Alternative NF-KB as a negative regulator of osteogenesis

替代 NF-KB 作为成骨负调节剂

• 批准号: 9124589
• 负责人: Jennifer Lynn Davis
• 金额: $ 3.02万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9124589
• 关键词: Address; Adult; Affect; Alleles; Anabolism; Animals; BMP2 gene; Biology; Biomechanics; Bone Marrow; Bone Matrix; Bone Resorption; Bone remodeling; Cell Lineage; Cells; Complex; Coupled; Disease; Equilibrium; Exhibits; Family; Fracture; Future; Gene Expression; Genetic Recombination; Growth; Homeostasis; Hormonal; Human; In Vitro; Individual; Injection of therapeutic agent; Intervention; Lead; LoxP-flanked allele; Malignant Neoplasms; Mechanical Stress; Mechanics; Modeling; Molecular; Monitor; Mus; NF-kappa B; Organ; Osteoblasts; Osteoclasts; Osteocytes; Osteogenesis; Osteoporosis; Ovariectomy; PTH gene; Pathology; Pathway interactions; Phenotype; Play; Process; Protocols documentation; Regulation; Reporting; Role; Signal Pathway; Signal Transduction; Skeleton; Staging; Stimulus; Stress; Stromal Cells; Testing; Time; Transgenic Mice; base; bone; bone loss; bone mass; bone quality; cell type; design; gain of function; in vivo; insight; microCT; mineralization; mouse model; mutant; osteoblast differentiation; osteogenic; overexpression; public health relevance; research study; response; transcription factor

 DESCRIPTION (provided by applicant): Regulation of bone mass is a highly complex process involving the coordinated actions of osteoblasts (OBs), osteoclasts, and osteocytes. Aberrant signaling is manifest in a host of disease states such as osteoporosis, fracture, and cancer. This proposal will examine for the first time the direct role of alternative NF-κB in the OB lineage during basal bone homeostasis and under stress conditions. Transgenic mouse models will be used to modulate alternative NF-κB in the OB lineage using Osx-Cre. Upon induction of alternative NF-κB, NIK phosphorylates IKKa. As a NIKf/f model does not currently exist, Osx-Cre;IKKaf/f animals (cKO) will be used to examine inhibition of alternative NF-κB in the OB lineage. NIKdeltaT3 (NT3) is a constitutively active allele of NIK placed downstream of a floxed STOP cassette. Recombination with Osx-Cre will activate alternative NF-κB in the OB lineage (cACT). In Aim 1a, microCT, histomorphometric and mechanical strength analyses of control, cKO, and cACT mice will establish how loss or activation of the alternative NF-κB pathway in the OB lineage impacts basal bone phenotype. In Aim 1b, the in vitro OB phenotype of cKO and cACT cultures will be compared to control. Addition of known activators (Wnt3a, BMP2) of osteogenesis will delineate possible stage-specific effects of alternative NF-κB and interactions with various osteogenic signals. In Aim 2, the response to either pathological mechanical or hormonal anabolic stimuli will be evaluated in the same animals used in Aim 1. Aim 2a will use an in vivo loading protocol to monitor OB lineage cell response to mechanical stress. In Aim 2b, the ability of OB lineage cells to respond to hormonal stimulus will be evaluated by treating animals with intermittent parathyroid hormone, a currently used osteoporosis therapy. Bone formation rate and bone mass will be quantified in both Aims 2a and 2b to assess how loss or activation of the alternative NF-κB pathway influences the ability of OB lineage cells to respond to these anabolic stimuli. Thus, both in vivo and in vitro experiments will test the overarching hypothesis that alternative NF-κB is a negative regulator of OB lineage differentiation and/or function. The results of this proposal will provide valuable insight into the molecular mechanisms of osteogenesis, which may help guide future interventions to treat bone-related pathologies.

 描述（申请人提供）：骨量调节是一个高度复杂的过程，涉及成骨细胞（OB）、破骨细胞和骨细胞的协调作用。异常信号传导在许多疾病状态中是明显的，如骨质疏松症、骨折和癌症。本研究将首次研究在基底骨稳态和应激条件下，替代性NF-κB在OB谱系中的直接作用。转基因小鼠模型将用于使用Osx-Cre调节OB谱系中的替代NF-κB。在诱导替代NF-κB后，NIK磷酸化IKKa。由于目前不存在NIKf/f模型，因此将使用Osx-Cre;IKKaf/f动物（cKO）检查OB谱系中替代NF-κB的抑制。NIKdeltaT 3（NT 3）是位于floxed STOP盒下游的NIK的组成型活性等位基因。用Osx-Cre修饰将激活OB谱系（cACT）中的替代NF-κB。在目标1a中，对照、cKO和cACT小鼠的microCT、组织形态计量学和机械强度分析将确定OB谱系中替代NF-κB通路的缺失或激活如何影响基底骨表型。在目标1b中，将cKO和cACT培养物的体外OB表型与对照进行比较。加入已知的成骨激活剂（Wnt 3a，BMP 2）将描述替代NF-κB的可能阶段特异性作用以及与各种成骨信号的相互作用。在目标2中，将在目标1中使用的相同动物中评价对病理性机械或激素合成代谢刺激的反应。目的2a将使用体内加载方案来监测OB谱系细胞对机械应力的响应。在目标2b中，将通过用间歇性甲状旁腺激素（一种目前使用的骨质疏松症疗法）治疗动物来评价OB谱系细胞对激素刺激的反应能力。将在目标2a和2B中定量骨形成率和骨量，以评估替代NF-κB途径的缺失或激活如何影响OB谱系细胞对这些合成代谢刺激的反应能力。因此，体内和体外实验都将检验替代性NF-κB是OB谱系分化和/或功能的负调节剂的总体假设。该提案的结果将为骨生成的分子机制提供有价值的见解，这可能有助于指导未来的干预措施来治疗骨相关的病理。