子激光角膜切削术后角膜创面的愈合及皮质类固醇对愈合的影响

目的研究准分子激光角膜切削术（ｐｈｏｔｏｒｅｆｒａｃｔｉｖｅｋｅｒａｔｅｃｔｏｍｙ，ＰＲＫ）后创面愈合过程及皮质类固醇对愈合的影响。方法６只猕猴双眼行ＰＲＫ后，右眼滴皮质类固醇激素眼液，左眼滴新霉素眼液。于术后４天、１、２周、１、３及６个月测量角膜厚度，并取角膜做免疫组化染色，检测Ⅲ、Ⅳ、Ⅶ型胶原、纤维连结蛋白和层粘连蛋白。结果术后１个月角膜Ⅶ型胶原的分布恢复正常状态；应用皮质类固醇可降低Ⅲ型胶原和纤维连结蛋白的沉积，且术后中央角膜厚度的变化较对照组更接近期望值。结论ＰＲＫ后角膜上皮细胞与基质可达到紧密的附着；皮质类固醇对减少术后角膜雾状混浊和屈光回退有一定作用。     

羊膜对激光角膜切除术后TGF—β1表达调控的初步研究

目的 探讨羊膜对激光角膜光学切除术（Ｐｈｏｔｏｒｅｆｒａｃｔｉｖｅ ｋｅｒａｔｅｃｔｏｍｙ ＰＲＫ）后角膜组织ＴＧＦ－β１表达的影响。方法 对６只新西兰白兔双眼行ＰＲＫ,术后１眼立即行角膜表面羊膜移植术,只眼作为对照。术后４周用原位杂交方法检测角膜上皮和基质ＴＧＦ－β１ｍＲＮＡ的表达。结果 ＰＲＫ后４周对照组角膜上皮和基质有较浓度的ＴＧＦ－β１ｍＲＮＡ表达,羊膜移植组角膜上皮和基质亦有ＴＧＦ－β１     

准分子激光屈光性角膜切削术后角膜地形图分析

目的分析准分子激光屈光性角膜切削术（ｐｈｏｔｏｒｅｆｒａｃｔｉｖｅｋｅｒａｔｅｃｔｏｍｙ,ＰＲＫ）术后角膜切削区的形态、偏心情况和屈光的稳定性。方法对３１２例（３６６只眼）ＰＲＫ手术患者进行术后１、３和６个月的角膜地形图检查。结果术后１个月切削区中心偏离瞳孔中心的距离为０．２６６ｍｍ,双眼平均偏离瞳孔中心的方向均为鼻上侧。切削区形态平滑型占４９．５％,半环型、钥匙洞型、肾型和哑铃型占４２．９％,中心岛型占６．０％。中心岛型对术后最佳矫正视力影响较大。术后１～３个月角膜屈折力变化较大,高度近视比低度近视回退明显。结论提示ＰＲＫ术中瞄准中心问题非常重要,直接影响术后的效果,同时也应长期随访角膜地形图,进一步观察术后的稳定性。     

PRK术后角膜基质内Ⅰ型和Ⅲ型胶原的mRNA表达

目的：从分子水平探讨ＰＲＫ术后前基质内Ⅰ型和Ⅲ型胶原的，ＲＮＡ表达情况。方法：２４只新西兰白兔，双眼行－５．０Ｄ近视治疗。于术后３天、１、２、３、４周取下角膜，用原位杂交方法观察Ⅰ型和Ⅲ型胶原的基因表达情况。结果：所有眼Ⅰ型胶原的ｍＲＮＡ表达均为阳性。术后２周时，Ⅲ型胶原ｍＲＮＡ表达阳性，到４周时表达较明显。讨论：ＰＲＫ术后上皮下混浊与新生胶原的形成有关，且Ⅲ型胶原是新生胶原的主要成分。     

