ORIGIN AND MAINTENANCE OF THE OCULAR SURFACE EPITHELIA

眼表上皮的起源和维持

• 批准号: 8514617
• 负责人: DAVID CY BEEBE
• 金额: $ 28.88万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8514617
• 关键词: Ablation; Address; Adhesions; Adult; Animals; Applications Grants; Binding; Blindness; Cell Adhesion Molecules; Cell Proliferation; Cell physiology; Cell-Cell Adhesion; Cells; Chemicals; Collaborations; Complement; Conjunctival Epithelium; Cornea; Corneal Stroma; Critical Pathways; Cytokine Signaling; Development; Differentiation Antigens; Ectoderm; Embryo; Embryonic Development; Epithelial; Epithelial Cells; Epithelium; Exons; Eye; Eye diseases; Eyelash; Eyelid structure; Film; Gene Expression Regulation; Gene Targeting; Generations; Genes; Genetic; Genome; Gland; Green Fluorescent Proteins; Hereditary Disease; Image; In Situ Hybridization; In Vitro; Inflammation; Injury; Knowledge; Label; Laboratories; Lacrimal gland structure; Lasers; Life; Limbus Corneae; Lipids; Maintenance; Mediating; Methods; Microarray Analysis; Microdissection; Mucous body substance; Mus; N-Cadherin; Pathway interactions; Pattern; Play; Property; Proteins; Publications; Reporter Genes; Reporting; Research Personnel; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Signaling Pathway Gene; Site; Specific qualifier value; Stem cells; Stromal Cell-Derived Factor 1; Surface; System; Tamoxifen; Testing; Time; Tissue Differentiation; Tissues; To specify; Undifferentiated; Vision; Work; adult stem cell; bone morphogenetic protein receptors; cell type; conjunctiva; corneal epithelial stem cells; corneal epithelium; eye dryness; immunocytochemistry; in vitro Assay; in vivo; induced pluripotent stem cell; injured; irritation; lacrimal; limbal; meibomian gland; neovascularization; notch protein; novel; ocular surface; prevent; recombinase; selective expression; stem; stem cell biology; stem cell niche; tool; transcription factor

DESCRIPTION (provided by applicant): The tissues on the surface of the eye and the secretory glands that are derived from these tissues are essential for vision. The lacrimal, Meibomian and conjunctival mucus glands secrete the components of the tear film. Insufficient function of any of these glands leads to dry eye disease. The corneal and limbal epithelia are responsible for maintaining the refractive surface of the eye. Deficiencies in the differentiation f the corneal epithelial cells or insufficient generation of corneal epithelial cells by the limbal sem cells leads to severe ocular irritation, inflammation, neovascularization of the corneal stroma and blindness. The aims of this proposal are 1) to identify the signaling systems that are responsible for the proper formation and function of the ocular surface epithelia and their derivatives during development and 2) to identify the signaling pathways that establish and maintain the limbal stem cells to provide functional corneal epithelial cells. The studies described in this proposal show that Pax6 and BMP signaling are key factors required for the formation and differentiation of all of the ocular surface epithelia. Laser microdissection and microarray analysis identified or confirmed four transcription factors that are early markers for the different ocular surface epithelia and are likely to play important roles in their differentiaton. Three of these factors depend on Pax6 for their expression. This proposal also describes a new method to genetically mark the limbal stem cells. At the same time, genes within these cells can be selectively deleted. This method will be used to inactivate critical pathways known to function in other adult stem cells. Three of these pathways, BMP and SDF-1 signaling and adhesion between niche and stem cells mediated by N-cadherin, have been shown by our collaborator on this project, Dr. Scheffer Tseng, to be involved in signaling between limbal stem and niche cells. The functions of these pathways will be tested in vivo by genetic ablation and confocal imaging. In each case, our vivo analyses will be complemented by in vitro genetic studies performed in Dr. Tseng's laboratory. This collaboration is possible because Dr. Tseng isolates limbal stem and adherent niche cells after overnight incubation. We will send eyes from our genetically-modified mice to Dr. Tseng by overnight courier. He will isolate the niche and stem cells and treat them with tamoxifen to delete the targeted genes. This collaboration will provide the most extensive analysis to date of the pathways that create and maintain the limbal stem cell niche. Information derived from both aims will be valuable for the replacement of injured or defective corneal and conjunctival epithelia using induced pluripotent cells or by "reprogramming" of other epithelial cell types.

描述（由申请人提供）：眼表面的组织和从这些组织中得出的分泌腺对视觉至关重要。泪，meibomian和结膜粘液腺体分泌泪膜的成分。这些腺体中的任何一个功能不足会导致眼睛疾病干燥。角膜和角膜缘上皮负责保持眼睛的折射表面。角膜上皮细胞的分化缺乏或角膜缘细胞的角膜上皮细胞不足会导致严重的眼部刺激，炎症，角膜基质和失明的新血管形成。该提案的目的是1）确定负责在开发过程中构成眼表上皮及其衍生物的正确形成和功能的信号传导系统，以及2）确定建立和维持边缘干细胞以提供功能性角膜上皮细胞的信号传导途径。该提案中描述的研究表明，PAX6和BMP信号传导是所有眼表上皮的形成和分化所需的关键因素。激光显微解剖和微阵列分析确定或确认了四个转录因子，这些转录因子是不同眼表上皮的早期标记，并且很可能在其区分中起重要作用。其中三个因素取决于PAX6的表达。该提案还描述了一种新的方法，可以在遗传上标记缘干细胞。同时，可以选择性地删除这些细胞内的基因。该方法将用于灭活其他成年干细胞中已知功能的关键途径。这些途径中的三个，即N-钙粘着蛋白介导的小裂和干细胞之间的BMP和SDF-1信号传导以及我们在该项目Scheffer Tseng博士上的介导的干细胞之间的粘附，并参与了边缘干细胞和小细胞之间的信号传导。这些途径的功能将通过遗传消融和共聚焦成像在体内进行测试。在每种情况下，我们的体内分析都将与Tseng博士实验室中进行的体外遗传研究相辅相成。这种合作之所以可能，是因为腾登博士在隔夜孵化后隔离了层长茎和粘附的小众细胞。我们将通过夜间快递将眼睛从遗传改性的小鼠眼中送往Tseng博士。他将分离利基和干细胞，并用他莫昔芬对其进行处理以删除靶向基因。这项合作将为创建和维护缘干细胞生态位的途径迄今为止提供最广泛的分析。从这两个目标中得出的信息对于使用诱导的多能细胞或“重新编程”其他上皮细胞类型的替代受伤或有缺陷的角膜和结膜上皮都很有价值。