Development of an Integrated 3D Human Osteo-Mucosal Model

摘要：硬/软组织界面的完整性和稳定性是再生过程中的主要挑战 以及构建由大相径庭的不同组织类型组成的结构。目前的做法是聘用 生物相容的天然粘合剂，如纤维蛋白胶。然而，使用这种粘合剂，粘接强度是 生理状态不够强壮，随着时间的推移会放松。此外，由于它们充当了额外的 两个组织之间的层/材料，组织之间健康的细胞通讯将受到干扰 对界面组织发育的不利影响。 在这个项目中，我们提出了一种新的软硬组织粘连方法，解决了这两个问题 缺点。我们将优化和应用我们的口腔粘膜/骨粘连技术，以开发一种工程化的 骨-粘膜复合体是一个包含大量不同组织的结构的关键例子。 我们提出的方法依赖于将粘连官能团原位结合到细胞负载的软支架中 基于一种生物兼容的方法，它很容易附着在表面处理过的硬支架上。更多 具体地说，可光固化的明胶基质包裹细胞将被酶处理以具有 可与聚己内酯骨上的胺基形成共价键的苯醌官能团 脚手架。界面结合将以Michael反应为基础。 我们认为，这种新技术与现有方法相比有三大优点：1)它直接将两种方法结合在一起 软硬组织隔间，不需要使用第三种材料。2)附着力很强 强大，不会随着时间的推移而减弱。3)该配方将导致细胞的深度渗透和 促进了支架内细胞的生长。 为了验证我们的假设，我们将在体外对粘附力进行全面的定量和定性评估。 随着时间的推移，骨-粘膜复合体的形成和生物学特性将被检查 详细地说。用纤维蛋白胶作为粘合剂处理的类似结构将被并行生产以供使用 作为对照样品。此外，我们的骨-粘膜结构的粘附力将与 新鲜屠宰的猪下巴中的天然口腔粘膜/骨界面组织。 建议的粘附法在骨-粘膜复合体中的成功结果将表明 该技术可潜在地应用于(经过相关修改)的界面组织工程 牙周复合体、骨/软骨、骨/韧带、骨/腱和牙本质/牙髓复合体。 已开发的骨-粘膜复合体的应用包括：A)临床移植，如牙槽骨移植 重建和口腔内移植，B)生产临床相关的体外测试系统和替代方案 动物实验模型，用于1)研究生物材料与口腔组织的相互作用，以及2)口腔疾病筛查 以及药物输送系统的评价。

SUMMARY: The integration and stability of the hard/soft tissue interface is a major challenge in regeneration and engineering of constructs composed of widely different tissue types. The current approach is employment of biocompatible natural adhesives, such as fibrin glue. However, using such adhesives, the adhesion strength is not strong enough in physiological condition and will loosen in time. Moreover, since they act as an additional layer/material between two tissues, the healthy cellular communication between tissues will be disturbed with adverse influence on interfacial tissue development. In this project, we propose a new method for adhesion of soft/hard tissues, which addresses both of the above drawbacks. We will optimize and apply our technique in oral mucosa/bone adhesion to develop an engineered osteo-mucosal complex as a key example of a construct containing widely dissimilar tissues. Our proposed approach relies on in situ incorporation of adhesive functional groups into a cell-laden soft scaffold based on a biocompatible method, which become readily adherent to a surface treated hard scaffold. More specifically, the photocurable gelatin-based matrix encapsulating cells will be enzymatically treated to have quinone functional groups which can form covalent bonds with the amine groups on the polycaprolactone bone scaffold. The interfacial adhesion will be based on Michael reaction. We claim that this new technique has three major advantages over current methods: 1) It directly integrates two compartments of soft/hard tissues without the need for employing a third material. 2) The adhesion is very strong and will not be diminished over time. 3) The formulation will lead to in depth penetration of cells and enhanced cell growth inside the scaffolds. To examine our hypotheses, the adhesion strength will be fully evaluated in vitro quantitatively and qualitatively over time after development of the osteo-mucosal complex, and the biological characteristics will be inspected in detail. A similar construct treated by fibrin glue as the adhesive agent will be produced in parallel to be used as the control sample. Moreover, the adhesion strength of our osteo-mucosal construct will be compared with the natural oral mucosa/bone interfacial tissue in a freshly slaughtered pig’s jaw. A successful outcome from the proposed adhesion method in the osteo-mucosal complex will suggest that the technique can potentially be applied (after relevant modifications) for engineering of interfacial tissues in periodontal complex, bone/cartilage, bone/ligament, bone/tendon, and dentin/pulp complexes. The applications of the developed osteo-mucosa complex include A) clinical transplantation such as alveolar reconstruction and intraoral grafting, B) production of a clinically relevant in vitro test system and an alternative to animal test models for 1) studying the interaction of biomaterials and oral tissue, and 2) oral disease screening and evaluation of drug delivery systems.