Transplantation of tissue-engineered human corneal epithelium in limbal stem cell deficiency rabbit models

AIM: To evaluate the biological functions of tissue-engineered human corneal epithelium (TE-HCEP) by corneal transplantation in limbal stem cell deficiency (LSCD) rabbit models. METHODS: TE-HCEPs were reconstructed with DiI-labeled untransfected HCEP cells and denuded amniotic membrane (dAM) in air-liquid interface culture, and their morphology and structure were characterized by hematoxylin-eosin (HE) staining of paraffin-sections, immunohistochemistry and electron microscopy. LSCD models were established by mechanical and alcohol treatment of the left eyes of New Zealand white rabbits, and their eyes were transplanted with TE-HCEPs with dAM surface outside by lamellar keratoplasty (LKP). Corneal transparency, neovascularization, thickness, and epithelial integrality of both traumatic and post transplantation eyes were checked once a week by slit-lamp corneal microscopy, a corneal pachymeter, and periodic acid-Schiff (PAS) staining. At day 120 post surgery, the rabbits in each group were sacrificed and their corneas were examined by DiI label observation, HE staining, immunohistochemistry and electron microscopy. RESULTS: After cultured for 5 days on dAM, HCEP cells, maintaining keratin 3 expression, reconstructed a 6-7 layer TE-HCEP with normal morphology and structure. The traumatic rabbit corneas, entirely opaque, conjunctivalized and with invaded blood vessels, were used as LSCD models for TE-HCEP transplantation. After transplantation, obvious edema was not found in TE-HCEP-transplanted corneas which became more and more transparent, the invaded blood vessels reduced gradually throughout the monitoring period. The corneas decreased to normal thickness on day 25, while those of dAM eyes were over 575 mu m in thickness during the monitoring period. A 45 layer of epithelium consisting of TE-HCEP originated cells attached tightly to the anterior surface of stroma was reconstructed 120 days after TE-HCEP transplantation, which was similar to the normal control eye in morphology and structure. In contrast, intense corneal edema, turbid, invaded blood vessels were found in dAM eyes, and no multilayer epithelium was found but only a few scattered conjunctiva-like cells appeared. CONCLUSION: The TE-HCEP, with similar morphology and structure to those of innate HCEP, could reconstruct a multilayer corneal epithelium with normal functions in restoring corneal transparency and thickness of LSCD rabbits after transplantation. It may be a promising HCEP equivalent for clinical therapy of corneal epithelial disorders.

In vitro reconstruction and characterization of tissue-engineered human corneal epithelium with seeder cells from an untransfected human corneal epithelial cell line

AIM: To demonstrate the morphology and structure of in vitro reconstructed tissue-engineered human corneal epithelium (TE-HCEP) with seeder cells from an untransfected HCEP cell line. METHODS: The TE-HCEPs were reconstructed in vitro with seeder cells from an untransfected HCEP cell line, and scaffold carriers of denuded amniotic membrane (dAM) in air-liquid interface culture for 3, 5, 7 and 9 days, respectively. The specimens were examined with hematoxylin-eosin (HE) staining of paraffin-section, immunocytochemical staining, scanning and transmission electron microscopy. RESULTS: During in vitro reconstruction of TE-HCEP, HCEP cells formed a 3-4, 6-7 and 8-10 layers of an HCEP-like structure on dAMs in air-liquid interface culture for 3, 5 and 7 days, respectively. But the cells deceased to 5-6 layers and the structure of straified epithelium became loose at day 9. And the cells maintained positive expression of marker proteins (keratin 3 and keratin 12), cell-junction proteins (zonula occludens-1, E-cadherin, connexin 43 and integrin beta 1) and membrane transport protein of Na+-K+ ATPase. The HCEP cells in TE-HCEP were rich in microvilli on apical surface and established numerous cell-cell and cell-dAM junctions at day 5. CONCLUSION: The morphology and structure of the reconstructed TE-HCEP were similar to those of HCEP in vivo. The HCEP cells in the reconstructed TE-HCEP maintained the properties of HCEP cells, including abilities of forming intercellular and cell-extracellular matrix junctions and abilities of performing membrane transportation. The untransfected HCEP cells and dAMs could promisingly be used in reconstruction HCEP equivalent for clinical corneal epithelium transplantation.

