Craniofacial skeletal cell lineage plasticity for reconstituting stem cells and their niches

颅面骨骼细胞谱系可塑性用于重建干细胞及其生态位

• 批准号: 10565884
• 负责人: Noriaki Ono
• 金额: $ 43.58万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-03-01 至 2026-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10565884
• 关键词: ATAC-seq; Adult; Alleles; Alveolar Bone Loss; Bar Codes; CRISPR screen; Cartilage; Cell Differentiation process; Cell Line; Cell Lineage; Cell Therapy; Cells; Cementoblast; Chondrocytes; Dedications; Defect; Deformity; Dental; Dental Sac; Development; Epiphysial cartilage; Event; Fibroblasts; Functional Regeneration; Future; Genes; Growth; In Vitro; Life; Ligature; Mastication; Mesenchymal Stem Cells; Modeling; Molecular; Natural regeneration; Periodontal Diseases; Periodontal Ligament; Periodontitis; Periodontium; Phase; Physiologic Ossification; Play; Recovery; Regenerative capacity; Reporting; Resolution; Rest; Role; Skeletal Development; Slide; Source; Surface; Surgical sutures; Technology; Testing; Time; Tissues; Tooth root structure; Tooth structure; bone; cell motility; comparative; craniofacial; developmental plasticity; epigenomics; experimental study; in vivo; innovation; insight; mineralization; overexpression; parathyroid hormone-related protein; postnatal; reconstitution; skeletal; skeletal tissue; skull base; stem cell niche; stem cells; transcriptome sequencing; transcriptomics; vector

PROJECT SUMMARY/ABSTRACT The craniofacial skeletal tissues are composed of multiple functional units, encompassing both mineralized and non-mineralized components. The non-mineralized tissues, such as sutures, cranial base synchondroses and periodontal ligaments, exist between mineralized tissues, and play important roles in craniofacial growth and regeneration by providing a niche for tissue-specific stem cells in postnatal life. Current cell-based therapies cannot effectively reconstitute stem cell niches; as a result, recovery of devastating skeletal conditions such as craniofacial deformities and advanced alveolar bone loss associated with periodontal diseases has not been made possible to date. Functional regeneration of craniofacial skeletal tissues requires an innovative approach to reestablish inherent stem cells and their supporting niches. In this proposal, we aim to define molecular and cellular mechanisms underlying developmental plasticity of the craniofacial skeletal lineage and explore the possibility to apply these mechanisms to enhance endogenous regeneration capacity. We hypothesize that functionally dedicated cells of the postnatal craniofacial skeletal cell lineage can reconstitute tissue-specific stem cells and their supporting niches through lineage plasticity. We will test this hypothesis using a combination of in vivo clonal lineage-tracing and single-cell and spatial transcriptomic approaches to unravel fundamental molecular and cellular events associated with formation of stem cells and their stem cell niche. We will focus on two models of the cranial base synchondrosis and the periodontium to investigate developmental craniofacial skeletal lineage plasticity. In Aim 1, we will characterize plasticity of Runx2+ perichondrial cells in establishing the cranial base synchondrosis niche. We hypothesize Runx2+ perichondrial fibroblasts generate both stem cells and their niches within postnatal synchondroses through developmental plasticity. We will use a combination of cell-lineage tracing experiments and single-cell transcriptomic analyses, high-resolution spatial transcriptomic analysis and CRISPR screens using the feature barcoding technology to define molecular mechanisms underlying developmental plasticity and stem cell-generating potential of Runx2+ perichondrial cells of the postnatal synchondrosis. In Aim 2, we will explore the possibility to reactivate PTHrP+ cementoblasts to regenerate functional periodontal attachment apparatus. We hypothesize that PTHrP+ cementoblasts on the adult root surface retain a dental follicle (DF) cell-like state, and can be experimentally reverted to dental root mesenchymal progenitor cells. We will use a combination of cell-lineage tracing experiments, single-cell and bulk transcriptomic and epigenomic analyses to define how PTHrP+ cementoblasts are related to PTHrP+ DF cells, and change their molecular identities during periodontal destruction and regeneration. We will also examine whether PTHrP overexpression is sufficient to revert mature skeletal cells to a mesenchymal progenitor-like state at a post-growth phase, as a proof-of-principle study to test the applicability of developmental lineage plasticity to adult stages.

项目概要/摘要 颅面骨骼组织由多个功能单元组成，包括矿化和 非矿化成分。非矿化组织，如缝线、颅底软骨组织和 牙周膜存在于矿化组织之间，在颅面生长和发育过程中发挥着重要作用。 通过为出生后的组织特异性干细胞提供一个利基来实现再生。当前基于细胞的疗法 不能有效地重建干细胞生态位；结果，破坏性骨骼状况的恢复，例如 与牙周疾病相关的颅面畸形和晚期牙槽骨丢失尚未得到证实 迄今为止已成为可能。颅面骨骼组织的功能再生需要创新方法 重建固有的干细胞及其支持生态位。在本提案中，我们的目标是定义分子和 颅面骨骼谱系发育可塑性的细胞机制并探索 应用这些机制来增强内源性再生能力的可能性。我们假设 出生后颅面骨骼细胞谱系的功能专用细胞可以重建组织特异性 干细胞及其通过谱系可塑性的支持生态位。我们将使用 结合体内克隆谱系追踪与单细胞和空间转录组学方法来解开谜团 与干细胞及其干细胞生态位形成相关的基本分子和细胞事件。 我们将重点研究颅底软骨联合和牙周组织的两种模型 发育性颅面骨骼谱系可塑性。在目标 1 中，我们将表征 Runx2+ 的可塑性 软骨膜细胞建立颅底软骨联合生态位。我们假设 Runx2+ 软骨膜 成纤维细胞通过发育过程产生干细胞及其在出生后软骨细胞内的生态位 可塑性。我们将结合使用细胞谱系追踪实验和单细胞转录组分析， 使用特征条形码技术进行高分辨率空间转录组分析和 CRISPR 筛选 定义 Runx2+ 发育可塑性和干细胞生成潜力的分子机制 产后软骨病的软骨膜细胞。在目标 2 中，我们将探索重新激活 PTHrP+ 的可能性 成牙骨质细胞再生功能性牙周附着装置。我们假设 PTHrP+ 成体牙根表面的成牙骨质细胞保留牙囊（DF）细胞样状态，并且可以通过实验 恢复为牙根间充质祖细胞。我们将结合使用细胞谱系追踪 实验、单细胞和批量转录组和表观基因组分析来定义 PTHrP+ 成牙骨质细胞与 PTHrP+ DF 细胞相关，并在牙周期间改变其分子特性 破坏和再生。我们还将检查 PTHrP 过度表达是否足以恢复 成熟的骨骼细胞在生长后阶段达到间充质祖细胞样状态，作为原理验证 研究测试发育谱系可塑性对成年阶段的适用性。