准分子激光角膜切削术后角膜伤口的愈合机制和几种生长因子的调节作用

准分子激光角膜切削术（ＰＲＫ）术后角膜伤口的愈合是一复杂的病理生理过程。受多种因素的调节和影响，一些生长因子在ＰＲＫ术后角膜薄雾的形成过程中起关键作用。本文重点对上皮生长因子（ＥＧＦ）、转化生长因子α（ＴＧＦ－α）、转化生长因子β（ＴＧＦ－β）、蛋白水解酶等特性加以综述，并探讨其在角膜愈合过程中的作用。     

准分子激光角膜切削术后角膜伤口的愈合机制和几种生…

准分子激光角膜切削术（ＰＲＫ）术后角膜伤口的愈合是一复杂的病理生理过程。受多种因素的调节和影响，一些生长因子在ＰＲＫ术后角膜薄雾的形成过程中起关键作用。本重点对上皮生长因子（ＥＧＦ）、转化生长因子α （ＴＧＦ－α）、转化生长因子β（ＴＣＦ－β）、蛋白水解酶等特性加以综述，并探讨其在角膜愈合过程中的作用。     

激光角膜光学切除术后兔角膜上皮细胞凋亡的研究

目的 探讨激光角膜光学切除术后（ｐｈｏｔｏｒｅｆｒａｃｔｉｖｅ ｋｅｒａｔｅｃｔｏｍｙ,ＰＲＫ）伤口愈合过程中角膜上皮细胞凋亡情况。方法 对６只兔双眼分别行Ｐ．Ｒ,激光切削参数为一８．０Ｄ、１５６μｍ深,消隔直径５．６ｍｍ。将兔分别于ＰＲＫ后１ｍｏ、３ｍｏ处死,取下角膜进行冰冻切片,用的位标记检测法（ＴＵＮＥＬ法）分别检测角膜上皮细胞凋亡情况。结果 正常兔角膜可见上皮上皮浅层有少量细胞亡,ＰＲＫ后     

准分子激光屈光性角膜切削术角膜神经损伤及再生的形态学研究

２２只新西兰白兔（４０眼）行－５．５０Ｄ，５．５ｍｍ直径的近视ＰＲＫ，术后不同时期取角膜进行氯化金染色以观察角膜神经的损伤及再生。ＰＲＫ术后切削区内上皮下神经丛被切除，术后１周可见神经纤维再生，２个月时新生神经纤维密度高于正常，术后３—４个月其密度达高峰，到术后６个月时角膜中央区神经再生过程尚未全部完成。结果表明，ＰＲＫ可以损伤切削区角膜神经，其再生在术后早期即可发生，但神经的修复还需一个较长的时间过程。     

影响准分子激光屈光性角膜切削术后眼压的因素

目的分析影响准分子激光屈光性角膜切削术（ｅｘｃｉｍｅｒｌａｓｅｒｐｈｏｔｏｒｅｆｒａｃｔｉｖｅｋｅｒａｔｅｃｔｏｍｙ,ＰＲＫ）术后眼压的因素。方法采用非接触式眼压计（ｎｏｎｃｏｎｔａｃｔｔｏｎｏｍｅｔｒｙ,ＮＣＴ）测量眼压,对ＰＲＫ前、后随访半年以上８６例（１５０只眼）患者眼压差与角膜切削厚度、术前术后角膜曲率差之间进行多元回归分析。结果术前眼压明显高于ＰＲＫ术后１周、３及６个月的眼压,差异有非常显著性（ｔ检验,Ｐ＜０．０１）,与术后１个月时眼压比差异无显著性（Ｐ＞０．０５）。术后１个月时的眼压高于术后其他时间眼压（Ｐ＜０．０１）。ＰＲＫ后眼压降低与角膜厚度减少及角膜前表面曲率的降低有关（ｒ＝０．３６１,Ｐ＜０．０１;ｒ＝０．１８８,Ｐ＜０．０５）,建立二元回归方程如下：Ｙ＝－０．０５９－０．０３８Ｘ１＋０．００９Ｘ２。Ｙ：术前术后眼压差（ｋＰａ）,Ｘ１：术前术后角膜曲率差（Ｄ）,Ｘ２：角膜切削厚度（μｍ）。结论ＰＲＫ后ＮＣＴ测量眼压低于术前,术后眼压与氟甲脱氧泼尼松龙（ｆｌｕｏｒｏｍｅｔｈｏｌｏｎｅ）的用药次数和时间、角膜切削厚度、角膜曲率有关。     

