角膜移植供体材料分析

本文对我中心１９９２－１９９４年９４０眼角膜移植供体材料状况及保存方法，眼库技术，供体材料的合理应用等问题进行了分析，并就上述问题提出了一些探讨性意见。     

远程投送对中期保存兔角膜内皮细胞的影响

目的:探讨模拟运输状态下机械振动对中期保存角膜内皮细胞(CEC)活性的影响.方法:DX中期保存液保存兔角膜片7d,实验室用摇床模拟远程运输环境,按作用不同时间、不同振动强度进行分组,分别进行CEC活性染色和CEC酶组织化学染色,对比不同组间模拟运输对角膜内皮的影响.结果:实验条件下观察指标并不随振动强度和时间变化,组间各项指标差异无显著性.结论:实验室模拟远程投送对中期保存兔CEC无明显影响.     

模拟远程投送对甘油长期冷冻保存兔角膜内皮细胞的影响

目的:探讨在实验室模拟远程运输状态下机械振动对改良后甘油长期保存兔角膜内皮细胞(CEC)活性的影响。方法:通过改良后的甘油保存角膜的方法,实验室中保存兔角膜片3mo,实验室用摇床模拟远程运输环境,结合实际按不同时间、不同振动强度进行分组,分别进行CEC活性染色并行实验性PKP术,对比各组间模拟运输对角膜内皮的影响。结果:实验条件下观察指标并不随振动强度和时间变化,组间各项指标差异无显著性。结论:实验室模拟远程投送对甘油长期保存兔角膜的CEC无显著性影响。     

角膜移植供体材料的现状与进展

从选材、材料改进以及临床试验等角度进行描述不同角膜移植供体材料的各自进展及特点.由于提高角膜与组织相容性,减少排斥反应和并发症是长期研究的重点领域,而材料选择、新材料的开发应用等是提高角膜移植成功率的关健.     

甘油冷冻保存角膜远程投送包装系统的温度控制

目的:研究甘油冷冻保存角膜远程投送包装系统的温度变化和保存角膜的安全性及远程投送的可行性。方法:常温下将-21℃冰盒与盛有-21℃冷冻甘油的保存瓶置于保温箱内,并用海绵填充箱体。实验组按照保存介质(甘油)体积分组,采用双通道温度记录仪,记录不同组别的瓶内瓶外温度变化曲线,观察该包装能否满足甘油冷冻保存-20℃以下的温度要求,及该温度范围的维持时间,并在各分组间进行两两比较分析。结果:常温下实验各组保存介质冷冻甘油24h温度几何均数均低于-20℃,而对照组24h温度均高于-17℃。保存介质体积≤300mL可以维持温度-20℃以下达17h以上,保存介质体积400mL可维持有效温度6h以上,各实验组与对照组比较,差异有显著性。结论:常温下该包装系统能有效控制温度于-20℃以下达20h以上,可以满足国内国际大部分地区远程投送的时间要求。     

角膜移植供体材料的现状与进展北大核心CSCD

从选材、材料改进以及临床试验等角度进行描述不同角膜移植供体材料的各自进展及特点。由于提高角膜与组织相容性,减少排斥反应和并发症是长期研究的重点领域,而材料选择、新材料的开发应用等是提高角膜移植成功率的关健。     

中期保存角膜远程投送包装系统温度控制的实验研究

目的 探讨应用于角膜中期保存的安全性及远程投送的可行性.方法 常温下,将冰点为4℃冰盒与盛有4℃中期保存液的保存瓶,一同置于保温箱内,并用海绵固定.按照保存介质(中期保存液)体积分组,采用双通道温度记录仪,记录不同组别的瓶内瓶外温度变化曲线,观察该包装能否满足中期保存>0℃,≤4℃的温度要求,及该温度范围的维持时间,并在各分组间进行两两比较分析.模拟远程运输前后观察角膜植片在包装系统中的固定情况.结果 常温下,保存介质在一定体积范围内,该远程投送系统可维持>0℃,≤4℃达20h,温度无剧烈波动,明显长于普通冰桶相关指标,差异有显著性.模拟远程投送前后角膜植片在本包装系统运输保存瓶中的固定情况良好.结论 常温条件下,该包装系统能有效控制温度满足中期保存角膜的温度要求和国内国际大部分地区投送的时间要求;并且此系统具有防震及保持植片固定特点,应用于中期保存角膜安全可靠,进行远程投送具有可行性.     

