MVNP, p62P392L and IL-6 in the Pathogenesis of PD

MVNP、p62P392L 和 IL-6 在 PD 发病机制中的作用

• 批准号: 7809011
• 负责人: Garson DAVID ROODMAN
• 金额: $ 61.03万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-28 至 2011-09-27
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7809011
• 关键词: Affect; Alkaline Phosphatase; Binding; Bone Development; Bone Diseases; Bone Resorption; Bone remodeling; Breeding; Calcium; Cell Nucleus; Cells; Characteristics; Cholecalciferol; Coupled; Development; Environmental Risk Factor; Genetic; Grant; Hyperactive behavior; In Vitro; Innate Bone Remodeling; Interleukin-6; Lesion; MAP Kinase Gene; MAPK14 gene; Marrow; Measles virus; Measles virus nucleocapsid protein; Mus; Nuclear; Osteoblasts; Osteoclasts; Osteogenesis; Paget' s Disease; Pathogenesis; Pathway interactions; Patients; Phenotype; Phosphorus; Physiological; Play; Production; Reporting; Research Design; Role; Signal Pathway; Site; TNFSF11 gene; Testing; Transgenic Mice; Vitamin D; Vitamin D3 Receptor; Whole Blood; absorption; bone; bone health; calcium phosphate; in vivo; insight; promoter; protein expression

DESCRIPTION (provided by applicant): Paget's disease (PD) is the second most common bone disease, affecting 1-2 million patients in the U.S. PD is one of the most exaggerated examples of normal bone remodeling and is a paradigm for studies of bone remodeling. Both genetic and environmental factors have been implicated in the pathogenesis of PD, and our current RO1 grant is examining the interaction of these two components in the development of PD in vivo. The primary cell involved in the pathogenesis of PD is the osteoclast (OCL). OCLs from PD patients have a distinct phenotype. They express measles virus nucleocapsid protein (MVNP), are hyper-responsive to 1,25-(OH)2D3 and RANKL, produce high levels of IL-6 and TAFII-17, a coactivator of VDR, and have an increased bone resorbing capacity per OCL. Importantly, targeted expression of MVNP to the OCL lineage in transgenic mice using the TRAP promoter (TRAP-MVNP mice) results in development of pagetic bone lesions and OCLs in these mice. However, the role that vitamin D receptor (VDR) hyperactivity plays in the development of PD is still unclear. We previously reported that TAFII-17 is induced by MVNP, and increased TAFII-17 levels are present in PD patients. Increased TAFII-17 levels result in hyper- responsivity of OCL precursors to 1,25-(OH)2D3. However, increased expression of TAFII-17 by itself is not sufficient to induce pagetic OCL in vitro but requires addition of IL-6 to the cultures. Importantly, MVNP cannot induce pagetic OCL in the absence of VDR. We found that binding of 1,25-(OH)2D3 to VDR in OCL precursors expressing MVNP increases activation of distinct signaling pathways (e.g. p38 and PLC32), which either increase IL-6 production, OCL formation or nuclear number per OCL. Objective/Hypothesis: It is our hypothesis that environmental factors implicated in the pathogeneses of PD, such as measles virus (MV), increase IL-6 levels in OCL precursors, which in turn increase TAFII17 expression. The increased expression of TAFII-17 enhances VDR activity at physiologic concentration of 1,25-(OH)2D3, that in turn activates both the p38 MAPK and PLC32/CREB pathways to increase IL-6 production, OCL numbers, nuclei per OCL and OCL bone resorption capacity. Further, elevated local levels of IL-6 induce normal OCL precursors to become hyper-responsive to vitamin D and form hyper- multinucleated OCL with increased bone resorbing capacity in the presence of vitamin D in vivo. To test this hypothesis we will pursue the following specific aims. Specific Aim 1: Determine if increased VDR activity in combination with high local levels of IL-6 in OCL precursors results in formation of hyper-multinucleated OCLs and increased bone resorption in vivo. Specific Aim 2: Determine if administration of vitamin D3 to TRAP-IL-6 mice results in formation of hyper- multinucleated OCLs and increased bone resorption. Study Design: Aim 1: We will characterize the effects of increased expression of TAFII-17 in the absence or presence of elevated IL-6 in OCL on pagetic OCL formation in vivo by analyzing TRAP-TAFII-17, TRAP-IL-6, and TRAP- TAFII-17/IL-6 (generated by breeding the first two lines) mice (followed over 18 months) for development of bone lesions, abnormal OCL activity, excessive new bone formation and other features of PD that we have reported in our TRAP-MVNP mice. We will compare these results to those obtained from TRAP-MVNP and WT mice. Aim 2: We will administer 1,25-(OH)2D3 (0.1 <g/kg IP per day) or vehicle daily to TRAP-IL-6 mice for 4 weeks, and determine if these mice develop pagetic OCL and bone lesions characteristic of PD in vivo by histomorphometry analysis as well as analysis of whole blood ionized calcium, phosphorus, alkaline phosphatase, and Pyd levels. Impact: This proposal will test for the role of increased VDR activity in the development of PD-OCL and bone lesions in vivo and if co-expression of increased levels of TAFII-17 and IL-6 in normal OCL precursors is sufficient to induce hyper-multinucleated OCLs characteristic of PD when mice are treated with vitamin D3.

