整合素α5及其配体在大鼠角膜创伤愈合中表达的研究

目的 观察创伤愈合中大鼠角膜整合素（integrin)α5及其配体－纤维连接蛋白（fibronectin,FN）表达的变化,探讨二者对角膜创伤愈合的影响。方法 大白鼠27只,随机分实验组和对照组。依左右眼手术不同又分左眼和右眼组。右眼刮除中央3mm直径区域内角膜上皮,保持前弹力膜完整;左眼同上操作后,用RK刀作长3mm,深100μm的非穿透切口。术后即刻、3、6、12、24、72h、1w,2w分别处死每组大鼠,取双眼角膜,用免疫荧光染色法观察角膜整合素α5及FN分布的改变。对照组不手术,处死后角膜标本按实验组同样步骤处理。结果 与对照组相比,术后3h双眼角膜创缘上皮表层及左眼基质切口处出现α5和FN荧光、之后荧光亮度升高,72h后下降,术后2w恢复至术前水平。α5与FN的表达相对应。结论 角膜创伤愈合早期整合素α5及其配体FN增多并相协调。二者能促进角膜创伤愈合早期的上皮迁移。     

核素示踪技术在角膜创伤愈合研究中的应用

角膜移植术后,植片上皮细胞最终全部被受体上皮细胞取代已成定论。但是,其他角膜成份的命运究竟如何?供体细胞究竟能否在受体中存活?受体细胞最终能否取代供体细胞?这一直是角膜病医生争论的同题,也是提出把核素示踪技术用于角膜创伤愈合研究的根据。一、核素示踪技术用于角膜创伤愈合研究的历史 1957年Dohlman首先用核素示踪技术对角膜移植术后植片中的细胞成份和非细胞成份进行了研究。Dohlman用的同位素是35S和32P。 1959年,Harding等首先使用氚标记的胸腺嘧啶     

体外培养人角膜基质细胞β1整合素的表达

目的探讨β1整合素在角膜创伤愈合中的作用。方法采用免疫荧光细胞计数仪检测法。结果体外培养的人角膜基质细胞β1整合素表达的阳性率平均为95．7％，阴性表达率为4．3％。结论在角膜创伤愈合中，角膜基质细胞β1整合素的表达可能极高，从而促进角膜基质细胞与细胞外基质的粘附。     

整合素-α5及其配体在大鼠角膜创伤愈合中的表达

整合素属细胞黏附分子，是由α和β亚单位通过非共价键形式结合成为跨膜糖蛋白，主要介导细胞与细胞和细胞与细胞外基质间的黏附，并在角膜创伤愈合中起重要作用。α5主要与β1结合。文献报道，配体纤维连接蛋白(fibronectin，FN)是以二聚体形式存在的大分子糖蛋白，经典受体为整合素-α5，大鼠角膜移植术后角膜创缘上皮细胞、迁移上皮细胞膜基底面及基质切口边缘α5及FN蛋白表达量明显增加；当创伤愈合后，其α5及FN蛋白表达消失。为此我们对大鼠角膜创伤愈合中整合素-α5及FN蛋白表达量进行检测，探讨其在角膜创伤愈合中的作用，现将结果报告如下。     

角膜缘部干细胞对角膜上皮创伤愈合的影响

利用角膜缘上皮移植，结膜移植和非手术的方法分别对角膜上皮创伤伴随膜缘损伤的兔眼进行实验研究，结果表明角膜缘上皮移植组上皮愈合时间短（平均８．５天），且无杯状细胞，无或极少量新生血管；结膜移植组，上皮愈合时间延长（平均１１．５天），含有杯状细胞和新生血管，非手术组形成角膜血管翳性混浊。而不件随角膜缘损伤仅有角膜中央皮皮创伤的兔眼愈合时间最短（平均３．５天），愈合后无新生血管及怀状细胞。结果证实角膜部     

胰岛素样生长因子-1在糖尿病角膜上皮创伤愈合中的研究进展

近年来,糖尿病角膜上皮创伤愈合越来越受到人们的重视,国内外已经对糖尿病角膜上皮创伤的病理、愈合过程及修复机制均有了比较深刻的认识。很多学者对角膜上皮愈合过程中各种因子的促进作用进行了系统的研究,其中胰岛素样生长因子-1（insulin-likegrowth factor-1,IGF-1）对糖尿病角膜上皮的创伤愈合有促进作用。在这里,我们采用综述的形式对IGF-1在糖尿病角膜上皮创伤愈合中的作用和机制进行阐述。     