同供体双眼穿透性角膜移植术

双眼均适宜角膜移植术的角膜病变并非少见，传统多采取双眼分次手术，故为两个供体，用同一供体同时行双眼穿透性角膜移植术未见报道。近年我院收治８例，取得远期满意疗效。资料与方法一、病例选择：本组８例，男３例，女５例，年龄１６～５６岁，平均３３．５岁。例１为...     

适宜中国国情的角膜移植技术的挑战和创新

角膜盲是我国仅次于白内障的致盲性疾病,角膜供体材料匮乏造成角膜盲患者绝对数量仍在逐年上升。与欧美发达国家不同,严重的感染性角膜炎、热烧伤、化学伤和各种眼内手术引起的大泡性角膜病仍然是我国角膜移植的主要适应证。我国角膜盲的防盲面临着患者病情重、依从性差、可支付能力低和角膜供体材料严重缺乏的多重挑战。近年来,我国的角膜专科医师根据特殊的国情,发展和创新了一系列深板层移植、内皮移植、人工角膜移植和组织工程角膜异种移植的新技术,从技术层面显著提高了供体角膜材料的利用率,降低了高危移植的排斥反应。然而,由于角膜供体材料极其缺乏的瓶颈依旧,能掌握当代角膜移植新技术的角膜专科医师数量稀少,目前推动角膜捐献的立法工作和深入组织工程异种角膜材料学研究、加强国际间眼库合作迫在眉睫,适宜国情的角膜移植技术的创新和推广工作仍然任重而道远。     

Optical development of the kitten cornea

The kitten eye undergoes considerable growth during a period when there is a high neural susceptibility to the effects of visual experience. We have monitored growth of the cornea in cats of different ages using photokeratoscopy. Qualitatively, the corneal surface of the youngest kittens studied (20 days old) is sometimes irregular and photokeratographs are correspondingly partially distorted. In older kittens the general surface quality appears to be improved. Quantitatively, corneal curvature is very steep centrally in the youngest kittens and it flattens with age at a higher rate than in the pericentral region. In the adult cats, radii of curvature of central and pericentral areas are approximately equal so that a relatively large region of the cornea is spherical.     

Iontophoresis-gentamicin delivery into the rabbit cornea, using a hydrogel delivery probe

PURPOSE: To evaluate the efficacy of penetration of gentamicin into the cornea of rabbits using iontophoresis with a hydrogel-gentamicin containing probe. METHODS: Eight of 10 groups (groups 3-10) of 6 rabbits (one eye per rabbit), underwent corneal iontophoresis using soft stable hydroxyethyl methacrylate hydrogel discs (80% water content) loaded with gentamicin sulphate which were mounted on an iontophoresis probe. The studied current intensities were 0, 0.1, 0.3 and 0.6 mAmp, and the durations of iontophoresis were 10 and 60 sec. Two control groups received 1.4% topical drops of gentamicin every 5 min for 1 hr (group 1) or sub-conjunctival injection of 10 mg gentamicin (group 2). Following sacrifice, aqueous humour was taken, corneas were excised, and gentamicin concentration was determined in aqueous humour and cornea samples. RESULTS: Post-iontophoresis, the concentration of gentamicin in the corneas ranged from high (88.60 +/- 38.64 microg ml(-1)) to very low (0.10 +/- 0.89 microg ml(-1)). Both the control groups and those rabbits treated with current intensity of 0.1 mAmp or greater obtained therapeutic gentamicin levels in the corneas. Use of iontophoresis for 60 sec or current intensity greater than 0.1 mAmp obtained corneal gentamicin levels not different from sub-conjunctival injection. Application of current intensity of 0.1 mAmp or greater gave corneal gentamicin concentrations comparable to topical application of the drug, except when 0.6 mAmp were used for 60 sec (p = 0.05). Increasing current intensity or duration of iontophoresis significantly increased (p = 0.001 for both) gentamicin penetration into the cornea. Current intensity had more influence (Beta2 = 0.40) than duration (Beta2 = 0.13) on drug penetration. A significant interaction was found between the duration of iontophoresis and the current intensity. Very small or no concentrations of the drug were discovered in the anterior chambers of rabbits. CONCLUSIONS: Iontophoresis using hydrogel-gentamicin probe may deliver therapeutic concentrations of gentamicin into the cornea.     

