ERK Signaling in Inflammatory Bone Loss

炎症性骨丢失中的 ERK 信号传导

• 批准号: 8793760
• 负责人: Francis Young-In Lee
• 金额: $ 48.07万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8793760
• 关键词: Address; Adverse effects; Affect; Anti-Inflammatory Agents; Anti-inflammatory; Biology; Bone Resorption; CSF1 gene; Calcium Oscillations; Caspase; Caspase-1; Cells; Data; Disease; Dissociation; Economic Burden; Elderly; Exhibits; Fracture; Functional disorder; Future; Gene Expression; Genes; Goals; Healed; Health; Health Care Costs; Human; Image; Immunoprecipitation; In Vitro; Inflammation; Inflammatory; Intervention; Knockout Mice; Knowledge; Macrophage Activation; Malignant Neoplasms; Measures; Mediating; Medical; Messenger RNA; Metastatic Neoplasm to the Bone; Methods; Molecular; Mus; Musculoskeletal Diseases; Osteoclasts; Osteolysis; Osteolytic; Osteoporosis; PTGS2 gene; Parents; Pathogenesis; Pathologic; Pathway interactions; Pharmacologic Substance; Play; Population; Process; Production; Proteins; Proteomics; Recombinants; Recruitment Activity; Research; Role; Signal Transduction; Site; Staining method; Stains; TNF gene; TNFSF11 gene; Testing; Therapeutic; Therapeutic Effect; Time; Tissues; Topical application; Translations; Treatment Efficacy; Wages; Work; base; bone; bone loss; bone mass; calreticulin; cathepsin K; clinical application; clinically relevant; cytokine; disability; healing; in vivo; in vivo Model; inflammatory bone resorption; inhibitor/antagonist; innovation; interest; interleukin-1beta-converting enzyme inhibitor; liquid chromatography mass spectrometry; mRNA Expression; macrophage; meetings; molecular imaging; monocyte; new therapeutic target; novel; osteoclastogenesis; pathologic bone resorption; prevent; protective effect; rapid growth; research study; screening; success; therapeutic target; transcription factor

DESCRIPTION (provided by applicant): Pathologic bone resorption due to osteoporosis, fragility fractures and cancers pose a significant economic burden to healthcare costs and lost wages. These osteolytic diseases share a similar pathophysiology, so understanding the mechanisms of bone resorption greatly aids the search for treatment options. The need to develop therapeutic options with minimal side effects is strong, but requires greater elucidation into the mechanisms of inflammatory bone resorption. We previously confirmed the role of pERK1/2 on the production of pro-osteoclastogenic cytokines such as RANKL, MCSF, COX-2 and IL-1b. Then, we focused on a seemingly overlooked target for inhibiting osteoclastogenesis, IL-1b, an inflammatory cytokine implicated in pathologic osteolysis. An IL-1b Converting Enzyme (ICE, or Caspase-1) inhibitor showed dramatic inhibition of osteoclastogenesis. However, Caspase-1inhibitor -/- osteoclast precursors did not demonstrate impaired osteclastogenesis, implying off-target components to this osteolysis-inhibition mechanism. Subsequently, our proteomic screening efforts yielded us Calregulin (CRG) as our osteoclastogenic-inhibition focus. Subsequent in vitro and in vivo studies met with great success as administration of recombinant human Calregulin (rhCRG) demonstrated inhibition of osteoclastogenesis in vitro in osteoclast-precursor macrophages and inhibition of LPS-induced osteolysis in vivo. rhCRG inhibited NFATc1 among several osteoclastogenic transcription factors. The new direction of this competitive renewal is to test a novel hypothesis that Calregulin inhibits osteoclastogenesis in vitro and inflammation-associated osteoclastogenesis in vivo. Therefore, our aims for this proposal employ 3 parallel Aims to determine mechanisms and therapeutic translation in vitro and in vivo. Aim 1 will determine mechanisms by which rhCRG interferes with key osteoclastogenic pathways such as NFATc1 in vitro. We will determine whether rhCRG interferes with osteoclastogenic gene expression, calcium oscillation, NFATc1 activity, and bone resorption. In addition, we will examine whether rhCRG/NFATc1 functionally interact with 6 candidate proteins which we identified on Liquid Chromatography/Mass Spectrometry of His-CRG pulled-down proteins. Aim 2 will determine whether CRG regulates macrophage activation in inflammatory osteolysis in vitro and in vivo. Macrophage activation of M1 or M2 plays pro- and anti-inflammatory roles, respectively. By determining the CRG's role in the polarization of M2 macrophages and inhibition of M1, we will establish an alternative mechanism. Aim 3 will establish a therapeutic role of topical rhCRG in inflammatory osteolysis in vivo. We will induce osteolysis with clinically relevant RANKL, TNF and LPS to determine whether rhCRG can prevent or treat inflammation-associated osteoclastogenesis and bone resorption. We will measure bone protective effects of rhCRG using Cathepsin K molecular imaging, dynamic bone histomorphometry and TRAP staining. Overall impact is high in that we expect to unravel novel anti-osteoclastogenic mechanisms and therapeutic promise of rhCRG in the context of inflammatory osteolysis.

