中文摘要：

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。

英文摘要：

BACKGROUND：There are two types of epithelial stem cells in the ocular surface tissue：corneal epithelial stem cells and conjunctival epithelial stem cells. The corneal epithelial stem cells play an important role in renewal of corneal epithelial cells and maintenance of corneal transparency. OBJECTIVE：To study the location of corneal epithelial stem cells using laser in vivo confocal microscopy and immunofluorescent staining. METHODS：Patients with unilateral limbal stem celldeficiency who went to Henan Eye Institute from September 2009 to September 2012 were enrol ed in this study. Bilateral eyes were scanned by laser in vivo confocal microscopy, and the healthy eye was imaged as a control. The central cornea and limbus were scanned and images were recorded for statistical analysis. The eye bal s were obtained from Henan Eye Bank, China. Central cornea and limbus were dissected and embedded in the OCT compound for frozen section and the proper thickness of the section was 5-7μm. Immunofluorescent staining was used to detect the expression of p63, ABCG2, K3 and Connexin 43 in the epithelial layers of central cornea and limbus. RESULTS AND CONCLUSION：Twenty-four patients diagnosed with unilateral limbal stem celldeficiency were recruited. Under confocal microscopy, in the affected eyes, the typical morphology of conjunctival epithelial cells＆amp;nbsp;and goblet cells was detected instead of corneal epithelial cells;in the limbus, a great amount of fiber scarring tissue was detected instead of Vogt palisade, rete pegs and pigment cells. Immunofluorescent staining showed the expression of p63, ABCG2 was mainly in the basal layer of limbal epithelium, especial y in the outer and middle parts, but the expression of p63 and ABCG2 was not detected in the epithelial celllayers of central cornea. K3 and Connexin43 were not expressed in suprabasal layers of limbal epithelium, but in central cornea, they were expressed highly in the whole epithelial celllayers. Laser in vivo confocal microscopy and immunofluor