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Regulation of Osteoclastogenesis by Calcium

钙对破骨细胞生成的调节

基本信息

批准号：
8609756
负责人：
Harry C. Blair
金额：
$ 33.38万
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/8609756
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

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 types, 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.

我们将研究钙对破骨细胞生成的调节，钙释放激活钙通道途径。破骨细胞再吸收的激活是最终的病理性骨破坏的共同途径，Ca 2+信号传导对RANKL刺激的骨细胞凋亡至关重要。破骨细胞生成在破骨细胞前体中，Ca 2+信号激活转录调节因子 NFATc 1，驱动破骨细胞特异性蛋白的表达。NFATc 1的其他作用包括：在破骨细胞前体融合中的作用，现在还不清楚。然而，空间和时间 Ca 2+通量调节分化的模式在很大程度上是未知的。我们确定了钙释放激活的钙通道Orai 1作为细胞外的介导剂，体外破骨细胞前体中的钙内流。在体外人破骨细胞分化中，我们表明 Orai 1的缺乏或抑制阻断了成熟多核破骨细胞的形成，减少骨矿物质吸收。对Orai 1-/-小鼠的研究显示出类似的体内变化，与骨骼异常相关，证实了Orai 1在骨调节中的重要性。我们假设Orai 1介导调节破骨细胞形成的关键Ca 2+信号，控制前体细胞的融合。在目标1中，我们将在体内定义Orai 1在破骨细胞前体中的功能。我们将生成一个小鼠允许从破骨细胞前体中组织选择性缺失Orai 1。这将避免以下缺陷： Orai 1完全缺乏，影响多种细胞类型，导致早期死亡。我们将比较一种新的组织选择性Orai 1基因敲除小鼠对整体Orai 1基因敲除小鼠的骨骼效应敲除，以区分Orai 1对破骨细胞生成和骨骼肌的直接和间接作用。调控我们将分析破骨细胞前体中选择性Orai 1缺乏对骨骼的影响，体内和鉴定由破骨细胞前体Orai 1缺失引起的分化异常。在目标2中，我们将确定Orai 1钙信号调节破骨细胞的机制前体基因转录调节破骨细胞形成，特别是细胞融合。这项工作将包括定义Orai 1依赖性Ca 2+通量响应破骨细胞分化信号的研究通过测量野生型破骨细胞分化和活性方面的Ca 2+振荡和Orai 1缺陷细胞。通路分析将进一步将NFATc 1调节的基因表达定义为一种新的途径。 Ca 2+激活在Orai 1缺陷和WT细胞中的功能，我们将鉴定和分析融合- 依赖Orai 1的相关基因。这些研究将确定Orai 1调节破骨细胞生成的机制，将指导控制病理性骨坏死活性的新策略的发展。