CSF-1 in Dental Biology

CSF-1 在牙科生物学中的应用

基本信息

批准号：
8294405
负责人：
SHERRY L ABBOUD-WERNER
金额：
$ 27.58万
依托单位：
UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-03-05 至 2014-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8294405
关键词：
Address Ameloblasts Amelogenesis Amelogenesis Imperfecta Animal Model Biology Cell Lineage Cell surface Cells Complementary DNA Data Defect Dental Dental Enamel Dentin Development Electrophoretic Mobility Shift Assay Epithelial Cells Gene Delivery Gene Expression Genes Genetic Genetic Crosses Genetic Transcription Goals Human In Situ Hybridization In Vitro Incisor Knock-out LacZ Genes Lentivirus Vector Link Macrophage Colony-Stimulating Factor Mandible Mediating Molecular Morphology Mus Newborn Infant Odontoblasts Oral health Osteocalcin Osteoclasts Phenotype Plant Roots Printing Protein Isoforms Regulatory Element Reporter Genes Resolution Scanning Series Testing Therapeutic Time Tooth Diseases Tooth eruption Tooth structure Transfection Transgenic Mice Transgenic Organisms Work base cis acting element enamel matrix proteins enamelin foot gene therapy improved in vivo kallikrein 4 lentiviral-mediated mRNA Expression mineralization mouse model novel novel therapeutics postnatal prevent promoter protein expression public health relevance research study selective expression transgene expression vector

项目摘要

DESCRIPTION (provided by applicant): Macrophage colony stimulating factor (CSF-1) is essential for tooth matrix formation and eruption. Ameloblasts and odontoblasts express soluble (s) and cell-surface (cs) forms of CSF-1; however, the precise biologic effects of these isoforms on amelogenesis and the regulatory elements in the CSF-1 gene that control their expression during tooth development have not been explored. The long-term goal of this proposal is to characterize the molecular mechanisms that control CSF- 1 expression in ameloblast lineage cells and determine the biologic effect of CSF-1 isoforms on enamel matrix formation using animal models. Our central working hypothesis is that CSF-1 is critical for amelogenesis during tooth development. Preliminary data show, for the first time, that a -774/+183 bp fragment of the CSF-1 promoter in transgenic mice confers high lacZ expression in the inner enamel epithelial (IEE) cells that differentiate into ameloblasts. Our first hypothesis is that cell-specific cis-acting elements in the -774 bp CSF-1 promoter direct gene expression in ameloblast lineage cells during tooth development. To address this issue, a series of -774 bp ]5'CSF-1 promoter deletion constructs will be tested for transcriptional activity in cultured ameloblast and non-ameloblast cells and relevant sequences will be analyzed in vivo by generating transgenic mice harboring these sequences linked to the lacZ reporter gene. In recent studies using op/op mice that lack both CSF-1 isoforms, we showed that absence of CSF-1 alters tooth matrix protein expression that, in turn, leads to enamel and dentin defects. Transgenic op/op mice expressing either csCSF-1 (op/opCS) or sCSF-1 (op/opS) in odontoblasts under the control of the osteocalcin (OC) promoter were generated and showed distinct tooth phenotypes. sCSF-1 corrected dentin and led to partial correction of enamel defects with op/opS mice showing unique features characterized by chalky white teeth and impaired root formation. These findings are novel and indicate that absence of CSF-1 in ameloblasts of op/opS teeth alters enamel matrix and root development. This is supported by our preliminary data in op/opS mice showing decreased enamelin and kallikrein-4 (KLK4, known as EMSP1) as well as shortened roots. Our second hypothesis is that CSF-1 isoforms differentially regulate enamel matrix and root formation and result in distinct phenotypes. For these experiments, the -774/+183 bp CSF-1 promoter will be used to selectively express sCSF-1 or csCSF-1 in ameloblasts. Double transgenic op/op mice carrying sCSF-1 under the OC promoter and harboring either sCSF-1 or csCSF-1 under the -774/+183 bp promoter will be established. Mice will be examined for resolution of enamel defects and teeth will be analyzed for morphology, enamel matrix protein expression, enamel integrity and mineralization. We will also test the hypothesis that lentiviral-mediated gene delivery of sCSF-1 to ameloblasts will rescue enamel/root defects in op/opS mice. Results from these studies should increase our understanding of the molecular mechanisms that regulate CSF-1 and identify distinct functional effects of sCSF-1 and csCSF-1 that may have therapeutic application for preventing enamel defects in acquired and genetic dental disorders such as amelogenesis imperfecta. PUBLIC HEALTH RELEVANCE: Macrophage colony stimulating factor (CSF-1) is a key regulatory molecule for tooth matrix formation and eruption. Work in this proposal plans to determine the biologic effect of soluble and cell surface forms of CSF-1 on enamel matrix formation and the molecular mechanisms that control CSF-1 expression during tooth development using animal models and gene therapy approaches. Results from these studies may suggest novel therapeutic strategies for enhancing enamel integrity and improving oral health in acquired and genetic dental disorders.

