Stem Cell Based Therapy for Regenerative Endodontics

基于干细胞的再生牙髓治疗

• 批准号: 7915543
• 负责人: GEORGE T.J HUANG
• 金额: $ 37.17万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7915543
• 关键词: Stem; Cell; Based; Therapy; Regenerative

DESCRIPTION (provided by applicant): The long-term goal of this proposal is to regenerate and repair dental pulp/dentin thereby reducing the need of endodontic procedures. Approximately 16 million teeth receive root canal treatment (~$14 billion) per year in the US-a procedure consisting of the removal of the entire pulp and replacing it with gutta percha (rubber-like material). There have been no other effective ways to repair the infected or injured pulp tissue besides complete amputation. Recent advances in pulp stem cell isolation and tissue engineering technologies have shed light on the possibility of pulp tissue regeneration. Removing the entire pulp poses two clinical problems: 1) further dental work such as crown and post that weaken the tooth are most often required; 2) devitalized immature teeth are weak with little dentin structure. Therefore the therapeutic capacity to regenerate pulp/dentin of the root would be clinically beneficial. Our preliminary data showed that human pulp-like structure constructed in vitro using stem cells and implanted in canals of human tooth fragments can be vascularized in the subcutaneous space of SCID mice. Therefore, we hypothesize that pulp-like tissue can be regenerated in tooth using stem cells aided with a scaffold system. In this proposal, we will characterize dental stem cells for regeneration of pulp/dentin and use mice and minipigs as ectopic and orthotopic animal study models respectively. There are three specific aims that stepwisely test the hypothesis to reach the goal: Aim 1- To characterize pulp stem cells of various sources for pulp regeneration. This aim will characterize prospective stem cells of various sources for pulp tissue regeneration. Human dental pulp stem cells (hDPSCs), human stem cells from apical papilla (hSCAP) and stem cells from human exfoliated deciduous teeth (SHED) will be isolated and characterized by examining their stem cell properties and gene expression profiles relevant to osteo/odontogenic potency using in vitro and in vivo experimentation. Procedures include flow cytometry, real-time RT-PCR, Western blot, immunohistochemical analysis, etc. Aim 2- To determine angiogenesis of human pulp-like tissues in vivo in SCID mice. The extent of angiogenesis is critical to the success of pulp regeneration. Stem cells seeded in D,L-lactide and glycolide (PLG) scaffolds containing angiogenic growth factors will be inserted into root canal space of human tooth fragments and implanted subcutaneously into SCID mice. The implanted teeth will then be harvested for histological analysis of cellularity, matrix production, formation of odontoblast-like cells and particularly vascularity. Gene expression profile of the regenerated tissue will be analyzed with and immunohistochemistry. Aim 3- To test pulp regeneration in a relevant tissue environment in swine. This experimental swine model simulates actual clinical applications in humans using autologous stem cells for pulp regeneration in pulp space that has been infected and then disinfected. Pulp stem cells from swine teeth will be seeded in scaffolds in vitro and planted back to the disinfected pulpless root canal space for de novo pulp regeneration or planted into pulp chamber space for partial pulp regeneration. The vascularization and innervations of the regenerated tissue will be determined with histology and immunohistochemistry. The success of this project would represent a potential breakthrough in clinical endodontics using stem cell therapy and serve as an example for other organ regeneration. PUBLIC HEALTH RELEVANCE: About 16 million teeth receive root canal treatment (~$14 billion) per year in the United States- a procedure that requires the removal of the entire dental pulp tissue which is then replaced with gutta percha (rubber-like material). This pulp tissue is important for the entire function and the health of the tooth, removing which renders the tooth weakened and susceptible to functional failures. There have been no other effective ways to repair the infected or injured pulp/dentin tissue besides complete amputation. The purpose of this proposal is to utilize and further characterize the adult dental stem cells for the regeneration of pulp/dentin tissues. Using animal study models, a method will be established and tested in this proposal to regenerate dental pulps using adult dental stem cells. If successful, a subsequent human trial may be carried out in the future and its success will promise a reduced need of traditional aggressive root canal therapies.