Limbal stem cells: Central concepts of corneal epithelial homeostasis

A strong cohort of evidence exists that supports the localisation of corneal stem cells at the limbus. The distinguishing characteristics of limbal cells as stem cells include slow cycling properties, high proliferative potential when required, clonogenicity, absence of differentiation marker expression coupled with positive expression of progenitor markers, multipotency, centripetal migration, requirement for a distinct niche environment and the ability of transplanted limbal cells to regenerate the entire corneal epithelium. The existence of limbal stem cells supports the prevailing theory of corneal homeostasis, known as the XYZ hypothesis where X represents proliferation and stratification of limbal basal cells, Y centripetal migration of basal cells and Z desquamation of superficial cells. To maintain the mass of cornea, the sum of X and Y must equal Z and very elegant cell tracking experiments provide strong evidence in support of this theory. However, several recent stud-ies have suggested the existence of oligopotent stem cells capable of corneal maintenance outside of the limbus. This review presents a summary of data which led to the current concepts of corneal epithelial homeostasis and discusses areas of controversy surrounding the existence of a secondary stem cell reservoir on the corneal

Evaluating alternative stem cell hypotheses for adult corneal epithelial maintenance

In this review we evaluate evidence for three different hypotheses that explain how the corneal epithelium is maintained. The limbal epithelial stem cell(LESC)hypothesis is most widely accepted. This proposes that stem cells in the basal layer of the limbal epithelium, at the periphery of the cornea, maintain themselves and also produce transient(or transit) amplifying cells(TACs). TACs then move centripetally to the centre of the cornea in the basal layer of the corneal epithelium and also replenish cells in the overlying suprabasal layers. The LESCs maintain the corneal epithelium during normal homeostasis and become more active to repair significant wounds. Second, the corneal epithelial stem cell(CESC) hypothesis postulates that, during normal homeostasis, stem cells distributed throughout the basal corneal epithelium, maintain the tissue. According to this hypothesis, LESCs are present in the limbus but are only active during wound healing. We also consider a third possibility, that the corneal epithelium is maintained during normal homeostasis by proliferation of basal corneal epithelial cells without any input from stem cells. After reviewing the published evidence, we conclude that the LESC and CESC hypotheses are consistent with more of the evidence than the third hypothesis, so we do not consider this further. The LESC and CESC hypotheses each have difficulty accounting for one main type of evidence so we evaluate the two key lines of evidence that discriminate between them. Finally, we discuss how lineage-tracing experiments have begun to resolve the debate in favour of the LESC hypothesis. Nevertheless, it also seems likely that some basal corneal epithelial cells can act as long-term progenitors if limbal stem cell function is compromised. Thus, this aspect of the CESC hypothesis may have a lasting impact on our understanding of corneal epithelial maintenance, even if it is eventually shown that stem cells are restricted to the limbus as proposed by the LESC hypothesis.

TREM-1 expression in rat corneal epithelium with Aspergillus fumigatus infection

AIM: To investigate the expression of triggering receptor expressed on myeloid cells-1(TREM-1) in the aberrant inflammation within the corneal epithelium at early period of fungal infection.METHODS: A total of 65 Wistar rats were randomly divided into control group, sham group and fungal keratitis(FK) group, in which the cornea was infected by Aspergillus fumigatus(A. fumigatus). After executed randomly at 8, 16, 24, 48 and 72 h after experimental model being established, the severity of keratomycosis in rats was scored visually with the aid of a dissecting microscope and slit lamp. Then corneas in three groups were collected to assess the expression of TREM-1through quantitative reverse transcription-polymerase chain reaction(RT-PCR), immunofluorescence technique and Western blot analysis. The correlation between FK inflammation and expression of TREM-1 was also analyzed.RESULTS: Corneal inflammation scores increased with time after fungal infection(F =49.74, P =0.000). The inflammation scores in FK group were obviously higher than those in sham group on the whole(F =137.78, P =0.000). Levels of TREM-1 in the infected rat corneal epithelium had elevated at 8h and peaked at 48h(P 【0.001,compared with control group). Western blot analysis also showed an obviously elevated TREM-1 level in rat corneal epithelium at 24 h and 48 h after fungal infection.Immunofluorescence technique showed that TREM-1mainly existed in corneal epithelium and infected corneal stoma of rat. TREM-1 protein expression was enhanced after fungal infection. Moreover, severity of FK inflammation was significantly related to TREM-1expression in FK(r =0.942, P =0.000).CONCLUSION: TREM-1 may contribute to amplify theinflammation in the cornea infected with A. fumigatus and play critical roles in the battle against A. fumigatus in the innate immune responses.