The immunohistochemical changes of TGF-beta, type I and III collagen in corneal healing after photorefractive keratectomy

OBJECTIVE: To observe the immunohistochemical expression of transforming growth factor-beta (TGF-beta), type I and III collagen in monkey corneal healing after photorefractive keratectomy (PRK) and find whether TGF-beta is involved in the process of corneal healing and correlated with the syntheses of type I and III collagen. METHODS: Three rhesus monkeys (6 eyes) were operated for myopic ablation of 10.0 diopters. The depth of ablation was 103 microm. Microscopy, transmission electron microscopy and immunohistochemistry were performed at 1, 3 and 6 months after PRK. RESULTS: TGF-beta staining was positive in the epithelium and a few fibroblasts under the epithelium at one month after the operation. The staining was negative at 6 months and in the control specimens. Postoperatively, type I and III collagen stainings were evident at 1, 3 and 6 months. Type I collagen staining was positive and type III was negative in the controls. CONCLUSION: TGF-beta is involved in the corneal healing after PRK, and perhaps correlated with the syntheses of type I and III collagen.     

准分子激光角膜切削术与准分子激光原位角膜磨镶术角膜创口愈合的对比实验研究

比较准分子激光角膜切削术(photorefractive keratectomy,PRK)与准分子激光原位角膜磨镶术(1aser in situ keratomileusis,LASIK)后激光对角膜组织的切削效应及角膜的愈合情况,从组织学角度探讨角膜雾状混浊(Haze)及屈光度数回退的成因。方法24只新西兰白兔按预矫屈光度数随机分为-4.00 D组和-8.00 D组,每只兔右眼行PRK,左眼行LASIK。术后10d,1、3及6个月观察Haze情况并验光,每组随机处死3只兔取角膜行光镜、电镜及免疫组化检查,检测胶原Ⅲ、胶原Ⅳ、纤维连结蛋白(fibronectine,FN)及转化生长因子-β(transforming growth factor-β1,TGF-β2)的含量。结果行PRK术的右眼术后出现不同程度的Haze及屈光度数回退,其程度与预矫正屈光度数成正比。行LASIK术的左眼术后除少数角膜瓣周围半环形混浊外,手术区域角膜透明,屈光度数回退较右眼轻。-4.00 D组右眼与左眼术后屈光度数均稳定,-8.00 D组右眼较左眼屈光度数回退明显。右眼术后角膜愈合反应重,恢复慢。6个月时角膜基质仍处于修复阶段。左眼术后除形成角膜上皮栓及对应处基质轻度增生外,手术区域角膜瓣与基质床间界面清晰,无明显增生,角膜基质愈合反应轻、恢复快。术后所有兔眼角膜均有TGF-β1表达及活化,持续时间与角膜愈合时间一致。Haze及屈光度数回退组织学改变为：角膜上皮细胞增生,基底膜不成熟,前基质角膜细胞活化、增殖,新生胶原Ⅲ合成、排列紊乱,细胞外基质FN在角膜上皮下沉积。结论LASIK矫正近视尤其是高度近视优于PRK;角膜伤口愈合特别是基质愈合的反应程度,是。Haze及屈光度数回退的关键;TGF-β1是角膜愈合过程中重要调节因子,可通过介导角膜上皮一基质相互作用,调节胶原Ⅲ及FN的含量,参与瘢痕形成。     

Expression of cyclooxygenase-2 in corneal cells after photorefractive keratectomy and laser in situ keratomileusis in rabbits