描述（由申请人提供）：巨噬细胞刺激因子（CSF-1）对于牙齿基质形成和喷发至关重要。 CSF-1的成成木细胞和Odontoblasts表达可溶性（S）和细胞表面（CS）形式；然而，尚未探索这些同工型对关节化的颗粒发生和控制其在牙齿发育过程中控制其表达的调节元素的精确生物学作用。该提案的长期目标是表征控制成友细胞谱系细胞中CSF-1表达的分子机制，并确定CSF-1同工型使用动物模型对搪瓷基质形成的生物学作用。我们的中心工作假设是，CSF-1对于牙齿发育过程中的休闲发生至关重要。初步数据首次表明，转基因小鼠中CSF -1启动子的-774/+183 bp片段赋予内胺上皮（IEE）细胞中的高LACZ表达，这些细胞分化为成蛋白细胞。我们的第一个假设是-774 bp CSF-1启动子在牙齿发育过程中的细胞特异性顺式作用元件直接基因表达中的直接基因表达。为了解决这个问题，将测试一系列-774 bp] 5'CSF-1启动子缺失构建体，以在培养的Amelbolast和非木质细胞细胞中的转录活性以及相关序列在体内通过产生与LACZ Reporter Gene相关的转基因小鼠，在体内分析相关序列。在最近使用两种CSF-1同工型的OP/OP小鼠的研究中，我们表明CSF-1的缺乏会改变牙齿基质蛋白的表达，从而导致牙釉质和牙本质缺陷。产生了表达CSCSF-1（OP/OPC）或SCSF-1（OP/OPS）的转基因OP/OP小鼠，在骨钙蛋白（OC）启动子的控制下的odontoblasts中产生并显示出明显的牙齿表型。 SCSF-1校正了牙本质，并通过OP/OPS小鼠对牙釉质缺陷进行了部分校正，其特征是粉笔白牙齿和根形成受损的特征。这些发现是新颖的，表明在OP/OPS牙齿的成成木中缺乏CSF-1会改变搪瓷基质和根发育。我们在OP/OPS小鼠中的初步数据支持了这一点，显示搪瓷蛋白降低和Kallikrein-4（KLK4，称为EMSP1）以及缩短根。我们的第二个假设是CSF-1同工型差异地调节了牙釉质基质和根形成，并导致不同的表型。对于这些实验，-774/+183 bp CSF-1启动子将用于选择性地表达成成布中的SCSF-1或CSCSF-1。将建立在-774/+183 bp启动子下携带SCSF-1或CSCSF-1下携带SCSF-1的双转基因OP/OP小鼠。将检查小鼠的牙釉质缺陷，将分析牙齿的形态学，搪瓷基质蛋白表达，搪瓷完整性和矿化。我们还将检验以下假设：慢病毒介导的SCSF-1递送至木材细胞的基因将挽救OP/OPS小鼠中的搪瓷/根缺陷。这些研究的结果应提高我们对调节CSF-1的分子机制的理解，并确定SCSF-1和CSCSF-1的不同功能效应，这些功能可能具有治疗性应用于预防在获得和遗传牙齿疾病（例如AmeLeation Indrefecta）中的牙釉质缺陷。公共卫生相关性：巨噬细胞刺激因子（CSF-1）是牙齿基质形成和喷发的关键调节分子。该提案中的工作计划确定CSF-1的可溶性和细胞表面形式对牙釉质基质形成的生物学作用以及使用动物模型和基因治疗方法在牙齿发育过程中控制CSF-1表达的分子机制。这些研究的结果可能表明，新的治疗策略可以增强牙釉质完整性并改善所获得和遗传牙齿疾病的口腔健康。