Mesenchymal Regulation of Jaw Bone Length

描述(申请人提供)：这是为加州大学旧金山分校的普通牙医Erin Ealba博士申请K08奖项。在头面部骨骼发育的基础科学研究中，艾尔巴博士正在确立自己作为一名年轻研究员的地位。这一K08奖项将为Ealba博士提供必要的支持，以实现以下目标：(1)成为发育、骨骼和颅面生物学方面的专家；(2)专注于教师技能；(3)提高她的手稿和拨款写作能力。为了实现这些目标，Ealba博士组建了一个指导团队，成员包括一名主要导师[颅面发育领域的权威Richard Schneider博士]、两名共同导师[专注于牙釉质矿化的领先牙医科学家Pamela DenBesten博士和转化生长因子？和三位合作者[Mary Nakamura博士、Ralph Marcucio博士和Ophir Klein博士]。Ealba博士的长期目标是发现新的基于分子的疗法来调节骨骼的长度和形状，以此作为解决颅面畸形非手术治疗需求的一种手段。目前这项研究的目标是通过了解构成面部和颌骨骨骼中所有元素的神经脊间充质(NCM)如何调节颌骨大小来实现这一目标。为了解决这个问题，我们在体内操纵了NCM，这是一个非常容易获得的胚胎种群。具体地说，我们将成熟较快的供体NCM移植到发育较慢的鸭子宿主中，从而产生嵌合QUCK；我们将较慢的供体鸭子NCM移植到相对较快的鹌鹑宿主中，产生嵌合二重体。利用鹌鹑和鸭子不同的发育程序提供了一种独特的方式来操纵NCM和邻近宿主组织之间的信号，并允许发现NCM依赖的过程。此外，所有的鹌鹑细胞都可以通过一种在鸭子中不存在的普遍存在的核标记来检测到。在已发表的工作和初步研究中，我们观察到NCM自主地同步和指导成骨诱导、增殖、分化、基质沉积、矿化和基质重塑。NCM如何完成如此复杂的任务，以及哪些因素足以复制这种现象，目前尚不清楚。可能的候选者可能包括转化生长因子-β2(TGF2)的成员和靶标，因为已知它们在成骨过程中发挥关键作用，并且它们的表达在嵌合体中发生变化。因此，我们假设通过调节转化生长因子的水平？NCM引导自己的成骨程序，并协调破骨细胞的活动来控制颌骨的长度。为了检验我们的假设，我们提出了三个互补且不相互依赖的具体目标。具体目标1将确定NCM使用TGF2信号来控制颌骨长度的程度。具体目标2将确定NCM在多大程度上依赖破骨细胞的行动来调节颌骨长度。具体目标3将确定NCM通过依赖于TGF2的RANKL/OPG信号影响破骨细胞活性和颌骨长度的程度。我们将使用增益和功能损失技术来识别赋予NCM控制颌长的能力的分子机制。这一提议的实验可以作为一个原则的证明，即可以设计出基于分子的疗法来治疗影响颌骨长度的疾病。此外，识别供体NCM转导其对破骨细胞等宿主细胞的作用的机制，对于修复和再生因创伤或疾病(如骨质疏松和骨坏死)而受伤的骨骼具有重要意义。我们希望我们的研究将为以生物学为基础的非手术方法治疗各种临床骨骼疾病提供基础。

DESCRIPTION (provided by applicant): This is an application for a K08 award for Dr. Erin Ealba, a general dentist at the University of California, San Francisco. Dr. Ealba is establishing herself as a young investigator in basic science research of craniofacial bone development. This K08 award will provide Dr. Ealba with the support necessary to accomplish the following goals: (1) to become an expert in developmental, skeletal, and craniofacial biology; (2) to focus on faculty skills; and (3) to enhance her manuscript and grant writing abilities. To achieve these goals, Dr. Ealba has assembled a mentoring team comprised of a primary mentor [Dr. Richard Schneider, an authority in the field of craniofacial development], two co-mentors [Dr. Pamela DenBesten, a leading dentist-scientist focused on enamel mineralization, and Dr. Tamara Alliston, an expert in TGF? signaling], and three collaborators [Dr. Mary Nakamura, Dr. Ralph Marcucio, and Dr. Ophir Klein]. Dr. Ealba's long-term goal is to discover novel molecular-based therapies for regulating the length and shape of bone as a means to address the need for non-surgical treatments of craniofacial malformations. The objective of the current study is to build toward this goal by understanding how neural crest mesenchyme (NCM), which forms all the elements in the facial and jaw skeletons, regulates jaw size. To address this issue, we manipulate in vivo the NCM, a highly accessible embryonic population. Specifically, we transplant faster- maturing quail donor NCM into a slower-developing duck host, which creates chimeric quck; and we transplant slower duck donor NCM into the relatively faster quail host, generating chimeric duail. Exploiting the divergent developmental programs of quail and duck provides a unique way to manipulate signaling between NCM and adjacent host tissues, and allows discovery of NCM-dependent processes. Also, all quail cells can be detected via a ubiquitous nuclear marker not present in duck. In published work and in preliminary studies, we observe that NCM autonomously synchronizes and directs osteogenic induction, proliferation, differentiation, matrix deposition, mineralization, and matrix remodeling. How NCM accomplishes such a complex task, and what factors are sufficient to replicate this phenomenon, is unknown. Likely candidates may include members and targets of the Transforming Growth Factor-Beta (TGF2) since they are known to play critical roles during osteogenesis, and their expression is altered in chimeras. Therefore, we hypothesize that by modulating levels of TGF? signaling, NCM directs its own osteogenic program and coordinates the activities of osteoclasts to control jaw length. To test our hypothesis, we propose three complementary and non-interdependent Specific Aims. Specific Aim 1 will determine the extent to which NCM employs TGF2 signaling to control jaw length. Specific Aim 2 will determine the extent to which NCM relies on the actions of osteoclasts to regulate jaw length. Specific Aim 3 will determine the extent to which NCM acts via TGF2-dependent RANKL/OPG signaling to affect osteoclast activity and jaw length. We will employ gain- and loss-of-function techniques to identify molecular mechanisms that endow NCM with the ability to control jaw length. Experiments from this proposal can serve as a proof-of-principle that molecular-based therapies can be devised to treat disorders that affect the length of the jaw. Moreover, identifying mechanisms through which donor NCM transduces its effects on host cells such as osteoclasts has implications for repair and regeneration of bones injured by trauma or diseases like osteoporosis and osteonecrosis. We are hopeful that our research will provide a foundation for biologically based, non-surgical methods to remedy a variety of clinical skeletal conditions.