应用活体共聚焦显微镜观察糖尿病患者的角膜细胞

活体角膜共聚焦显微镜是一种无创、快速且全层面的技术,可以实时、动态地观察角膜组织的所有层面。共聚焦显微镜可以通过直接可视化检查角膜不同层面的形态和细胞密度。随着糖尿病患病率的不断上升,眼部并发症已经变得越发常见,并引起眼科临床工作者和科研工作者越来越多的兴趣和逐渐深入的研究。文章旨在通过采用活体角膜共聚焦显微镜观察糖尿病患者角膜组织各层的研究进展进行综述。

Putative epidermal stem cell convert into corneal epithelium-like cell under corneal tissue in vitro

Rhesus putative epidermal stem cells are being investigated for their potential use in regenerative corneal epithelium-like cells, which may provide a practical source of autologous seed cells for the construction of bioengineered corneas. The goal of this study was to investigate the potential of epi-dermal stem cells for trans-differentiation into corneal epithelium-like cells. Rhesus putative epidermal stem cells were isolated by type IV collagen attachment method. Flow cytometry analysis, immuno-histology and RT-PCR were conducted to identify the expression of specific markers (β1, α6 integrin, K15, K1/K10, K3/K12 and CD71) on the isolated rapid attaching cells. The isolated cells were cocultured with human corneal limbal stroma and corneal epithelial cells. After coculture, the expression of the same specific markers was evaluated in order to identify expression difference caused by the coculture conditions. K3/K12 expression was analyzed in coculture cells on day 2, 4, 6, 8 and 10. Putative epi-dermal stem cells in conditioned culture media were used as control. Putative epidermal stem cells were predominant in rapid attaching cells by type IV collagen attachment isolation. Before being co-cultured, the rhesus putative epidermal stem cells expressed K15, α6 and β1 integrin, but no CD71, K1/K10 and K3/K12. After coculture, these cells expressed K3/K12 (a marker of corneal epithelial cells), K15 and β 1 integrin, but no K1/K10. Cells being not coculture converted into terminally differentiated cells expressing K1/K10. These results indicate that rhesus putative epidermal stem cells can trans-differentiate into corneal epithelium-like cells and, therefore, may have potential therapeutic application as autologous seed cells for the construction of bioengineered corneas.

Differentiation of embryonic stem cells into corneal epithelium

Our project was to determine whether embryonic stem (ES) cells could be induced to differentiate into corneal epithelia by superficial corneoscleral limbal stroma. To achieve this goal, ES-GFP cell line D3 was pre-induced by retinoic acid (RA). The pre-induced cells were seeded on deepithelialized superficial corneoscleral slices (SCSS) to form a monolayer, and di-vided into three groups. Group 1 was cultured and passaged in vitro for direct detection. Group 2 was exposed to air-liquid interfaces for 10 days and implanted into the subcutaneous layer of nude mice for 2 weeks for further induction in vivo. Group 3 was cultured in vitro without any in-ducing factors for control. There were no teratomas found in nude mice which were implanted with differentiated ES cells after two weeks. The differentiated cells showed an appearance of epithelia both in vitro and in vivo. Expression of CK3, P63 and PCNA was detected by immuno-histochemical staining in the differentiated cells in group 1 and 2. Microvillis and zonula oc-cludens were observed on the surface of the differentiated cells under an electron microscope. In the control group, ES cells differentiated freely without any inducing factors. Most cells were shed and formed a neuronal dendrite-like structure, and a minority of cells appeared polymorphic. These results demonstrate that ES cells can differentiate into corneal epithelia on the surface of SCSS under the controlled condition. Differentiated ES cells could be used as epithelial seeding cells for the reconstruction of ocular surface and corneal tissue engineering in the future.

