Yang Z-H, Zhang X-J, Dang N-N, Ma Z-F, Xu L, Wu J-J, Sun Y-J, Duan Y-Z, Lin Z, Jin Y
Department of Orthodontics, School of Stomatology, Fourth Millitary Medical University, 145 West Changle Road, Xi'an, Shaanxi, China.
J Periodontal Res. 2009 Apr;44(2):199-210. doi: 10.1111/j.1600-0765.2008.01106.x. Epub 2008 Jun 25.
Limitations of current periodontal regeneration modalities in both predictability and extent of healing response, especially on new cementum and attachment formation, underscore the importance of restoring or providing a microenvironment that is capable of promoting the differentiation of periodontal ligament stem cells (PDLSCs) towards cementoblast-like cells and the formation of cementum/periodontal ligament-like tissues. The aim of this study was to investigate the biological effect of conditioned medium from developing apical tooth germ cells (APTG-CM) on the differentiation and cementogenesis of PDLSCs both in vitro and in vivo.
Using the limiting dilution technique, single-colony-derived human PDLSCs were isolated and expanded to obtain homogeneous populations of PDLSCs. Morphological appearance, cell cycle analysis, bromodeoxyuridine incorporation, alkaline phosphatase (ALP) activity, mineralization behavior, gene expression of cementoblast phenotype and in vivo differentiation capacities of PDLSCs co-cultured with APTG-CM were evaluated.
The induced PDLSCs exhibited several characteristics of cementoblast lineages, as indicated by the morphological changes, increased proliferation, high ALP activity, and the expression of cementum-related genes and calcified nodule formation in vitro. When transplanted into immunocompromised mice, the induced PDLSCs showed tissue-regenerative capacity to produce cementum/periodontal ligament-like structures, characterized by a layer of cementum-like mineralized tissues and associated periodontal ligament-like collagen fibers connecting with the newly formed cementum-like deposits, whereas control, untreated PDLSCs transplants mainly formed connective tissues.
Our findings suggest that APTG-CM is able to provide a cementogenic microenvironment and induce differentiation of PDLSCs along the cementoblastic lineage. This has important implications for periodontal engineering.
目前牙周再生方式在愈合反应的可预测性和程度方面存在局限性，尤其是在新牙骨质和附着形成方面，这凸显了恢复或提供一种能够促进牙周膜干细胞（PDLSCs）向成牙骨质细胞样细胞分化并形成牙骨质/牙周膜样组织的微环境的重要性。本研究的目的是在体外和体内研究发育中的根尖牙胚细胞条件培养基（APTG-CM）对PDLSCs分化和成牙骨质作用的生物学效应。
采用有限稀释技术分离并扩增单克隆来源的人PDLSCs，以获得均一的PDLSCs群体。评估了与APTG-CM共培养的PDLSCs的形态外观、细胞周期分析、溴脱氧尿苷掺入、碱性磷酸酶（ALP）活性、矿化行为、成牙骨质细胞表型的基因表达以及体内分化能力。
诱导的PDLSCs表现出成牙骨质细胞谱系的几个特征，体外表现为形态变化、增殖增加、高ALP活性、牙骨质相关基因表达以及钙化结节形成。当移植到免疫缺陷小鼠体内时，诱导的PDLSCs显示出产生牙骨质/牙周膜样结构的组织再生能力，其特征是一层牙骨质样矿化组织以及与新形成的牙骨质样沉积物相连的相关牙周膜样胶原纤维，而对照的未处理PDLSCs移植主要形成结缔组织。
我们的研究结果表明，APTG-CM能够提供一个促成牙骨质的微环境，并诱导PDLSCs沿成牙骨质细胞谱系分化。这对牙周组织工程具有重要意义。
