Vitamin D metabolism and function in the cornea and anterior segment

描述（由申请人提供）：据估计，超过50%的美国人口维生素D （Vit D）不足。在数以万计的关于维生素D的发表研究中，只有少数研究了角膜和前段的维生素D，而且没有一项研究测量了这些部位的维生素D浓度、代谢或活性。系统性维生素D缺乏症可导致或加剧前段维生素D缺乏症。鉴于我们的初步数据显示缺乏维生素D受体导致伤口愈合延迟，这可能是角膜损伤后值得关注的问题。我们也证明了维生素D增强角膜上皮屏障功能。除了在眼睛中可能的免疫作用外，也有研究表明维生素D在眼压控制和年龄相关性黄斑变性中起作用。在过去的两年里，我们进入了这个新的令人兴奋的研究领域，即检查眼睛前段的维生素D （Vit D）。拟议的项目将继续为未来角膜和眼睛的所有Vit D工作奠定基础。我们的主要假设是Vit D促进眼睛细胞分化，并且Vit D存在于整个眼睛中，并被特异性地转运到泪液中，从而影响上皮细胞的分化。基于这些假设，本提案的三个具体目标是：(1)确定Vit D是否影响角膜上皮分化；(2)检测VDR基因敲除对角膜上皮创面愈合的影响；(3)确定Vit D如何转运到泪液中。在Aim 1中，体内实验将检测Vit D受体（VDR）敲除小鼠的分化标记。体外实验将检测暴露于不同浓度25(OH)D3和125 (OH)2D3的培养角膜上皮细胞的分化标志物。我们还将研究occludin和其他Vit D代谢物在角膜上皮生理学中的作用。对于Aim 2，将在出生后的不同时间检测-/-和-/+ VDR敲除小鼠的角膜上皮伤口愈合情况。相同组饲喂特殊补充饲料，并检查角膜上皮伤口愈合情况。连接复合物和分化标记也将在所有这些小鼠中进行检查。在第三阶段，将收集正常或高维生素D饲粮兔的泪液，并测量维生素D代谢物。将在所有样品中检测Vit D结合蛋白以及meggalin和cubilin（其他重要的Vit D蛋白载体），以确定可能的Vit D进入眼室的运输途径。

DESCRIPTION (provided by applicant): It is estimated that more than 50% of the United States population has vitamin D (Vit D) insufficiency. Of the tens of thousands of published studies devoted to Vit D, less than a handful have examined Vit D in the cornea and anterior segment and none of these measured Vit D concentrations, metabolism, or activity at those sites. Systemic hypovitaminosis D could lead to or exacerbate anterior segment hypovitaminosis D. This may be of concern following corneal injury given that our preliminary data showing that lack of the vitamin D receptor leads to delayed wound healing. We have also demonstrated that Vit D enhances corneal epithelial barrier function. Apart from its likely immune role in the eye, it has also been suggested that Vit D plays a role in intraocular pressure control and age related macular degeneration. Over the last 2 years we have entered into this new and exciting area of research, namely examining vitamin D (Vit D) in the anterior segment of the eye. The proposed project will continue to lay the foundation for all future Vit D work in te cornea and eye. Our primary hypotheses are that Vit D promotes cell differentiation in the eye, and that Vit D is present throughout the eye and is specifically transported into tear fluid where t can influence epithelial differentiation. The three Specific Aims of this proposal, based on these hypotheses, are (1) Determine if Vit D affects corneal epithelial differentiation; (2) Examine the effects VDR knockout on corneal epithelial wound healing; and (3) Determine how Vit D is transported into the tear fluid. For Aim 1, in-vivo experiments will examine differentiation markers in Vit D Receptor (VDR) knockout mice. In-vitro experiments will examine differentiation markers in cultured corneal epithelial cells exposed to different concentrations of 25(OH)D3 and 1,25(OH)2D3. We will also examine the role of occludin and other Vit D metabolites in corneal epithelial physiology. For Aim 2, corneal epithelial wound healing will be examined in -/- and -/+ VDR knockout mice at different times after birth. Identical groups will be fed a special replenishment diet and corneal epithelial wound healing will be examined. Junctional complexes and differentiation markers will also be examined in all of these mice. In Aim 3, lacrimal fluid wil be collected from the rabbits fed a normal or high Vit D diet and Vit D metabolites will be measured. Vit D binding protein as well as megalin and cubilin (additional significant Vit D protein carriers) will be assayed in all samples to determine possible Vit D transport pathways into the ocular compartments. PUBLIC HEALTH RELEVANCE: It is estimated that more than 50% of the United States population has vitamin D (Vit D) insufficiency (2, 4, 5). Of the tens of thousands of published studies devoted to Vit D, less than a handful have examined Vit D in the cornea and anterior segment and none of these measured Vit D concentrations, metabolism, or activity at those sites. This work will address some of these topics.