PURPOSE: To compare the expression pattern of cyclooxygenase-2 (COX-2) in rabbit corneal cells after photorefractive keratectomy (PRK) and laser in situ keratomileusis (LASIK) with the same refractive correction. SETTING: Department of Ophthalmology, Wakayama Medical University, Wakayama, Japan. METHODS: Thirty adult albino rabbits were used in the study. Photorefractive keratectomy or LASIK was performed in 1 eye of each animal for the same refractive correction. Each animal was killed after healing intervals up to 6 months. Paraffin sections of the cornea were processed for immunohistochemistry for COX-2 and NFkappaB (p65). RESULTS: After PRK, the central and peripheral corneal epithelia up-regulated COX-2 at 3 days; the central epithelium was positive at 4 weeks. Central and peripheral epithelia returned to negative 3 months later. After LASIK, the central epithelium on the corneal flap up-regulated COX-2 at 1 and 2 weeks; it returned to negative at 4 weeks. The peripheral epithelium was labeled with the antibody. Keratocytes around the stromal incision between the flap and the stromal bed up-regulated COX-2 and returned to negative at 3 months. COX-1 was not detected immunohistochemically in corneal tissue during the healing intervals after both procedures. Nuclear factor kappaB was detected in the cytoplasm and nuclei of migrating corneal epithelial cells 1 day after PRK, was positive in the cytoplasm at 3 days and negative in cytoplasm and nuclei at week and later. CONCLUSIONS: Migrating injured epithelium expressed COX-2 until week 4 during post-PRK healing. Central uninjured epithelium as well as stromal keratocytes expressed COX-2 from 3 days to 2 weeks after LASIK. Uninjured peripheral epithelium also expressed COX-2 at 4 weeks. Activation of stromal keratocytes may induce expression of COX-2 in overlying uninjured epithelium via the inflammatory cytokine(s)/NFkappaB pathway.     

准分子激光术后大鼠角膜上皮下雾状混浊中Ⅰ型前胶原的测定

目的探讨准分子激光术后大鼠角膜中Ⅰ型前胶原的变化,比较角膜非手术区、手术野透明区与上皮下雾状混浊（haze）区的Ⅰ型前胶原含量的差异。方法32只6月龄清洁级SD大鼠按照手术前后被处死的时间不同平均分为8组。28只大鼠的一侧眼被施行准分子激光角膜切削术（PRK）。术后每日裂隙灯显微镜下观察术眼角膜情况,角膜haze的分级参照Seiler标准。分别于手术前和术后1、3、7、15d及1、2、3个月处死动物,行苏木精-伊红染色,观察手术区角膜的形态变化,采用免疫组织化学法检测角膜Ⅰ型前胶原在术眼角膜组织中的表达,并以光密度（OD）值表示。对不同时间采集的角膜非手术区、手术野透明区与haze区中Ⅰ型前胶原含量进行定量分析。结果臼术后3d起,术眼角膜上皮开始移行覆盖切削区并逐渐增厚,基质纤维细胞增生活跃;术后1个月时术眼角膜上皮基底膜开始形成,基质胶原纤维排列紊乱;术后3个月角膜基质胶原重塑过程基本完成,角膜结构趋于正常。大鼠角膜术后haze分级与Ⅰ型前胶原含量呈正相关（r=0．406,P〈0．05）;非手术区各时间段角膜平均OD值均无明显变化（t=5．719,P〉0．05）;手术区角膜中的Ⅰ型前胶原含量明显高于非手术区（P〈0．05）;haze区平均OD值明显高于非haze区（t=5．578,P〈0．05）。免疫组织化学检测表明,角膜上皮中Ⅰ型前胶原的阳性表达在术后7d最多,并终止于术后1个月,而角膜细胞间质中Ⅰ型前胶原持续至整个修复过程。结论Ⅰ型前胶原含量的增加可能是haze形成的主要因素之一,为进一步研究准分子激光术后角膜前胶原的变化提供理论支持。     