诱导骨髓间质干细胞分化为角膜上皮样细胞的初步研究

目的:观察人骨髓间质干细胞(MSCs)移植到兔角膜基质后的分化发育情况,探讨MSCs分化为角膜上皮细胞的可行性。方法:24只新西兰兔随机分为2组。实验组:将人MSCs接种在保存人羊膜上培养4d,用5-溴脱氧尿嘧啶(BrdU)标记后移植到兔角膜基质;对照组:采用保存羊膜移植到兔角膜基质。分别于移植后1、2、3、4、6和8周,摘取各组实验眼行组织学和免疫组织化学检查,检查移植到兔角膜基质的MSCs的存活、形态变化以及移植局部的反应等情况;免疫组织化学检测移植到角膜基质的带有BrdU标记的细胞角蛋白K3/12(CK3/CK12)和角蛋白K13(CK13)的表达。结果:MSCs接种到羊膜后能在羊膜上生长,与羊膜共培养4d后,MSCs贴附羊膜生长迅速,组织学特征无明显改变。羊膜负载MSCs移植到兔角膜基质表面,术后免疫组织化学检测角膜上皮层CK3/CK12表达阳性,CK13表达阴性,在重建的角膜上皮层可检测到BrdU核阳性细胞并同时表达角膜上皮细胞特异性表面标志蛋白K3/K12,未发生免疫排斥反应,未见异常增殖细胞。结论:羊膜负载MSCs移植到兔眼表角膜基质后,MSCs能存活、增殖并向角膜上皮样细胞分化。

重组人表皮生长因子促人角膜缘上皮细胞生长的研究

目的:探讨重组表皮生长因子(rhEGF)对人角膜缘上皮细胞生长的影响。方法:采用rhEGF作用于体外培养的人角膜缘上皮细胞,记录细胞生长曲线,并用四唑氮盐(MTT)法测定细胞增殖状况。结果:rhEGF浓度为10ng/ml时对人角膜缘上皮细胞的促增殖作用最强(P<0.05)。MTT法48h的吸光度值实验组均高于对照组(P<0.01)。结论:rhEGF对体外培养人角膜上皮细胞有明显促增生作用。

苯扎氯胺对体外培养的人角膜上皮细胞毒性作用的研究

目的观察滴眼液中最常用的防腐剂-苯扎氯胺(BAC)对体外培养的人角膜上皮细胞的毒性作用。方法建立人角膜上皮细胞的体外培养模型。不同质量浓度的BAC(0.1%、0.01%、0.001%、0.0001%)与细胞共同孵育,时间分别为10min、30min,2、6、24、48h。评估细胞毒性的方法:MTT分析,光镜和扫描电镜观察细胞形态学改变。结果0.1%BAC导致细胞立即死亡,MTT值在暴露时间为10min时显著下降(P<0.01),30min降至最低(P<0.01);光镜下细胞迅速脱壁。0.01%BAC在24h内导致细胞缓慢死亡,MTT值逐渐下降;扫描电镜示细胞的形态学改变为表面微绒毛脱落,细胞膜破损;0.001%～0.0001%BAC对细胞的损害出现在24h后。结论BAC对体外培养的人角膜上皮细胞具有明确的毒性作用,即使在滴眼液中含有的常规质量浓度(0.01%)及低于常规质量浓度下仍可出现。部分临床长期使用含有BAC滴眼液患者可出现眼表问题。

脱细胞猪角膜基质体外支持角膜上皮和基质细胞的生长(英文)

目的：探讨脱细胞猪角膜基质体外能否支持兔角膜细胞的生长。方法：体外培养兔角膜上皮细胞和基质细胞,并接种到制备的脱细胞猪角膜基质上,倒置相差显微镜和组织学观察细胞生长情况。结果：上皮细胞能在脱细胞猪角膜基质上贴附生长,10d时可形成2～3层的复层结构。基质细胞在脱细胞猪角膜基质上贴附生长后可向材料深层迁徙。结论：制备的脱细胞猪角膜基质体外可支持兔角膜上皮细胞和基质细胞的生长。

自体角膜缘部上皮移植治疗翼状胬肉

采用自体角膜缘部上皮移植治疗原发性胬肉５５眼，复发性胬肉６眼。经３～６个月随访，原发性胬肉全部治愈无复发，复发性胬肉有一例复发。研究表明本手术法与结膜移植法相比有复发率低、眼部不充血、角膜上皮恢复时间短、角膜无新生血管等优点，并对自体角膜缘部上皮移植治疗翼状胬肉的机理进行了探讨。