The effect of therapeutic soft contact lenses on antibiotic delivery to the cornea

The effects of high (71%) and low (38.6%) water content lenses on antibiotic delivery to the cornea were studied in rabbits by measuring corneal tobramycin concentrations 1, 2, 4 and 6 hours after topical application at intervals of 15 minutes. In the presence of the low water content lens corneal drug levels were higher than in control corneas for every time point assayed. This difference was only significant at 4 hours (P less than 0.05). In eyes wearing the high water content contact lens corneal drug concentration was also higher at every time point tested except one hour. The difference was significant only at 4 hours (P less than 0.01). The data suggest that in the normal, noninflammed eye, the presence of a therapeutic soft contact lens will not compromise aminoglycoside delivery to the cornea.     

角膜发育的分子调控机制研究进展

角膜的发育异常可引起先天性角膜内皮营养不良、Peters异常、Axenfeld—Rieger综合征及眼表鳞状上皮瘤（OSSN）等先天性眼病。角膜的发育经历了从诱导、发生到逐渐成熟的过程,在此期间的细胞增生、凋亡、分化等一系列过程受到多种信号转导通路和转录因子的精确调控。近年来通过胚胎的组织和基因操作技术,人们发现了一系列信号转导通路,如骨形态发生蛋白（BMP）、成纤维细胞生长因子（FGF）、Notch等,和多种转录因子,如Pax6、Krfippel样因子6（KLF6）、转化生长因子8（TGF-β）、TGF—α等均参与了角膜的发育调控过程,它们分别或协同作用于角膜发育的不同阶段,发挥各自的重要功能。主要介绍几种已知在角膜胚胎发育过程中起重要作用的信号转导通路和细胞因子。     

鸡胚发育过程中角膜神经的分布和变化

背景 了解动物或人的角膜神经分布和发育过程对于角膜疾病的临床和基础研究具有重要意义,目前关于动物角膜神经发育和特异性定位的研究结果已有发表,但是有关胚胎发育阶段角膜神经纤维的分布规律和角膜神经纤维的定量研究结果尚少见. 目的 了解鸡胚发育过程中角膜神经纤维的分布,并定量评价随着鸡胚龄增长其角膜神经纤维长度和密度的变化规律. 方法 选取胚胎期6 ～20 d(E6～E20)的鸡胚作为角膜胚胎发育模型,获取相应胚期的带有角膜缘的完整鸡角膜,使用β-tubulinⅢ抗体进行角膜免疫荧光染色,以标记角膜神经,将角膜上皮面朝上做角膜上皮的放射状切开,制备全角膜铺片,用含DAPI抗荧光淬灭缓冲甘油封片.利用正置荧光显微镜拍摄获取整个角膜的神经纤维图像,使用Photoshop CS4测量不同胚龄的鸡胚角膜表面积和神经纤维束数量,采用Imaris x64 7.4.2软件测量不同胚龄的鸡胚角膜神经纤维总长度和密度.结果 全角膜铺片显示,鸡胚E6 ～ E8可见神经束从颞侧巩膜进入角膜缘,E9 ～ E10时神经纤维在角膜缘呈环状分布,E11～ E15时延伸进入角膜中央,E16 ～ E20时角膜形成神经纤维丛.E6～E20期间,鸡胚角膜表面积、角膜神经纤维长度、角膜神经纤维密度均随着胚龄的增长而逐渐增加,总体比较差异均有统计学意义(F=127.007、227.051、67.748,均P＜0.01),鸡胚角膜表面积与角膜神经纤维长度间呈强正相关(r=0.863,P＜0.01).鸡胚角膜神经纤维束从E13开始出现,为(59.00±1.14)/mm2,此后缓慢增加,至E18达到高峰,数量为(576.75±29.16)/mm2,至E20时角膜神经纤维束数量减少,为(299.67±25.46)/mm2,不同胚龄期鸡胚角膜神经纤维束数量的总体比较差异有统计学意义(F=13.759,P=0.000).结论 鸡胚角膜神经发育从E9开始,随着发育鸡胚角膜表面积、角膜神经纤维总长度及密度均快速增加.     

