Scaffold-free Tissue Engineering: Using Principles from Developmental Biology to Support Craniofacial Regeneration

 描述（由申请人提供）：该职业发展奖将促进主要研究者（PI）过渡到终身教职所需的培训，并建立一个独立的研究计划，专注于使用发育生物学原理来生成再生颅面治疗的仿生结构。这项研究的最终目标是设计定制的多组织结构来替代牙齿组织。 组织工程结构可以恢复功能和美观的患者遭受创伤，先天性缺陷，疾病的颅面区域。然而，颅面区域包括复杂的多组织结构，由于难以引导正确的空间组织和产生适当的界面，使用传统的组织工程方法重建这些结构具有挑战性。在自然组织发育过程中，细胞迁移和形态梯度的表达是组织图案化的两个关键组成部分;理解和控制这些过程是改善组织工程设计的关键。PI将接受培训，以研究和操纵细胞迁移和形态发生梯度的表达，以研究促进工程组织中有组织的多组织结构形成的机制，并定制组织图案。PI先前已经表明，从牙髓细胞（DPC）形成新的三维无支架组织工程化构建体（SFC）导致空间组织化的牙本质-牙髓复合体样结构，其包含封闭在牙本质样外围内的牙髓样核心。在目标1中，PI将接受先进成像技术的培训，以跟踪3D组织内的细胞迁移。在从牙髓细胞的总群体产生的SFC中，CD 146+干/祖细胞迁移到外周以产生外牙本质样组织的假设将被测试。在目标2中，PI将获得微流控设备的培训，以产生跨SFC的分子梯度，从而改变组织模式。钙离子和转化生长因子β 1（参与牙源性分化的已知因子）的替代物将在整个SFC中产生，以检验牙本质样结构将在最高浓度的化学梯度中形成并过渡到牙髓样组织的假设。作为独立研究者，PI将使用目标1中接受的培训来跟踪SFC中干/祖细胞迁移，以响应微流体装置产生的分子梯度。在目标3中，作为独立研究者，PI将在牙本质-牙髓复合体样SFC的外表面上生成牙骨质样组织层。DPC生成的SFC将用牙周韧带干/祖细胞（PDLC）片包裹，以检验PDLC在空间定位于牙本质-牙髓复合体样结构的外表面时将向成牙骨质细胞表型分化的假设。此外，利用在 目的2，微流控装置将用于产生梯度的骨形态发生蛋白2或胰岛素样生长因子1，已知的成牙骨质细胞分化的诱导剂，通过SFC改变牙骨质图案。

 DESCRIPTION (provided by applicant): This career development award will facilitate the training needed for the principle investigator (PI) to transition to a tenure-track faculty positio and establish an independent research program focused on using developmental biology principles to generate biomimetic constructs for regenerative craniofacial therapy. The ultimate goal of this research is to engineer customized, multi-tissue structures to replace dental tissues. Tissue-engineered constructs could restore function and aesthetics to patients suffering from trauma, congenital defects, and disease to the craniofacial region. However, the craniofacial region comprises complex, multi-tissue structures that are challenging to recreate using traditional tissue engineering methods due to difficulties in directing correct spatial organizatio and producing proper interfaces. During natural tissue development, cell migration and the expression of morphogen gradients are two critical components of tissue patterning; understanding and controlling these processes is key for improving tissue engineering design. The PI will acquire training to study and manipulate cell migration and the expression of morphogen gradients to investigate mechanisms facilitating the formation of organized multi-tissue structures in engineered tissues and to customize tissue patterning. The PI has previously shown that forming novel three-dimensional, scaffold-free tissue-engineered constructs (SFC) from dental pulp cells (DPC) results in spatially organized dentin-pulp complex-like structures containing a pulp-like core enclosed within a dentin-like periphery. In Aim 1, the PI will receive training in advanced imaging techniques to track cell migration within 3D tissues. The hypothesis that in SFCs generated from the total population of dental pulp cells, the CD146+ stem/progenitor cells migrate to the periphery to produce the outer dentin-like tissue will be tested. In Aim 2, the PI will acquire training in microfluidic devices to generate molecular gradients across SFCs to alter tissue patterning. Gradients of calcium and transforming growth factor 1, known factors involved in odontogenic differentiation, will be generated across the SFCs to test the hypothesis that a dentin-like structure will form in in the highest concentrations of the chemical gradients and transition to a pulp-like tissue. As an independent investigator the PI will use the training received in Aim 1 to track stem/progenitor cell migration in SFCs in response to the molecular gradients generated by the microfluidic devices. In Aim 3, as an independent investigator, the PI will generate a cementum-like tissue layer on the outer surface of the dentin-pulp complex-like SFC. SFCs generated by DPCs will be wrapped with a periodontal ligament stem/progenitor cell (PDLC) sheet to test the hypothesis that PDLC will differentiate towards a cementoblast phenotype when spatially positioned on an outer surface of a dentin-pulp complex-like structure. Additionally, using the training received in Aim 2, microfluidic devices will be used to generate gradients of bone morphogenic protein 2 or insulin-like growth factor 1, known inducers of cementoblast differentiation, across the SFCs to alter cementum patterning.