NELL-1, A Cbfa1 DOWNSTREAM TARGET, IN BONE FORMATION

NELL-1，Cbfa1 下游靶标，在骨形成中

• 批准号: 6899379
• 负责人: KANG TING
• 金额: $ 34.76万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6899379
• 关键词: bone development; cell differentiation; cell growth regulation; cell type; gel mobility shift assay; genetic regulatory element; genetic transcription; laboratory mouse; mesenchyme; osteoblasts; polymerase chain reaction; protein binding; protein quantitation /detection; protein structure function; site directed mutagenesis; southern blotting; tissue /cell culture; western blottings

DESCRIPTION (provided by applicant): Cbfa1 mediates mesenchymal cell commitment to an osteochondroprogenitor lineage and supports continued differentiation and function in committed osteoblasts. However, definitive downstream mediators of Cbfa1 that critically impact osteoblast differentiation and bone formation have yet to be described. Ne1 (a protein strongly expressed in neural tissue encoding epidermal growth factor like domain) was first identified in chicken embryos. The preliminary data shows that Nell-1 (Ne1-like molecule-1) in vivo is associated with normal and pathological suture fusion, while Nell- 1 in vitro is associated with increased osteoblast differentiation and mineralization. Furthermore, the human Nell-1 promoter contains three osteoblast-specific cis-acting element 2 (OSE2) response elements for Cbfa1 and that Nell-1 overexpression mice can functionally compensate for some aspects of intramembranous bone deficiency in heterozygous Cbfa1 deficient mice. These data have led to the hypothesis that Nell-1 is a central downstream target of Cbfa1 that supports continued differentiation and function in committed osteoblasts and proper Nell-1 expression is required for normal membranous bone growth and formation. To test this hypothesis, this proposal will analyze the following: 1) the regulation of Nell-1 transcription by Cbfa1 during osteoblast formation and function and by Cbfa1 through OSE2 response elements; 2) the functional compensation of Cbfa1 deficiency by Nell-1 in vivo and in vitro; and 3) the effects of targeted Nell-1 disruption in vivo. If Nell-1 is proven to be a critical mediator of intramembranous bone formation, it can further aid in understanding the development of intramembranous vs. endochondral bone. In addition, since sutures are major sites of calvarial intramembranous bone growth the close association of Nell-1 with calvarial sutures can also impact present knowledge on cranial vault development. Lastly, Nell-1 can be tremendously useful as a clinical tool to inhibit or induce intramembranous bone growth, with the secretory nature of Nell-1 making this utility even more feasible. Controlled inhibition of intramembranous bone growth by anti-Nell-1 strategies in CS patients may offer a less invasive, biologically based, therapeutic alternative to radical surgical craniofacial reconstruction. In addition, the ability of Nell- 1 to induce calvarial osteoblast differentiation may be clinically applied to situations where membranous bone formation and regeneration are desirable (e.g., cleft lip or cleft palate repair and craniofacial distraction osteogenesis).

描述（由申请人提供）：Cbfa1介导间充质细胞向骨软骨祖细胞谱系的分化，并支持成骨细胞的持续分化和功能。然而，Cbfa1的下游介质对成骨细胞分化和骨形成的决定性影响尚未被描述。Ne1（一种在神经组织中强烈表达的编码表皮生长因子样结构域的蛋白）首次在鸡胚胎中被发现。初步数据显示，体内的Nell-1 （ne1样分子-1）与正常和病理缝合融合有关，而体外的Nell-1则与成骨细胞分化和矿化增加有关。此外，人类Nell-1启动子含有3个Cbfa1的成骨细胞特异性顺式作用元件2 （OSE2）应答元件，并且Nell-1过表达小鼠可以在功能上补偿杂合子Cbfa1缺陷小鼠的膜内骨缺乏的某些方面。这些数据导致假设Nell-1是Cbfa1的中心下游靶点，支持成骨细胞的持续分化和功能，正常的膜性骨生长和形成需要适当的Nell-1表达。为了验证这一假设，本提案将分析以下内容：1)Cbfa1在成骨细胞形成和功能过程中对Nell-1转录的调控，以及Cbfa1通过OSE2应答元件对Nell-1转录的调控；2) Nell-1在体内外对Cbfa1缺乏症的功能代偿；3)体内靶向破坏Nell-1的影响。