Regulation of Lens Fiber Cell Organization

晶状体纤维细胞组织的调节

• 批准号: 8487408
• 负责人: RENPING ZHOU
• 金额: $ 34.3万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8487408
• 关键词: Adherens Junction; Age; Antibodies; Blindness; Cadherins; Cataract; Cell Adhesion; Cell Communication; Cell Differentiation process; Cell Shape; Cells; Characteristics; Chimeric Proteins; Connexins; Cytoplasm; Defect; Development; Differentiation Antigens; Disease; Electron Microscope; Embryo; Ensure; Eph Family Receptors; EphA2 Receptor; Ephrin-A5; Ephrins; Family; Fiber; Future; Health; Human; Immunohistochemistry; In Situ Hybridization; Knockout Mice; Knowledge; Lead; Lens Fiber; Lens Opacities; Ligands; Light Microscope; Mediating; Molecular; Morphology; N-Cadherin; Nervous system structure; Organelles; Pathogenesis; Phenotype; Prevention; Refractive Indices; Regulation; Role; Signal Transduction; Staging; Structure; Techniques; Testing; Time; Western Blotting; axon guidance; fiber cell; insight; lens; lens transparency; light transmission; mutant; novel; postnatal; receptor; receptor-mediated signaling; research study; Adherens Junction; Age; Antibodies; Blindness; Cadherins; Cataract; Cell Adhesion; Cell Communication; Cell Differentiation process; Cell Shape; Cells; Characteristics; Chimeric Proteins; Connexins; Cytoplasm; Defect; Development; Differentiation Antigens; Disease; Electron Microscope; Embryo; Ensure; Eph Family Receptors; EphA2 Receptor; Ephrin-A5; Ephrins; Family; Fiber; Future; Health; Human; Immunohistochemistry; In Situ Hybridization; Knockout Mice; Knowledge; Lead; Lens Fiber; Lens Opacities; Ligands; Light Microscope; Mediating; Molecular; Morphology; N-Cadherin; Nervous system structure; Organelles; Pathogenesis; Phenotype; Prevention; Refractive Indices; Regulation; Role; Signal Transduction; Staging; Structure; Techniques; Testing; Time; Western Blotting; axon guidance; fiber cell; insight; lens; lens transparency; light transmission; mutant; novel; postnatal; receptor; receptor-mediated signaling; research study

DESCRIPTION (provided by applicant): Lens transparency is made possible by a combination of the highly ordered organization of the lens fiber cells, their unique refractive index, and the lack of organelles in the fiber cells. The highly ordered arrangement of the lens fiber cells is critical for proper light transmission, and disruption of this structure by alterations of cell-cell interactions is likely to lead to cataracts. However, signals that regulate lens fiber cell interaction remain largely unknown. Our preliminary studies have identified a new class of molecules, the Eph tyrosine kinase receptor family that regulates lens cell organization. Inactivation of ephrin-A5, a ligand of the Eph receptors, leads to the disruption of N-cadherin localization, change in lens fiber cell shape, disorganization of lens cells, and the development of cataracts. We hypothesize that ephrin-A5, interacting with its receptor(s), regulates N-cadherin-mediated fiber cell adhesion to maintain proper lens cell organization. To test this hypothesis, we will: (1) Examine the spatial and temporal characteristics of the ephrin-A5-null lens, determine when and where during development the lens defects first occur, and whether the loss of ephrin-A5 results in disruptions of lens fiber cell differentiation. The morphology of the lens at different developmental stages will be analyzed using both light and electron microscope techniques. Antibodies against markers of lens cell differentiation will be used in immunohistochemical experiments to study the expression of differentiation markers. (2) Elucidate receptor mechanisms of ephrin-A5 in lens development by examining which specific Eph receptors are expressed in the developing lens and where they are expressed, using Real-Time PCR, in situ hybridization, and immunohistochemistry. Since the interaction between Eph receptors and ligands leads to bidirectional signaling, we plan to analyze whether receptor- mediated signaling, the ligand-mediated reverse signaling or both are required for lens development using selective inactivation of different receptor domains. (3) Study the molecular alterations that lead to cataracts in ephrin-A5-null mice. Preliminary studies have revealed a disruption of N-cadherin distribution in the lens fiber cells. We will determine whether ephrin-A5 receptors interact physically with adherens junction molecules, and analyze effects of the ligand on N-cadherin functions. To critically evaluate roles of N-cadherin in mediating ephrin-A5 function and lens cell adhesion, we plan also to examine expression of a N-cadherin-2-catenin fusion protein in a phenotypic rescue experiment. The proposed studies will establish roles of a previously unsuspected family of molecules in lens development and reveal novel regulations of N-cadherin functions. These studies will enhance our understanding of how lens cell interaction is regulated to ensure lens transparency and provide insights into the mechanisms of cataractogenesis.

描述（由申请人提供）：透镜透明度是通过晶状体纤维细胞的高度有序组织，其独特的折射率以及纤维细胞中缺乏细胞器的组合来实现的。透镜纤维电池的高度有序排列对于适当的光传输至关重要，并且通过细胞 - 细胞相互作用的改变来破坏这种结构可能导致白内障。但是，调节透镜纤维相互作用的信号在很大程度上尚不清楚。我们的初步研究已经确定了一种新的分子，即调节晶状体细胞组织的EPH酪氨酸激酶受体家族。 EPHRIN-A5（EPH受体的配体）的失活导致N-钙粘蛋白定位的破坏，透镜纤维细胞形状的变化，透镜细胞的混乱和白内障的发展。我们假设ephrin-A5与其受体相互作用，调节N-钙粘蛋白介导的纤维细胞粘附以保持适当的晶状体细胞组织。为了检验这一假设，我们将：（1）检查晶状体A5-NULL透镜的空间和时间特征，确定在发育期间何时何地出现镜头缺陷，以及ephrin-A5的丧失是否导致透镜纤维细胞分化的破坏。将使用光和电子显微镜技术分析不同发育阶段的镜头的形态。针对晶状体细胞分化标记的抗体将用于免疫组织化学实验，以研究分化标记的表达。 （2）通过检查在发育中的晶状体中表达哪些特异性EPH受体以及使用实时PCR，原位杂交和免疫组织化学表达的特定EPH受体，从而阐明了晶状体发育中Ephrin-A5的受体机制。由于EPH受体和配体之间的相互作用导致双向信号传导，因此我们计划分析受体介导的信号传导，配体介导的反向信号传导是否需要使用不同受体结构域的选择性失活晶状体开发。 （3）研究导致ephrin-A5-Null小鼠白内障的分子改变。初步研究表明，晶状体纤维细胞中N-钙粘蛋白分布的破坏。我们将确定ephrin-A5受体是否与粘附分子进行物理相互作用，并分析配体对N-钙粘蛋白功能的影响。为了批判性评估N-钙粘蛋白在介导Ephrin-A5功能和透镜细胞粘附中的作用，我们还计划在表型救援实验中检查N-钙粘蛋白-2-catenin融合蛋白的表达。拟议的研究将在晶状体发育中建立先前未经引起的分子家族的作用，并揭示N-钙粘蛋白功能的新法规。这些研究将增强我们对晶状体细胞相互作用的调节方式的理解，以确保晶状体透明度并提供有关白内生生成机制的见解。