兔LASEK与PRK术后角膜Ⅰ、Ⅲ、Ⅴ、Ⅵ型胶原的Western Blot研究

目的:用WesternBlot法比较准分子激光上皮下角膜磨削术(LASEK)与准分子激光角膜切削术(PRK)术后角膜Ⅰ、Ⅲ、Ⅴ、Ⅵ型胶原的动态变化情况。方法:52只新西兰白兔分为8组,对每只兔右眼行LASEK,左眼行PRK。术后1d、7d、1个月、3个月、4个月、5个月、6个月观察Haze情况,处死动物取角膜行Ⅰ、Ⅲ、Ⅴ、Ⅵ型胶原的动态变化过程的免疫组织化学检查与WesternBlot检测。结果:经免疫组化和WesternBlot研究发现:LASEK组术后角膜基质中Ⅰ、Ⅲ型胶原开始增生的时间早于PRK组,表达的强度与PRK组有显著差异;两组的Ⅴ、Ⅵ型胶原动态变化曲线相似,达到表达高峰的时间一致,PRK组Ⅴ、Ⅵ型胶原的表达均明显高于LASEK组,术后6个月时LASEK组表达显著弱于PRK组。结论:PRK手术后角膜基质中Ⅰ、Ⅲ、Ⅴ、Ⅵ型胶原的表达强度、达到高峰及恢复正常的时间与LASEK相比存在显著差异,表明PRK术后基质内有胶原的过量沉积,可能是临床PRK术后Haze严重及屈光回退的组织学基础。WesternBlot法是一种可比较准确地半定量检测胶原含量动态变化的研究方法。     

Intact corneal epithelium is essential for the prevention of stromal haze after laser assisted in situ keratomileusis

AIMS: To determine the effect of intact corneal epithelium on stromal haze and myofibroblast cell formation after excimer laser surgery. METHODS: Denuded epithelium alone, photorefractive keratectomy (PRK), laser in situ keratomileusis (LASIK), or LASIK with denuded epithelium was performed in rabbit eyes. Postoperative anterior stromal haze was assessed employing a standard scale. Immunohistochemical methods were used to detect alpha smooth muscle actin (alpha-SMA), a marker for myofibroblastic cells, and type III collagen in subepithelial corneal tissue. RESULTS: Three weeks after surgery, the presence of alpha-SMA positive long extended and spindle-shaped stromal cells, and synthesis of type III collagen were observed in the subepithelial stromal layer corresponding to corneal haze in PRK and LASIK with denuded epithelium, but not in denuded epithelium alone and LASIK. CONCLUSION: The intact corneal epithelium may play an important part curbing subepithelial haze and differentiation of myofibroblasts in corneal wound healing.     

Wound healing in rabbit corneas after photorefractive keratectomy and laser in situ keratomileusis

PURPOSE: To compare the wound-healing process in the rabbit cornea after photorefractive keratectomy (PRK) and laser in situ keratomileusis (LASIK) with the same refractive correction. SETTING: Department of Ophthalmology, Wakayama Medical University, Wakayama, Japan. METHODS: Adult albino rabbits (N = 24) were used. One eye of each animal had PRK or LASIK with the same refractive correction. Each animal was killed after an interval of up to 6 months. The expression pattern of corneal stromal injury-related molecules with the 2 treatments were compared. Paraffin sections of the cornea were processed immunohistochemically for alpha-smooth muscle actin (alpha-SMA), collagen type IV [alpha1(IV)](2),alpha2(IV), and heat shock protein (HSP) 47 as well as other HSPs. Sections were also examined after hematoxylin and eosin or periodic acid-Schiff staining. RESULTS: Hematoxylin and eosin staining showed the central epithelium to be thick in PRK-treated corneas. The thick epithelium was restricted to the area around the corneal flap edge adhesion in LASIK-treated corneas at 3 months. Periodic acid-Schiff staining showed an absence of or interruption in the epithelial basement membrane in PRK-treated corneas for up to 6 months. Heat shock protein 47 was detected in keratocytes on day 3 but not after that in PRK-treated corneas. There was no difference in the expression of other HSPs. Alpha-smooth muscle actin was expressed in keratocytes repopulated in the central anterior cornea of PRK-treated corneas at 28 days. Keratocytes with immunoreactivity for these 2 proteins were not seen in LASIK-treated corneas. Collagen IV [alpha1(IV)](2),alpha2(IV) was not detected in either group of corneas. The central epithelium became transiently thicker in PRK-treated corneas. CONCLUSION: Keratocyte responses to laser stromal ablation were more marked in corneas treated with PRK than in those treated with LASIK.     

