CORNEAL CELL-SURFACE ANTIGENS

角膜损伤和失调是损伤的常见原因 幻象。为了更好地管理和治疗角膜愈合 问题，重要的是要了解正常 角膜组织的发育和再生。角膜上皮 已确定的蛋白质(HEBM1)，似乎参与了 胎儿发育过程中的角膜上皮分化 维持正常的成人角膜，并在愈合中。上一次 研究表明，HEBM1是一种新的丝氨酸/苏氨酸蛋白激酶 (主键)。拟议的研究将检验这样的假设：这个新的PK 参与a)介导角膜缘的信号转导通路 正常成人的角膜上皮表型转变，以及b)副 在会痊愈的过程中。HEBM1的特征将是确定 其底物和抑制剂的专一性，涉及的机制 它的合成和酶活性的调节及其在 调节角膜上皮细胞的表型特征。研究ITS 监管角色，其变化之间的时间关系 角膜缘到角膜上皮表型的表达和激活 过渡，将在体外使用兔和人的角膜缘进行评估 外植体培养。另一种方法是以不同的方式表达HEBM1 细胞类型(如角膜成纤维细胞或晶状体上皮细胞) 正常情况下不表达HEBM1(通过将编码的cDNA导入它们 HEBM1)，然后检查形态和表型变化 在这些确实表达HEBM1的转基因细胞中。 细胞骨架结构现在被公认为动态结构 它们参与各种细胞活动，这些活动对 胚胎发育并会痊愈。两个免疫相关的 识别的肌动蛋白细丝(微丝)相关蛋白 由相同的单抗(AFAP50和180)和波形蛋白 (中间丝)相关蛋白(IFAP130)已被鉴定 并被发现由激活的成纤维细胞表达 伤人。以下是关于这些蛋白质功能的假设 (基于初步研究)将进行测试：a)AFAP50是链 延伸因子，EF1a；b)AFAP180是肌动蛋白结合蛋白；c) IFAP130是一种新的结合肌动蛋白和波形蛋白的蛋白质；d) AFAP50/180和IFAP130的表达增加与 角膜成纤维细胞的迁移活性。这些实验将包括 这些蛋白质与蛋白质相互作用的直接生化分析 肌动蛋白和/或波形蛋白细丝(使用纯化蛋白)， 通过分析表征它们的结构-功能关系 它们的cDNA和推导的氨基酸序列及其作用的评价 这些蛋白质在伤口愈合过程中的迁移活性(即， 这些基因的表达和细胞内分布的变化 蛋白质及其相关的细胞骨架网络)。这些研究是关于 调节重要细胞活动的分子机制 在分化和再生过程中，角膜组织会产生 可能导致更好地诊断和治疗角膜的见解 发展和治愈问题。

Injuries and disorders of the cornea are common causes of impaired vision. For the better management and treatment of corneal healing problems, it is important to understand the mechanisms of normal development and regeneration of corneal tissues. A corneal epithelial protein has been identified (HEBM1), that appears to be involved in corneal epithelial differentiation during fetal development, in maintaining normal adult cornea, and in would healing. The previous study indicates that HEBM1 is a novel serine/threonine protein kinase (PK). The proposed studies will test the hypotheses that this novel PK is involved in a signal transduction pathway that a) mediates limbal to corneal epithelial phenotypic transition in normal adult, and b) vice versa during would healing. HEBM1 will be characterized by determining its substrate and inhibitor specificity, the mechanisms involved in regulation of its synthesis and enzymatic activity, and its role in regulating phenotypic features of corneal epithelial cells. To study its regulatory role, the temporal relationship between changes in its expression and activation, and limbal to corneal epithelial phenotypic transition, will be evaluated in vitro using rabbit and human limbal explant cultures. Another approach will be to express HEBM1 in different cell types (e.g. corneal fibroblasts or lens epithelial cells) that normally do not express HEBM1 (by transfecting them with cDNA encoding for HEBM1) and to then examine the morphological and phenotypic changes in these transfected cells that do express HEBM1. Cytoskeletal structures are now well-recognized as dynamic structures that are involved in various cellular activities which are important for embryonic development and would healing. Two immunologically related actin filament (microfilament)-associated proteins that are recognized by the same monoclonal antibody (AFAP50 and 180) and a vimentin (intermediate filament)-associated protein (IFAP130) have been identified and are found to be expressed by activated fibroblasts in response to wounding. The following hypotheses about the functions of these proteins (based on preliminary studies) will be tested: a) AFAP50 is the chain elongation factor, EF1a; b) AFAP180 is an actin binding protein; c) IFAP130 is a novel protein that binds both actin and vimentin; d) increased expressions of AFAP50/180 and IFAP130 are associated with the migratory activity of corneal fibroblasts. The experiments will include direct biochemical analyses of the interactions of these proteins with actin and/or vimentin filaments (using purified proteins), characterization of their structure-function relationship by analyzing their cDNA and deduced amino acid sequences, and evaluation of the role of these proteins in migratory activity during wound healing (i.e., changes in the expression and intracellular distribution of these proteins and their associated cytoskeletal networks). These studies of the molecular mechanisms that regulate important cellular activities during differentiation and regeneration of corneal tissues will yield insights that may lead to better diagnosis and treatment of corneal development and healing problems.