Cell Cycle Mechanisms of PTH/PTHrP Action in Osteoblasts

成骨细胞中 PTH/PTHrP 作用的细胞周期机制

• 批准号: 6956254
• 负责人: Nabanita S Datta
• 金额: $ 7.63万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-07 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6956254
• 关键词: AP1 protein; RNA interference; bone metabolism; cell cycle; cell differentiation; cyclins; hormone regulation /control mechanism; immunocytochemistry; laboratory mouse; osteoblasts; parathyroid hormone related protein; parathyroid hormones; protooncogene; tissue /cell culture

DESCRIPTION (provided by applicant): The physiological process of bone remodeling is tightly regulated by local and endocrine factors, and any disturbance will lead to various skeletal diseases including craniofacial deformities and orthopedic disorders. Parathyroid hormone (PTH) has been shown clinically to induce bone formation. However the exact biological mechanisms of bone formation and regeneration by PTH remain elusive. Our broad, long-term goal is to identify molecular mechanisms of PTH/PTHrP (PTH related protein) action and bone turnover. Preliminary data suggest that cell cycle control may be a mechanism through which PTHrP impacts the life span and bone forming activity of osteoblasts. Down-regulation of cyclin D1 and elevated expression of JunB suggested involvement of AP-1 transcription factors to the cell cycle machinery of osteoblasts and led us to hypothesize that the action of PTHrP on cyclin D1 is developmental stage specific and associated with specific signal transduction mediators. Using in vitro and in vivo models, 2 specific aims will test this hypothesis. In specific aim 1 cyclin D1 promoter constructs and Luciferase assays will determine if PTHrP effects on cyclin D1 are dependent on osteoblast proliferation and differentiation stage. We will determine if JunB plays a role in modulating cyclin D1 expression utilizing RNA interference (RNAi) technology, over expression of dominant negative AP-1 protein (TAM 67) and CDK1. Specific Aim 2 will utilize a novel ectopic ossicle model system to verify cyclin D1 as 1 of the downstream mediators of PTHrP signaling in vivo. Finally, using immunohistochemistry, effects of PTHrP on cyclin D1 will be elucidated at various stages of osteoblast differentiation in vivo. The outcome of this investigation will advance our knowledge of PTH/PTHrP action in bone and contribute to understanding the anabolic actions of PTH. Identifying these mechanisms related to PTH action will have applicability as potential targets for new therapeutic agents for the treatment of diseases which adversely affect bone, including osteoporosis and periodontal diseases.

描述（申请人提供）：骨重建的生理过程受局部和内分泌因素的严格调节，任何干扰都会导致各种骨骼疾病，包括颅面畸形和骨科疾病。甲状旁腺激素（PTH）已被证明在临床上诱导骨形成。然而，骨形成和再生的PTH的确切生物学机制仍然是难以捉摸的。我们广泛的长期目标是确定PTH/PTHrP（PTH相关蛋白）作用和骨转换的分子机制。初步数据表明，细胞周期控制可能是PTHrP影响成骨细胞寿命和骨形成活性的机制。细胞周期蛋白D1的下调和JunB的表达升高表明AP-1转录因子参与成骨细胞的细胞周期机制，并使我们假设PTHrP对细胞周期蛋白D1的作用是发育阶段特异性的，并与特定的信号转导介质相关。使用体外和体内模型，2个特定目标将检验这一假设。在具体的目标1中，细胞周期蛋白D1启动子构建和荧光素酶测定将确定PTHrP对细胞周期蛋白D1的作用是否依赖于成骨细胞的增殖和分化阶段。我们将利用RNA干扰（RNAi）技术确定JunB是否在调节细胞周期蛋白D1表达、显性失活AP-1蛋白（TAM 67）和CDK 1的过度表达中发挥作用。具体目标2将利用一种新的异位小骨模型系统，以验证细胞周期蛋白D1作为PTHrP信号的下游介质在体内。最后，使用免疫组化，PTHrP对细胞周期蛋白D1的影响将在体内成骨细胞分化的各个阶段阐明。本研究的结果将促进我们对PTH/PTHrP在骨中作用的认识，并有助于理解PTH的合成代谢作用。鉴定与PTH作用相关的这些机制将具有作为用于治疗对骨有不利影响的疾病（包括骨质疏松症和牙周病）的新治疗剂的潜在靶标的适用性。