羊膜移植对家兔PRK术后伤口愈合作用的研究

目的：观察羊膜对家兔PRK术后角膜伤口愈合反应的影响。方法：对30只家兔常规施行双眼PRK手术（－8．0D切削程序），即行羊膜移植，观察术后早期角膜切削区炎性细胞的浸润和角膜上皮愈合情况，晚期角膜成纤维细胞增生和上皮增生情况，观察haze并评分；早期测量角膜SOD活力和MDA含量，对结果进行统计学分析。结果：早期移植相炎性细胞的数目比对照组少，上皮愈合时间较短，晚期上皮增生较轻，晚期移植组成纤维细胞数目较少；移植组haze评分较低；早期移植组自由基反应产物MDA的含量较低，而SOD含量较高，差异均具有显著性。结论：羊膜能促进术后上皮愈合，减轻炎性反应和自由基反应，减少成纤维细胞增生，减轻haze的形成。     

In vivo evaluation of corneal structure changes after refractive procedures

PURPOSE: The aim of the study was to evaluate and compare in vivo the corneal structure changes after refractive procedures (PRK, LASIK, LASEK). MATERIAL AND METHOD: The analysed group of patients consisted of 226 corneas in 126 patients, who underwent correction of myopia and myopic astigmatism, using the procedures: PRK (120 eyes), LASIK (56 eyes) and LASEK (50 eyes). The photoablation of the corneas was performed with the Excimer Laser MEL 60 and MEL 70 G-Scan Aesculap Meditec. Postoperative observations were made using a confocal microscopes Confoscan P4 (Tomey) and ConoScan 2 (Fortune Technologies). The evaluations were performed in the early (up to 3 months) and late postoperative period (after PRK and LASIK-up to 2 years; after LASEK-up to 6 months). RESULTS: The confocal microscopy revealed some changes within the corneal epithelium and anterior part of stroma after PRK, LASIK and LASEK. After PRK, there was increased desquamation of superficial epithelial cells in early postoperative period. These cells were elongated after LASIK and LASEK procedures. After PRK and LASEK, the Bowman's membrane was absent in the central part of the cornea, during the whole observation period. After all these procedures, the anterior part of the corneal stroma in the ablation zone, showed increased background illumination of collagen fibres and an irregular pattern of elongated keratocytic nuclei, in the early postoperative period. No scar tissue--"haze" was found in cases after LASEK, what may occur after PRK procedure. The findings kept changing in the course of the follow up time. CONCLUSIONS: Confocal microscopy enables in vivo monitoring of changes, which occur in the corneal structure after refractive procedures; this facilitates the evaluation of corneal healing. LASEK is the least invasive refractive procedure, which allows prompt stabilization of the corneal structure.     

Corneal opacity after repeated photorefractive keratectomy

Corneal opacity developed in an eye that had photorefractive keratectomy (PRK) with a 193 nm excimer laser 5 times over 3 years. Six months after the last PRK, a partial penetrating keratoplasty was performed. The cornea was stained and immunohistochemically evaluated for collagen types. Light microscopy showed thickening of epithelial layers, proliferation of subepithelial fibroblasts, and the absence of Bowman's membrane. Transmission electron microscopy showed irregular collagen lamellae and electron-dense deposits adjacent to keratocytes. The staining was positive for Alcian blue, and immunohistochemistry was positive for type IV and VI collagen. This case suggests that corneal opacity after repeated PRK is the result of deposits of type IV and VI collagen and acidic mucoprotein in the extracellular matrix.