描述（由申请人提供）：骨质疏松症、脆性骨折和癌症导致的病理性骨吸收对医疗费用和工资损失造成了重大经济负担。这些溶骨性疾病具有相似的病理生理学，因此了解骨吸收的机制有助于寻找治疗方案。开发具有最小副作用的治疗方案的需求是强烈的，但需要进一步阐明炎性骨吸收的机制。我们先前证实了pERK 1/2在前破骨细胞生成细胞因子如RANKL、MCSF、考克斯-2和IL-1b的产生中的作用。然后，我们集中在一个似乎被忽视的抑制破骨细胞生成的靶点，IL-1b，一种与病理性骨质溶解有关的炎性细胞因子。IL-1b转化酶（ICE或Caspase-1）抑制剂显示出对破骨细胞生成的显著抑制作用。然而，Caspase-1抑制剂-/-破骨细胞前体并没有表现出破骨细胞生成受损，这意味着这种溶骨抑制机制的脱靶成分。随后，我们的蛋白质组学筛选工作产生了我们钙调蛋白（CRG）作为我们的破骨细胞生成抑制的重点。随后的体外和体内研究取得了巨大成功，因为重组人钙调蛋白（rhCRG）的给药证明了体外破骨细胞前体巨噬细胞中破骨细胞生成的抑制和体内LPS诱导的骨质溶解的抑制。在几种破骨细胞生成转录因子中，rhCRG抑制NFATc 1。这种竞争性更新的新方向是测试一种新的假设，即钙调蛋白抑制体外破骨细胞生成和体内炎症相关的破骨细胞生成。因此，我们的这一建议的目的采用3个平行的目标，以确定机制和治疗翻译在体外和体内。目的1将确定rhCRG在体外干扰关键破骨细胞生成途径如NFATc 1的机制。我们将确定rhCRG是否干扰破骨细胞基因表达、钙振荡、NFATc 1活性和骨吸收。此外，我们将研究rhCRG/NFATc 1是否与我们在His-CRG下拉蛋白的液相色谱/质谱分析中鉴定的6个候选蛋白质在功能上相互作用。目的2将确定CRG是否调节巨噬细胞活化的炎症性骨溶解在体外和体内。M1或M2的巨噬细胞活化分别起促炎和抗炎作用。通过确定CRG在M2巨噬细胞极化和抑制M1中的作用，我们将建立另一种机制。目的3建立局部应用rhCRG对炎性骨溶解的治疗作用。我们将用临床相关的RANKL、TNF和LPS诱导骨质溶解，以确定rhCRG是否可以预防或治疗炎症相关的破骨细胞生成和骨吸收。我们将使用组织蛋白酶K分子成像、动态骨组织形态计量学和TRAP染色来检测rhCRG的骨保护作用。总体影响是高的，因为我们期望在炎性骨质溶解的背景下解开新的抗破骨细胞生成机制和rhCRG的治疗前景。