Collagen organization in the secondary chick cornea during development

PURPOSE: The latter stages of morphogenesis in the embryonic chick cornea are instrumental in the establishment of a properly formed corneal stroma. This study was designed to provide better appreciation of collagen reorganization in the avian corneal stroma during the latter stages of embryogenesis. METHODS: High-angle synchrotron x-ray diffraction patterns were obtained from 47 developing chick corneas daily at developmental days 13 through 18 (n = 7 or 8 at each time point) and analyzed to establish collagen molecular spacing and fibril orientation. RESULTS: Collagen intermolecular x-ray reflections were of approximately constant intensity between days 13 and 15 of development, but thereafter became progressively more intense, suggesting that extra collagen is deposited in embryonic chick corneas after day 16 of development. At all times, the mean collagen intermolecular spacing measured approximately 1.43 nm. X-ray intensity was not uniform around the intermolecular x-ray reflections at earlier time points. Rather, a fourfold symmetry was evident, indicative of an orthogonal array of collagen fibrils. An index of this symmetry was essentially unchanged between developmental days 13 and 15, but thereafter diminished considerably. CONCLUSIONS: The lateral spacing of fibril-forming collagen molecules does not change as the chick cornea develops between days 13 and 18. An orthogonal array of collagen fibrils is present in the corneas of developmental day-13 to -18 chicks, but starting at developmental day 16, additional collagen is deposited in a less well-oriented manner and thus acts to obscure the overall orthogonality, with implications for the biomechanical strength and shape of the cornea.     

Rapid ocular angiogenic control via naked DNA delivery to cornea

PURPOSE: To determine the efficacy and safety of naked plasmid gene therapy to the corneal stroma and epithelium. METHODS: Naked plasmid DNA was injected under pressure into the cornea of mice. The expression of genes coding for beta galactosidase (beta-gal), enhanced green fluorescent protein (EGFP), vascular endothelial growth factor (VEGF), and soluble Flt-1 (s-Flt) was recorded and measured with regard to dose, time course, and bioactivity. RESULTS: LacZ gene expression of the protein beta-gal was demonstrated as early as 1 hour, with expression persisting for 10 days. Plasmid-injected corneas remained clear and free of inflammation. EGFP was bicistronically expressed with VEGF to demonstrate the practicality of simultaneous in vivo analysis of gene expression and growth factor bioactivity. Corneal injection of a plasmid containing VEGF cDNA induced corneal and anterior chamber neovascularization. Moreover, corneal injection of plasmid containing the cDNA for the soluble form of the VEGF receptor Flt-1 effectively prevented corneal neovascularization. CONCLUSIONS: The cornea is readily accessible for gene therapy in the laboratory and in the clinic. The method described is safe, effective, titratable, and easily monitored. Naked DNA delivery to the cornea has the potential to alter the treatment of a wide variety of corneal and anterior segment diseases.