描述(申请人提供)：Paget病(PD)是第二种最常见的骨骼疾病，在美国有100-200万患者。PD是正常骨重建最夸张的例子之一，是骨重建研究的范例。遗传和环境因素都与帕金森病的发病机制有关，我们目前的RO1资助正在研究这两种成分在体内帕金森病发生发展中的相互作用。参与帕金森病发病的主要细胞是破骨细胞(OCL)。PD患者的OCL具有明显的表型。它们表达麻疹病毒核衣壳蛋白(MVNP)，对1，25-(OH)2D3和RANKL高度反应，产生高水平的IL-6和VDR的共激活因子TAFII-17，并增加OCL的骨吸收能力。重要的是，在使用TRAP启动子的转基因小鼠(TRAP-MVNP小鼠)中，MVNP定向表达到OCL谱系中会导致这些小鼠出现Pagtic骨损伤和OCL。然而，维生素D受体(VDR)过度活跃在帕金森病发生发展中的作用尚不清楚。我们以前报道过TAFII-17是由MVNP诱导的，在PD患者中存在TAFII-17水平升高。TAFII-17水平升高导致OCL前体对1，25-(OH)2D3的高反应性。然而，增加TAFII-17的表达本身并不足以在体外诱导Pagtic OCL，而需要在培养中添加IL-6。重要的是，在缺乏VDR的情况下，MVNP不能诱发Pagtic OCL。我们发现，在表达MVNP的OCL前体中，1，25-(OH)2D3与VDR的结合增加了不同信号通路(如p38和PLC32)的激活，这些信号通路要么增加IL-6的产生，要么增加OCL的形成，要么增加OCL的核数量。目的/假设：我们的假设是，与帕金森病发病有关的环境因素，如麻疹病毒(MV)，会增加OCL前体细胞中IL-6的水平，进而增加TAFII17的表达。当生理浓度为1，25-(OH)2D3时，TAFII-17的表达增强VDR的活性，进而激活p38MAPK和PLC32/CREB通路，增加IL-6的产生、OCL数量、OCL的核数和OCL的骨吸收能力。此外，局部IL-6水平的升高会诱导正常的OCL前体细胞对维生素D产生高反应性，并在体内形成超多核OCL，并增加骨吸收能力。为了验证这一假设，我们将追求以下具体目标。具体目的1：确定OCL前体细胞中VDR活性增加和局部IL-6水平升高是否导致体内超多核OCL的形成和骨吸收的增加。特定目的2：确定给予TRAP-IL-6小鼠维生素D3是否导致超多核OCL的形成和骨吸收增加。研究设计：目的1：我们将通过分析TRAP-TAFII-17、TRAP-IL-6和TRAP-TAFII-17/IL-6(前两个品系饲养产生)小鼠(18个月以上)的骨损伤、OCL活性异常、过度新骨形成和我们在TRAP-MVNP小鼠中报道的其他特征，来表征在OCL中没有或存在IL-6升高的情况下TAFII-17的表达增加对活体Pagtic OCL形成的影响。我们将把这些结果与TRAP-MVNP和WT小鼠的结果进行比较。目的：给TRAP-IL-6小鼠每天ip 1，25-(OH)2D3(0.1&1g/kg)或赋形剂4周，通过组织形态计量学分析和全血钙、磷、碱性磷酸酶及PYD水平的分析，确定这些小鼠是否发生了寻常型OCL和体内帕金森病特有的骨病变。影响：这项提案将测试VDR活性增加在体内PD-OCL和骨损伤发展中的作用，以及当小鼠接受维生素D3治疗时，正常OCL前体中TAFII-17和IL-6水平的共同表达是否足以诱导具有PD特征的超多核OCL。