描述（由申请人提供）：该提案的长期目标是再生和修复牙髓/牙本质，从而减少了牙髓方法的需求。在US-A程序中，大约有1600万颗牙齿接受根管治疗（约140亿美元），包括去除整个纸浆并用Gutta Percha（橡胶样材料）代替。除完全截肢外，没有其他有效的方法来修复受感染或受伤的果肉组织。纸浆干细胞分离和组织工程技术的最新进展揭示了纸浆组织再生的可能性。去除整个纸浆会带来两个临床问题：1）最需要削弱牙齿的牙冠和邮政等进一步的牙科工作； 2）失去活力的未成熟牙齿较弱，牙本质结构很少。因此，在临床上，再生纸浆/牙本质的再生能力在临床上是有益的。我们的初步数据表明，使用干细胞在体外构建并植入人牙碎片运河的人类纸浆样结构可以在SCID小鼠的皮下空间中进行血管化。因此，我们假设可以使用脚手架系统帮助的干细胞在牙齿中再生纸浆样组织。在此提案中，我们将表征牙髓/牙本质再生的牙科干细胞，并分别将小鼠和小鼠作为异位和原位动物研究模型。有三个特定的目的是对假设实现目标的三个特定目的：目标1-以表征各种来源的纸浆干细胞以进行纸浆再生。该目的将表征各种来源的前瞻性干细胞用于纸浆组织再生。人牙浆干细胞（HDPSC），来自顶端乳头（HSCAP）的人干细胞和来自人类去角质的落叶牙齿（SHED）的干细胞，通过使用体外和体外和In Vivo实验来检查与骨/氧化的效力相关的干细胞特性和基因表达图谱，以检查和表征其干细胞特性和基因表达谱。程序包括流式细胞仪，实时RT-PCR，Western印迹，免疫组织化学分析等。目标2-确定SCID小鼠体内人类浆状组织的血管生成。血管生成的程度对于纸浆再生的成功至关重要。在含有血管生长因子的D，L-甲状腺素和糖醇（PLG）支架中的干细胞将插入人牙齿碎片的根管空间中，并皮下植入SCID小鼠中。然后，将收集植入的牙齿，以进行细胞性，基质产生，形成Odontoblast样细胞，尤其是血管性的组织学分析。将分析再生组织的基因表达谱和免疫组织化学。目标3-在猪中相关组织环境中测试果肉再生。该实验性猪模型使用自体干细胞在人类中模拟了实际的临床应用，以在已感染然后被消毒的纸浆空间中的纸浆再生中再生。猪牙齿中的纸浆干细胞将在体外播种在脚手架中，并将其播种回消毒的果皮根管空间，以进行从头开始再生或种植到纸浆腔室空间中以进行部分纸浆再生。再生组织的血管化和神经的神经将由组织学和免疫组织化学确定。该项目的成功将代表使用干细胞疗法的临床牙齿牙髓学的潜在突破，并作为其他器官再生的例子。 公共卫生相关性：在美国，约有1600万牙齿每年接受根管治疗（约140亿美元） - 这项程序需要去除整个牙浆组织，然后用Gutta Percha（橡胶样材料）代替。该纸浆组织对于整个功能和牙齿的健康很重要，从而使牙齿削弱并容易受到功能失败的影响。 除完全截肢外，没有其他有效的方法来修复感染或受伤的纸浆/牙本质组织。该提案的目的是利用并进一步表征成年牙科干细胞的果肉/牙本质组织的再生。使用动物研究模型，将在此提案中建立和测试一种使用成年牙科干细胞再生牙髓的方法。如果成功，将来可能会进行随后的人类试验，其成功将有望减少对传统侵略性根管疗法的需求。