Regulation of Osteoclastogenesis by Calcium

钙对破骨细胞生成的调节

基本信息

批准号：
8735616
负责人：
Harry C. Blair
金额：
$ 32.39万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-16 至 2018-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8735616
关键词：
Affect Animals Ants Arthritis Binding Bone Resorption Calcium Calcium Channel Calcium Signaling Cell fusion Cells Clinical Complex Development Disease Figs - dietary Gene Expression Genes Genetic Transcription Human ITAM ITPR1 gene In Vitro Knock-out Knockout Mice Life Malignant Neoplasms Measurement Measures Mediating Mediator of activation protein Minerals Modeling Molecular Molecular Conformation Mus Normalcy Osteoclasts Osteoporosis Pathologic Pathway Analysis Pathway interactions Pattern Postmenopausal Osteoporosis Process Proteins Regulation Rheumatoid Arthritis Role Signal Pathway Signal Transduction TNFSF11 gene Tissues Work bone cell type cytokine design extracellular in vivo mortality novel osteoclastogenesis precursor cell public health relevance receptor response skeletal skeletal abnormality tartrate-resistant acid phosphatase

项目摘要

DESCRIPTION (provided by applicant): We will study regulation of osteoclastogenesis by calcium, focusing on a recently discovered calcium-release activated calcium channel pathway. Activation of osteoclast resorption is the final common pathway in pathologic bone destruction, and Ca2+ signaling is critical to RANKL-stimulated osteoclastogenesis. In osteoclast precursors, Ca2+ signals activate the transcriptional regulator NFATc1, which drives expression of osteoclast-specific proteins. Other effects of NFATc1 include a role, now poorly defined, in fusion of osteoclast precursors. However, the the spatial and temporal patterns of Ca2+ fluxes regulating differentiation were largely unknown. We identified the calcium release activated calcium channel Orai1 as a mediator of extracellular Ca2+ influx in osteoclast precursors in vitro. In human osteoclast differentiation in vitro, we showed that absence or inhibition of Orai1 blocked the formation of mature multinucleated osteoclasts and greatly reduced bone mineral resorption. Studies of Orai1-/- mice showed similar changes in vivo, with associated with skeletal abnormalities, confirming the importance of Orai1 in bone regulation. We hypothesize that Orai1 mediates critical Ca2+ signals that regulate osteoclast formation by controlling the fusion of precursor cells. In Aim 1 we will define in vivo the function of Orai1 in osteoclast precursors. We will generate a mouse allowing tissue-selective deletion of Orai1 from osteoclast precursors. This will avoid pitfalls of complete Orai1 deficiency, which affects multiple cell type, leading to early mortality. We will compare the skeletal effects of a novel tissue-selective Orai1 knockout mouse to those of the global Orai1 knockout, to distinguish direct from indirect effects of Orai1 on osteoclastogenesis and skeletal regulation. We will analyze the skeletal effects of selective Orai1 deficiency in osteoclast precursors in vivo and identify abnormalities of differentiation caused by loss of Orai1 from osteoclast precursors. In Aim 2 we will determine the mechanisms by which Orai1 calcium signals modulate osteoclast precursor gene transcription to regulate osteoclast formation, particularly cell fusion. This work will include studies defining Orai1-dependent Ca2+ fluxes in response to osteoclast differentiation signals via measurement of Ca2+ oscillations with respect to osteoclast differentiation and activity in wild type and Orai1 deficient cells. Pathway analysis will, further, define NFATc1-regulated gene expression as a function of Ca2+ activation in Orai1-deficient and WT cells, and we will identify and analyze fusion- related genes dependent on Orai1. These studies will define mechanisms through which Orai1 regulates osteoclastogenesis, which will guide the development of new strategies to control pathologic osteoclastic activity.

描述（由申请人提供）：我们将研究钙对骨细胞生成的调节，重点是最近发现的钙释放激活的钙通道途径。破骨细胞吸收的激活是病理骨破坏的最终常见途径，Ca2+信号传导对于RANKL刺激的破骨细胞生成至关重要。在破骨细胞前体中，Ca2+信号激活转录调节剂NFATC1，该调节剂NFATC1驱动破骨细胞特异性蛋白的表达。 NFATC1的其他影响包括在破骨细胞前体融合中的作用，现在定义不明。但是，调节分化的Ca2+通量的空间和时间模式在很大程度上是未知的。我们确定钙释放活化的钙通道ORAI1是体外破骨细胞前体中细胞外Ca2+流入的介体。在人类破骨细胞分化的体外分化中，我们表明ORAI1的缺失或抑制阻止了成熟的多核破骨细胞的形成，并大大降低了骨矿物质的吸收。 ORAI1 - / - 小鼠的研究表现出类似的体内变化，与骨骼异常有关，证实了Orai1在骨调节中的重要性。我们假设Orai1介导了通过控制前体细胞融合来调节破骨细胞形成的关键Ca2+信号。在AIM 1中，我们将在体内定义Orai1在破骨细胞前体中的功能。我们将产生一种小鼠，允许从破骨细胞前体中的组织选择性缺失ORAI1。这将避免完全ORAI1缺乏的陷阱，这会影响多个细胞类型，从而导致早期死亡。我们将比较一种新型组织选择性Orai1敲除小鼠与全球Orai1敲除的骨骼效应，以区分Orai1对破骨细胞生成和骨骼调节的间接影响。我们将分析选择性ORAI1缺乏症在体内破骨细胞前体中的骨骼效应，并确定由破骨细胞前体失去Orai1引起的分化异常。在AIM 2中，我们将确定ORAI1钙信号调节破骨细胞前体基因转录以调节破骨细胞形成，尤其是细胞融合的机制。这项工作将包括通过测量野生型和ORAI1缺陷细胞中的破骨细胞分化以及活性的CA2+振荡来响应破骨细胞分化信号来定义ORAI1依赖性Ca2+通量的研究。途径分析将进一步将NFATC1调节的基因表达定义为ORAI1缺陷型和WT细胞中Ca2+激活的函数，我们将识别和分析依赖于Orai1的融合相关基因。这些研究将定义ORAI1调节破骨细胞生成的机制，该机制将指导发展新策略以控制病理破坏性肿瘤活性。