角膜创伤愈合中上皮细胞的作用初探

目的 探讨角膜创伤愈合中上皮细胞与角膜基质成纤维细胞的作用关系。方法 用形态学方法观察兔角膜损伤后上皮细胞与角膜基质成纤维细胞动态变化相关现象。结果 发现基质成纤维细胞的增生活跃程度与上皮基底细胞的细胞层数和细胞大小密切相关。结论角膜上皮细胞在角膜创伤个性中起主导作用。     

小鼠角膜上皮创伤再上皮化过程基本特征的动力学研究

目的 研究小鼠角膜上皮机械性创伤再上皮化过程炎性细胞迁移、血小板聚集的动力学特征.方法 计算小鼠角膜2 mm上皮缺损创面改变;免疫荧光染色观察角膜基底细胞分裂和中性粒细胞、血小板、γδT细胞及巨噬细胞迁移特征;电子显微镜观察角膜上皮细胞形态学、基质PMNs和角膜缘血小板的变化.结果 创伤后24 h基本再上皮化,96 h基底细胞恢复正常;伤后18 h和30 h分裂细胞达峰值;PMNs迁移峰为12 h和30 h,血小板聚集峰于12 h;γδT细胞的两峰为6 h和24 h,MΦ两峰滞后为24 h和42 h.结论 角膜上皮创伤修复过程分3期.基底细胞分裂增生和PMNs、MΦ以及γδT细胞迁移呈双相式;早期PMNs迁移与血小板聚集同步.     

P38 MAPK在角膜创伤愈合过程中的作用

丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)级联是细胞内重要的信号传导系统。细胞通过这一信号传导系统将细胞外信号传递到细胞核内,介导细胞产生反应,调节细胞生长、增殖、分化和凋亡等生理过程及创伤愈合和炎症反应等病理过程。近年来发现一类新的MAPK通路—p38MAPK信号通路,对其结构和功能以及在角膜创伤愈合修复和炎症反应中的作用已有了进一步的了解,在信号通路水平调控p38MAPK的表达和活性,可能成为临床角膜创伤治疗的新途径。     

蛋白多糖及胶原与角膜创伤愈合

蛋白多糖及胶原为角膜细胞外基质的重要组成成分,在角膜创伤愈合中起重要作用,本文就其结构功能,与细胞外基质的关系,在某些疾病情况下的改变以及研究进展方面作了概述。     

糖尿病患者角膜上皮损伤愈合延迟的研究进展

糖尿病患者角膜上皮损伤后愈合延迟较常见,持续不愈合的角膜损伤可导致复发性上皮糜烂、角膜溃疡甚至角膜穿孔,影响视功能。本文就糖尿病患者角膜上皮损伤愈合延迟的病理基础、发病机制以及相关治疗的研究进展做一综述。概述了糖尿病患者角膜上皮结构和功能的改变,阐述了高糖、蛋白酶、泪膜、生长因子和细胞因子、角膜神经以及microRNA等在发病机制中的作用,总结了治疗糖尿病患者角膜上皮损伤愈合延迟的最新思路。     

P-选择素对角膜上皮创伤修复及其中性粒细胞迁移的影响

目的观察P-选择素对角膜上皮创伤修复及其中性粒细胞迁移的影响。方法选用P-选择素基因敲除小鼠与野生型小鼠进行对比,观察角膜上皮创伤修复过程,同时用免疫组织化学方法观察中性粒细胞在角膜的迁移,并用酶联免疫吸附测定方法检测角膜中细胞因子和趋化因子的变化。结果与野生型小鼠相比,P-选择素基因敲除小鼠的角膜创伤修复延迟,向角膜迁移的中性粒细胞数量减少;中性粒细胞的迁移与趋化因子和细胞因子有密切的关系。结论P-选择素表达缺陷可导致角膜上皮创伤修复的延迟。这种延迟可能与创伤后中性粒细胞向创伤区迁移数量减少有关。     

Delayed corneal wound healing following radial keratotomy

Four human corneal specimens obtained 5 to 47 months following radial keratotomy were evaluated by correlative microscopy. Repeat radial keratotomies had been performed in two cases. We documented gaping keratotomy incisions, epithelial plugs, and epithelial-lined incisions. In all cases, Bowman's layer was malapposed with slight to moderate fibroblastic activity at the incision sites. Epithelial and endothelial radial ridges were seen in two cases. No endothelial damage was seen under the radial incisions. The morphological evaluation of these specimens show the potential for a poor wound-healing response when corneas with previous corneal surgery and/or pathologic states undergo radial keratotomy.     

