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

点击翻译按钮获取中文摘要

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.