1,25-Dihydroxyvitamin D and vitamin D Receptor Function in the Zebrafish Skeleton

描述(由申请人提供)：本申请的目的是利用斑马鱼Danio rerio作为模式生物，阐明1a，25-二羟基维生素D3(1a，25(OH)2D3)及其受体1a，25(OH)2D3受体(VDR)在其不参与跨上皮钙(Ca)和磷(PI)运输(“非经典”效应)的组织中的功能。我们最近发现，VDR在成年和发育中的斑马鱼Danio rerio的组织中表达。VDR存在于斑马鱼受精后早期发育中的软骨、骨、脑、眼和耳朵中。假设1a，25(OH)2D3和VDR通过调节骨形态发生蛋白及其受体的表达来改变斑马鱼软骨的发育和成熟。在目标1中，我们将研究1a，25(OH)2D3对Danio rerio骨骼发育的影响。我们将1a，25(OH)2D3应用于斑马鱼胚胎的发育中，以确定1a，25(OH)2D3是否改变了BMP及其受体的表达，BMP信号转导以及软骨发育和骨骼形态。在目标2中，我们将用反义吗啉寡核苷酸的方法来确定VDR在斑马鱼胚胎发育中的表达下调的效果。将评估VDR基因敲除对斑马鱼胚胎BMP及其受体表达、BMP信号转导以及软骨发育和骨骼形态的影响。意义：1a，25(OH)2D3及其受体在哺乳动物和人类中对钙转运的影响已被广泛研究。然而，与钙转运无关的影响还没有被详细研究。通过研究1a，25(OH)2D3和VDR通过BMP途径对斑马鱼胚胎骨骼发育的影响，我们将能够揭示1a，25(OH)2D3和VDR在软骨中的直接和非钙依赖的作用。我们还将建立斑马鱼作为一种新的模式生物，在其中研究1a，25(OH)2D3的影响。 与公众健康相关：维生素D对维持骨骼健康很重要。在这个项目中，我们将通过在发育中的斑马鱼模型生物中研究维生素及其受体如何在骨骼和软骨中发挥作用。通过研究活性维生素D及其受体对影响骨骼发育的生长因子的影响，我们将深入了解维生素在骨骼中的作用。这些信息将在预防随年龄增长而发生的骨丢失和治疗骨质疏松症方面有用。

DESCRIPTION (provided by applicant): The objective of this application is to elucidate the functions of 1a, 25-dihydroxyvitamin D3 (1a,25(OH)2D3) and its receptor, the 1a,25(OH)2D3 receptor (VDR), in tissues in which it does not have a role in trans-epithelial calcium (Ca) and phosphate (Pi) transport ("non-classical" effects) using the zebrafish, Danio rerio as a model organism. We have recently shown that the VDR is expressed in tissues of the adult and developing zebrafish, Danio rerio. The VDR is present in the developing cartilage, bone, brain, eye, and ear of the zebrafish early after fertilization. The hypothesis is that 1a,25(OH)2D3 and the VDR alter cartilage development and maturation in zebrafish by regulating the expression of bone morphogenetic proteins (BMPs) and BMP receptors. In aim 1 we will examine the effects of 1a,25(OH)2D3 on skeletal development in Danio rerio. We will administer 1a,25(OH)2D3 to developing zebrafish embryos to determine whether 1a,25(OH)2D3 administration alters BMP and BMP receptor expression, BMP signaling and cartilage development and skeletal morphology. In aim 2, we will determine the effects of "knocking down" VDR expression in the developing zebrafish embryo with anti-sense morpholino oligonucleotides methods. The effect of VDR knock- down on zebrafish embryo BMP and BMP receptor expression, BMP signaling and cartilage development and skeletal morphology will be assessed. Significance: The effects of 1a,25(OH)2D3 and its receptor have been extensively examined in mammals and humans with respect to their effects on Ca transport. Effects that are independent of Ca transport have not, however, been examined in detail. By examining the effects of 1a,25(OH)2D3 and the VDR on skeletal development in zebrafish embryos through BMP pathways, we will be able to delineate the direct and Ca-independent effects of 1a,25(OH)2D3 and the VDR in cartilage. We will also establish the zebrafish as a new model organism in which to examine the effects of 1a,25(OH)2D3. PUBLIC HEALTH RELEVANCE: Vitamin D is important for the maintenance of bone health. In this project we will examine how the vitamin and its receptor function in bone and cartilage by studying their effects in a model organism, the developing zebrafish. By examining the effects of active vitamin D and its receptor on growth factors that influence skeletal development we will gain insights into how the vitamin functions in bone. This information will be of use in preventing bone loss that occurs with aging and in treating osteoporosis.