表皮生长因子对碱烧伤角膜伤口愈合的实验研究

目的　观察鼠表皮生长因子 (m ouse epiderm al grow th factor,m EGF)对角膜上皮创伤修复的作用 ,并探讨其量效关系。方法　采用兔角膜碱烧伤后上皮缺损模型 ,分别给予 0 .5、1、2 g· L- 1m EGF滴眼液治疗 ,生理盐水作对照组 ,每日荧光染色裂隙灯观察、记录 ,疗程 1周。结果　 2 g· L - 1m EGF治疗组与其他 3组有显著性差异 ,0 .5及 1g· L- 1m EG F与对照组之间无显著性差异。结论　尽管量效关系复杂 ,2 g· L- 1m EGF对角膜碱烧伤后上皮缺损的修复起到一定的促进作用。     

FK506与环孢霉素A对角膜损伤愈合的对比研究

目的　探讨他克莫司 (FK5 0 6 )对兔角膜上皮愈合的影响并与环孢霉素A(CsA)对比。方法　 18只新西兰白兔建立角膜上皮损伤模型 ,分 3组 ,每组 6只 ,随机分为 4只和 2只 ,均于去除角膜上皮后即照像 1次 ,4只兔右眼滴 1g·L-1FK5 0 6眼液 ,左眼滴 5g·L-1CsA眼液 ,另 2只兔眼不滴眼液。 3组分别于滴眼后 12、2 4、36、4 8、72h各照像 1次。结果　统计学检验显示FK5 0 6组与CsA组及空白组相比较均有显著性差异 (P <0 .0 5 ) ;而CSA组与空白组无显著性差异。结论　局部应用 1g·L-1FK5 0 6滴眼液能明显缩短兔角膜上皮愈合时间 ;而 5g·L-1CsA对角膜上皮愈合无影响。     

The conjunctiva in corneal epithelial wound healing

BACKGROUND/AIMS—During the healing of corneal epithelial wounds with limbal involvement, conjunctival epithelium often migrates across the denuded limbus to cover the corneal surface. It is believed that, over a period of time, conjunctival epithelium covering the cornea assumes characteristics of corneal epithelium by a process referred to as conjunctival transdifferentiation. The purpose of this study was to examine, clinically, the fate of conjunctival epithelial cells covering the cornea and to assess the healing of corneal epithelial wounds when the conjunctival epithelium was removed or actively prevented from crossing the limbus and extending onto the cornea.
METHODS—10 patients with conjunctivalisation of the cornea were followed for an average of 7.5 months. Five patients in this group had their conjunctival epithelium removed from the corneal surface and allowed to heal from the remaining intact corneal epithelium. In another four patients with corneal epithelial defects, the conjunctival epithelium was actively prevented from crossing the limbus by mechanically scraping it off.
RESULTS—The area of cornea covered by conjunctival epithelium appeared thin, irregular, attracted new vessels and was prone to recurrent erosions. Conjunctivalisation of the visual axis affected vision. Removal of conjunctival epithelium from the cornea allowed cells of corneal epithelial phenotype to cover the denuded area with alleviation of symptoms and improvement of vision. It was also established that migration of conjunctival epithelium onto corneal surface could be anticipated by close monitoring of the healing of corneal epithelial wounds, and prevented by scraping off conjunctival epithelium before it reached the limbus.
CONCLUSION—This study shows that there is little clinical evidence to support the concept that conjunctival transdifferentiation per se, occurs in humans. "Replacement" of conjunctival epithelium by corneal epithelial cells may be an important mechanism by which conjunctival "transdifferentiation" may occur. In patients with partial stem cell deficiency this approach can be a useful and effective alternative to partial limbal transplantation, as is currently practised.