活体及体外条件下对角膜上皮干细胞的定位

背景:眼表存在两种形式的上皮干细胞即角膜上皮干细胞和结膜上皮干细胞,角膜上皮干细胞在角膜上皮细胞更新和角膜透明的维持方面起着重要作用。目的:采用活体激光扫描角膜共焦显微镜和免疫荧光染色技术相结合的方法,从活体和体外层面上对角膜上皮干细胞进行定位研究。方法:收集2009年9月至2012年9月来河南省眼科研究所就诊的单侧角膜缘干细胞缺乏患者,使用活体激光扫描角膜共焦显微镜检查患者双眼,健侧眼为对照。扫描方位依次为中央角膜及上、下、左、右方的角膜缘,记录扫描图像并分析。眼球材料来自于河南省眼库,切取角膜中央和角膜缘组织,组织包埋剂包被、冰冻切片,切片厚度5-7μm;免疫荧光染色技术检测p63、ABCG2、K3和Connexin 43在角膜中央及角膜缘上皮层的表达。结果与结论:共有24例患者确诊为单侧角膜缘干细胞缺乏,活体激光扫描角膜共焦显微镜下患侧眼角膜病变区可见结膜细胞及杯状细胞;角膜缘区域Vogt栅栏状结构消失,色素细胞消失,取而代之的是大量纤维瘢痕化组织。免疫荧光染色示表达ABCG2和p63的细胞主要在角膜缘上皮基底层,尤其在近结膜侧的角膜缘及角膜缘中间部表达相对较高,而中央角膜上皮层细胞不表达;K3及Connexin43在角膜缘上皮基底细胞层不表达,中央角膜上皮全层表达。通过活体激光扫描角膜共焦显微镜观察及干细胞标记物检测显示角膜上皮干细胞主要存在于角膜缘外2/3区域的Vogt栅栏基底部及钉突结构中。

小牛血去蛋白提取物眼用凝胶对LASIK术后角膜损伤修复的疗效观察

目的探讨小牛血去蛋白提取物眼用凝胶对准分子激光原位角膜磨镶术(laser in situ keratomileusis,LASIK)后角膜损伤修复的影响。方法选取在我院行LASIK患者114例(228眼)。患者左右眼随机分为治疗组和对照组,治疗组应用小牛血去蛋白提取物眼用凝胶,对照组不应用。测定两组术前、术后1d、1个月、3个月时泪液分泌量、BUT、角膜上皮下神经丛密度和角膜知觉。对上述4个指标采用SPSS13.0软件进行统计分析比较。结果治疗组术前泪液分泌量和BUT分别为(26.35±0.90)mm、(9.88±0.48)s;术后1d分别为(11.59±1.04)mm、(1.76±0.20)s,较术前明显降低(均为P<0.05);术后3个月分别为(25.82±0.89)mm、(8.76±0.40)s,与术前比较差异无统计学意义(均为P>0.05)。对照组术前泪液分泌量和BUT分别为(26.59±0.10)mm、(10.06±0.45)s;术后1d、术后3个月时变化趋势与治疗组相同,与治疗组同时期数据比较差异无统计学意义(均为P>0.05)。术后1个月治疗组泪液分泌量和BUT分别为(20.35±0.89)mm、(6.18±0.45)s;对照组分别为(17.24±0.77)mm、(4.71±0.23)s;治疗组两个指标的恢复程度均明显快于对照组,差异有统计学意义(t=7.012、3.293,均为P<0.01)。治疗组术前角膜上皮下神经丛密度和角膜知觉分别为(1588.00±173.53)μm、(60.00±0.00)mm;术后1d分别为(254.00±53.29)μm、(0.59±0.04)mm,较术前明显降低(均为P<0.05);术后1个月为(208.00±32.58)μm、(15.59±1.81)mm,术后3个月为(588.00±65.18)μm、(41.18±3.44)mm,均低于术前水平(均为P<0.05)。对照组术前角膜上皮下神经丛密度和角膜知觉分别为(1134.00±68.50)μm、(60.00±0.00)mm;术后1d、1个月、3个月时变化趋势与治疗组相同,与治疗组同时期数据比较差异无统计学意义(均为P>0.05)。结论 LASIK术后早期使用小牛血去蛋白提取物眼用凝胶有利于角膜表面损伤的修复,缓解术后干眼症状;但对于角膜神经的损伤修复无明显作用。

