Functions of extracellular matrix proteins in dental and skeletal mineralization

细胞外基质蛋白在牙齿和骨骼矿化中的功能

基本信息

批准号：
9980842
负责人：
Brian Lee Foster
金额：
$ 35.5万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-08-01 至 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9980842
关键词：
Ablation Affect Amino Acids Animals Arginine Aspartic Acid Atomic Force Microscopy Binding Binding Sites Biological Bone Regeneration Bone Resorption Bone remodeling Cell Line Cells Cementoblast Cementogenesis Cementum Formation Collagen Collagen Fibril Complex Data Dental Dental Cementum Deposition Development Epithelial Epithelial Cells Epithelium Exhibits Exons Extracellular Matrix Proteins Fiber Fluorescence-Activated Cell Sorting Foundations Genes Genetically Engineered Mouse Glycine Goals Growth Factor Harvest Health Human Impaired healing In Vitro Integrin Binding Knock-out Knowledge Literature Location LoxP-flanked allele Maps Minerals Modeling Mus Mutate Mutation Natural regeneration Neural Crest Oral Osteoblasts Osteotomy Outcome Peptides Periodontal Diseases Periodontal Ligament Planet Earth Plant Roots Play Polyglutamic Acid Population Process Proteins Quality of life RGD (sequence)RNA Sequences Recombinants Role Root Resorption Signal Transduction Skeletal Development Systemic disease Testing Tissues Tooth Loss Tooth structure Topaz alveolar bone base bone bone cell bone healing bone sialoprotein burden of illness design experimental study healing human subject in vivo insight mineralization mouse model novel novel therapeutic intervention osteoprogenitor cell programs protein expression regenerative repaired selective expression skeletal tool transcriptome transcriptomics

项目摘要

PROJECT SUMMARY/ABSTRACT The periodontal complex, including cementum, periodontal ligament (PDL), and alveolar bone, is critical for tooth attachment and function. Periodontal diseases are among the most prevalent on earth, causing periodontal destruction and tooth loss, and affecting health and quality of life. Periodontal regeneration is possible, however, current therapies are unpredictable, few are truly regenerative, and many lack a biologic foundation. The path to regeneration remains unclear, in part, because origins and differentiation of cementoblasts (cells that produce cementum), and regulatory processes in cementogenesis, remain poorly understood. Bone sialoprotein (Ibsp gene; BSP protein) is a multifunctional extracellular matrix (ECM) protein associated with mineralized tissues, skeletal formation, and bone remodeling. Ibsp knockout (Ibsp-/-) mice feature absence of functional acellular cementum, PDL detachment, defective alveolar bone and cellular cementum mineralization, and alveolar bone resorption and tooth loss. The underlying mechanisms of BSP function remain unknown, although BSP harbors three functional domains, including a collagen-binding domain, polyglutamic acid (polyE) motifs that promote mineralization, and an arginine-glycine-aspartic acid (RGD) integrin-binding domain that initiates cell signaling. We propose that BSP is a unique candidate factor for studying cementum formation and alveolar bone healing because it is selectively expressed, essential for proper function, and operates by non-redundant mechanism(s) distinct from other growth factors. Based on our preliminary data, our hypotheses are that cementoblasts are BSP-expressing ectomesenchymal cells distinct from osteoblasts; BSP signals bone cells via the RGD domain in bone remodeling and directs mineral deposition onto collagen fibrils via the collagen-binding domain; and BSP promotes alveolar bone healing. These hypotheses will be tested by 3 specific aims: (1) To define the origin and transcriptome of cementoblasts using conditional ablation of Ibsp from ectomesenchymal vs. epithelial cell populations, and use endogenous yellow fluorescent protein expression in Ibsp-topaz mice to ex vivo purify cementoblasts by fluorescence-activated cell sorting (FACS) and perform transcriptomic analysis; (2) To analyze the mechanism by which BSP functions in bones and teeth by defining the binding site of BSP on collagen and analyzing inactivation of BSP RGD and collagen-binding domains in cementoblasts in vitro and genetically engineered mouse lines in vivo; and (3) To evaluate the role of BSP in alveolar bone repair using a molar socket healing model in mice and an osteotomy healing model in human subjects to map BSP expression, and determine healing outcomes in mice when BSP is ablated or specific functional domains have been inactivated.

项目概要/摘要牙周复合体，包括牙骨质、牙周膜 (PDL) 和牙槽骨，对牙齿至关重要附件和功能。牙周病是地球上最常见的疾病之一，导致牙周病破坏和牙齿脱落，影响健康和生活质量。牙周再生是可能的，但是目前的疗法是不可预测的，很少有真正能够再生的，而且许多疗法缺乏生物学基础。路径再生的作用仍不清楚，部分原因是成牙骨质细胞（产生牙骨质细胞的细胞）的起源和分化牙骨质）以及牙骨质形成的调节过程仍然知之甚少。骨唾液蛋白（Ibsp 基因; BSP 蛋白）是一种与矿化组织相关的多功能细胞外基质（ECM）蛋白，骨骼形成和骨重塑。 Ibsp 基因敲除 (Ibsp-/-) 小鼠缺乏功能性非细胞功能牙骨质、PDL 脱离、牙槽骨缺陷和细胞牙骨质矿化以及牙槽骨吸收和牙齿脱落。尽管 BSP 具有三个功能域，包括胶原蛋白结合域、促进矿化作用，以及启动细胞信号传导的精氨酸-甘氨酸-天冬氨酸 (RGD) 整合素结合域。我们认为 BSP 是研究牙骨质形成和牙槽骨愈合的独特候选因子因为它是选择性表达的，对于正常功能至关重要，并且通过非冗余机制运行与其他生长因子不同。根据我们的初步数据，我们的假设是成牙骨质细胞是表达 BSP 的外间充质细胞与成骨细胞不同； BSP 通过 RGD 结构域向骨细胞发出信号参与骨重塑，并通过胶原蛋白结合域将矿物质沉积到胶原纤维上；和央行促进牙槽骨愈合。这些假设将通过 3 个具体目标进行检验：（1）定义起源和使用来自外间充质细胞和上皮细胞的 Ibsp 的条件消融来分析成牙骨质细胞的转录组群体，并使用 Ibsp-topaz 小鼠中的内源黄色荧光蛋白表达进行离体纯化通过荧光激活细胞分选（FACS）对成牙骨质细胞进行分选并进行转录组分析； (2) 分析通过定义 BSP 在胶原蛋白上的结合位点，BSP 在骨骼和牙齿中发挥作用的机制体外和遗传学分析成牙骨质细胞中 BSP RGD 和胶原蛋白结合域的失活体内工程小鼠品系； (3) 评估 BSP 在使用磨牙槽修复牙槽骨中的作用小鼠愈合模型和人类受试者截骨愈合模型，以绘制 BSP 表达图谱，以及确定 BSP 被消除或特定功能域失活时小鼠的愈合结果。