Keywords: corneal epithelium; conjunctiva; stem cells; transdifferentiation     

重组人生长激素对兔角膜损伤早期修复的研究

目的：探讨重组人生长激素(rHGH)对兔角膜上皮损伤早期的修复作用。

方法：选取32只健康雄性新西兰兔制作双眼角膜上皮损伤模型,随机选取1眼滴无菌生理盐水作为对照组,另1眼滴20nmol/L重组人生长激素作为rHGH组,分别于造模前、造模后24、48、72h采用荧光素钠染色法观察角膜上皮愈合情况,采用Cochet-Bonnet角膜知觉测量仪测量检查中央角膜敏感度,同时收集各时间点泪液检测炎症因子的表达。

结果：造模后48h对照组和rHGH组角膜上皮愈合率分别为62.52%±6.73%和79.62%±10.62%(P<0.05),造模后72h分别为90.56%±9.57%和98.43%±3.65%(P<0.05)。造模后48h,对照组和rHGH组中央角膜敏感度分别为3.22±0.42、4.22±0.26cm(P<0.05)。造模后各时间点两组泪液TNF-α和IL-1α表达均较造模前增加,且对照组均高于rHGH组。造模后24、48h,两组泪液MMP-9表达均较造模前增加,且造模后48h对照组高于rHGH组(均P<0.05)。造模后24、48h两组泪液IL-21表达均较造模前增加(P<0.05)。造模后各时间点两组泪液IL-17α、Leptin表达与造模前相比及两组之间比较均无差异(P>0.05)。

结论：重组人生长激素有利于兔角膜上皮损伤早期修复。     

Comparison of cytotoxicity and wound healing effect of carboxymethylcellulose and hyaluronic acid on human corneal epithelial cells

Abstract AIM: To investigate the cytotoxic effect on human corneal epithelial cells (HCECs) and the ability to faciliate corneal epithelial wound healing of carboxymethylcellulose (CMC) and hyaluronic acid (HA). METHODS: HCECs were exposed to 0.5% CMC (Refresh plus®, Allergan, Irvine, California, USA) and 0.1% and 0.3% HA (Kynex®, Alcon, Seoul, Korea, and Hyalein mini®, Santen, Osaka, Japan) for the period of 30min, and 4, 12, and 24h. Methyl thiazolyl tetrazolium (MTT)-based calorimetric assay was performed to assess the metabolic activity of cellular proliferation and lactate dehydrogenase (LDH) leakage assay to assess the cytotoxicity. Apoptotic response was evaluated with flow cytometric analysis and fluorescence staining with Annexin V and propiodium iodide. Cellular morphology was evaluated by inverted phase-contrast light microscopy and electron microscopy. The wound widths were measured 24h after confluent HCECs were scratch wounded. RESULTS: The inhibitory effect of human corneal epithelial proliferation and cytotoxicity showed the time-dependent response but no significant effect. Apoptosis developed in flow cytometry and apoptotic cells were demonstrated in fluorescent micrograph. The damaged HCECs were detached from the bottom of the dish and showed the well-developed vacuole formations. Both CMC and HA stimulated reepithehlialization of HCECs scratched, which were more observed in CMC. CONCLUSION: CMC and HA, used in artificial tear formulation, could be utilized without any significant toxic effect on HCECs. Both significantly stimulated HCEC reepithelialization of corneal wounds.     

Genomics of corneal wound healing: a review of the literature

Corneal wound healing is a complex process: its mechanisms and the underlying genetic control are not fully understood. It involves the integrated actions of multiple growth factors, cytokines and proteases produced by epithelial cells, stromal keratocytes, inflammatory cells and lacrimal gland cells. Following an epithelial insult, multiple cytokines are released triggering a cascade of events that leads to repair the epithelial defect and remodelling of the stroma to minimize the loss of transparency and function. In this review, we examine the literature surrounding the genomics of corneal wound healing with respect to the following topics: epithelial and stromal wound healing (including inhibition); corneal neovascularisation; the role of corneal nerves in wound healing; the endothelium; the role of aquaporins and aptamers. We also examine the effect of ectasia on corneal wound healing with regard to keratoconus and following corneal surgery. A better understanding of the cellular and molecular changes that occur during repair of corneal wounds will provide the opportunity to design treatments that selectively modulate key phases of the healing process resulting in scars that more closely resemble normal corneal architecture.