人胚肺成纤维细胞饲养兔角膜上皮和内皮细胞的体外培养

目的 研究人胚肺成纤维饲细胞体外培养兔角膜上皮和内皮细胞。方法 人胚肺成纤维细胞系细胞，经丝裂霉素Ｃ处理成饲细胞。兔角膜上皮和内皮细胞组织块原代培养，消化接种于人胚肺饲细胞瓶传代培养。体外培养的角膜上皮和内皮细胞行常规病理形态学检查，角膜上皮细胞行角蛋白、内皮细胞行神经烯醇化酶免疫组化染色检测。结果人胚肺成纤维细胞形态均一，易于分辨，经丝裂霉素处理后失去增殖能力。角膜上皮和内皮细胞原代培养５～７天，细胞密集，经人胚肺饲细胞共培养５代，细胞无衰老现象。检测角膜上皮细胞角蛋白、内皮细胞神经烯醇化酶表达阳性，人胚肺饲细胞表达阴性。结论 人胚肺饲细胞能够明显增强角膜上皮和内皮细胞的体外生存能力，提高细胞的增殖能力。

培养的角膜缘上皮细胞增殖能力的动态研究

目的 探讨组织块培养角膜缘上皮细胞在不同时间以及第 2、3代培养的角膜缘上皮细胞早期 (6～ 8d)的增殖能力。方法 应用组织块培养法培养角膜缘上皮细胞 ,在d8、12、16、2 0、2 4用流式细胞仪检测其细胞周期 ,同时对组织块培养 16d的角膜缘上皮细胞进行传代 ,用同样方法检测第 2、3代细胞出现汇合时 (6～ 8d)的细胞周期。结果 组织块培养角膜缘上皮细胞增殖指数 (ProliferativeIndex ,PI)在高水平上呈下降趋势 (分别为 31 2 %、2 0 1%、17 6 %、15 6 %、13 4 % ) ;2、3代培养细胞早期的增殖指数较高 (分别为 30 3%、2 6 % )。结论 组织块培养的角膜缘上皮细胞增殖能力在 8～ 2 4d内在高水平上呈下降趋势 ,第 2、3代培养的细胞在培养早期的增殖能力也较高 。

胚胎干细胞体外培养及在眼科的研究进展

胚胎干细胞(ESC)具有体外无限增生和自我更新并能够分化为体内各种细胞的潜能。ESC独特的生物学特性被广泛应用于生物和医学研究领域,因此,成功地培养ESCs对于更深入地探讨ESCs的作用机制是非常有帮助的。目前,培养和分化的ESCs已用于一些眼科疾病的治疗,如视网膜变性性疾病和严重的角膜疾病等。ESC在动物和人的胚胎发育、致畸实验、组织工程和移植治疗等研究领域具有重要的科学意义和巨大的应用前景。对胚胎干细胞的发展、体外培养体系及在眼科研究进展进行综述。

角质细胞生长因子促人角膜上皮细胞生长的研究

目的寻找促进角膜上皮损伤修复的有效方法。方法用３Ｈ胸腺嘧啶核苷（３ＨＴｄＲ）掺入及液体闪烁技术，观察角质细胞生长因子（ＫＧＦ）对体外培养的人角膜上皮细胞ＤＮＡ合成的影响，并计算细胞倍增时间。结果１～１００ｎｇ／ｍｌＫＧＦ有明显促进人角膜上皮细胞ＤＮＡ合成的作用，且呈剂量依赖性（ｒ＝０．９２３３，Ｐ＜０．００１）。１０ｎｇ／ｍｌＫＧＦ明显缩短了细胞倍增时间（３１．５９±４．８８ｈ，与对照组４０．９８±５．２０ｈ比较，Ｐ＜０．０５）。结论外源性ＫＧＦ对体外培养的人角膜上皮细胞有明显的促细胞增生作用。表明ＫＧＦ具有应用于临床，促角膜上皮损伤修复的可能性。

自体角膜缘部上皮移植治疗眼球表面疾病

本文采用自体角膜缘上皮移植术治疗145例眼表疾病，并用常规手术方法治疗50例单纯性胬肉。术后观察6～18月，结果：上皮移植组新生角膜上皮平均7天覆盖全部角膜表面，而对照组则需要12天；上皮移植组4～5周角膜表面新生血管逐渐退缩，而对照组则需15～18周。采用自体角膜缘上皮移植组145例有1例复发，而对照组50例中有17例复发。统计学处理有极显著差异P＜0．01。讨论了自体角膜缘上皮移植手术的机制，认为此手术是一个有推广意义的新术式。